U.S. patent application number 13/435927 was filed with the patent office on 2012-07-19 for novel human micrornas associated with cancer.
This patent application is currently assigned to Exiqon A/S. Invention is credited to Soren Morgenthaler Echwald, Anders Martin Bogild Jacobsen, Anders St.ae butted.rmose Krogh, Morten Lindow, Thomas Litman, Soren Moller, Sanne Nygaard, Rolf Sokilde.
Application Number | 20120184603 13/435927 |
Document ID | / |
Family ID | 41089527 |
Filed Date | 2012-07-19 |
United States Patent
Application |
20120184603 |
Kind Code |
A1 |
Litman; Thomas ; et
al. |
July 19, 2012 |
NOVEL HUMAN MICRORNAS ASSOCIATED WITH CANCER
Abstract
The invention provides new sequences for human microRNAs
associated with cancer which may be used as molecular markers for
cancer diagnostics or as therapeutic targets or agents.
Inventors: |
Litman; Thomas; (Vaerlose,
DK) ; Moller; Soren; (Holte, DK) ; Echwald;
Soren Morgenthaler; (Humlebaek, DK) ; Lindow;
Morten; (Kobenhavn V, DK) ; Jacobsen; Anders Martin
Bogild; (Kobenhavn V, DK) ; Krogh; Anders St.ae
butted.rmose; (Kobenhavn O, DK) ; Nygaard; Sanne;
(Bronshoj, DK) ; Sokilde; Rolf; (Kobenhavn N,
DK) |
Assignee: |
Exiqon A/S
Vedbaek
DK
|
Family ID: |
41089527 |
Appl. No.: |
13/435927 |
Filed: |
March 30, 2012 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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11975644 |
Oct 19, 2007 |
8188255 |
|
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13435927 |
|
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|
60900081 |
Feb 7, 2007 |
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60853410 |
Oct 20, 2006 |
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Current U.S.
Class: |
514/44R ; 506/16;
536/23.1; 536/24.3 |
Current CPC
Class: |
C12Q 1/6886 20130101;
C07H 21/02 20130101 |
Class at
Publication: |
514/44.R ;
536/23.1; 536/24.3; 506/16 |
International
Class: |
C07H 21/02 20060101
C07H021/02; C40B 40/06 20060101 C40B040/06; A61K 31/7088 20060101
A61K031/7088 |
Claims
1. A compound which comprises a contiguous nucleobase sequence of
between a total of 8 to 30 nucleobases, wherein the contiguous
sequence is identical or complementary to a nucleotide sequence
present in a nucleic acid sequence selected from the group
consisting of SEQ ID NOS 538, 539, and 540.
2. The compound according to claim 1, wherein the contiguous
nucleobase sequence is in the form of an oligonucleotide.
3. The compound according to claim 1, wherein the contiguous
nucleotide sequence comprises at least one nucleotide analogue.
4. The compound according to claim 2, wherein the oligonucleotide
comprises a total of less than 21 nucleobases.
5. The compound according to claim 4, wherein the oligonucleotide
comprises a total of between 12 and 20 nucleobases.
6. A conjugate comprising the compound according to claim 1 and at
least one non-nucleobase moiety covalently attached thereto.
7. A composition comprising a compound according to claim 1 or the
conjugate according to claim 6, and at least one pharmaceutically
acceptable diluent, carrier, salt or adjuvant.
8. The compound of claim 1, wherein said compound is a detection
probe.
9. The compound of claim 8, wherein the contiguous nucleobase
sequence forms a recognition sequence which is able to specifically
hybridize or is complementary to, a RNA selected from the group
consisting of SEQ ID NOs: 538, 539, and 540.
10. The compound of claim 8, wherein the oligonucleotide comprises
nucleoside analogues with regular spacing over part or the entire
nucleobases sequence.
11. The compound of claim 10, wherein the regular spacing is a
nucleotide analogue at every second, third or fourth nucleobase
position, or combination thereof.
12. A collection of detection probes which comprises at least one
compound according to claim 8, and at least one further detection
probe.
13. The collection of detection probes according to claim 12, which
comprises at least one detection probe pair, and optionally at
least one further detection probe, wherein one first member of the
detection probe pair specifically hybridizes to, or is
complementary to, a mature miRNA selected from the odd numbered SEQ
IDs No 1-407, or the SEQ IDs listed in the first column of table 3,
and the second member of the detection probe pair hybridizes, or is
complementary to, the corresponding pre-mature miRNA even numbered
SEQ IDs NO 2-408, or the SEQ IDs listed in the second column of
table 3, wherein either the first member of the detection probe
pair is unable to specifically hybridize to, or is not
complementary to, the pre-mature miRNA, and/or the second member of
the detection probe pair is unable to specifically hybridze to, or
is not complementary to, the mature miRNA.
14. The collection of detection probes according to claim 13, which
comprises at least two of said nonidentical detection probe
pairs.
15. The collection of detection probes according to claim 12, which
comprises at least one further detection probe comprising a
recognition sequence consisting of nucleobases, wherein the
detection probe is able to specifically hybridze to at least one
RNA selected from the group comprising: hsa mIR 21, hsa-Let 7i, hsa
miR 101, hsa miR 145, hsa miR 9, hsa miR122a, hsa miR 128b, hsa miR
149, hsa miR 125a, hsa miR 143, hsa miR 136, and hsa-miR 205.
16. The collection of detection probes according to claim 12, which
further comprises at least one detection probe comprising a
recognition sequence consisting of nucleobases, wherein the
detection probe is able to specifically hybridize to at least one
mRNA or DNA sequence associated with cancer.
17. The collection of detection probes according to claim 12 which
comprises at least 5 detection probes.
18. The collection of detection probes according to 17, which
comprises at least 30 detection probes.
19. A kit for the characterization of cancer, the kit comprising at
least one compound according to claim 8, or a collection of
detection probes according to claim 12.
20. A kit according to claim 19, wherein the kit is in the form of,
or comprises, an oligonucleotide array.
21. The compound according to claim 3, wherein said nucleotide
analogue is a LNA.
22. The compound of claim 1, wherein said compound has a total of
12 to 30 nucleobases.
23. The compound of claim 1, wherein said contiguous sequence is
identical to a contiguous sequence present in a nucleic acid
sequence selected from the group consisting of SEQ ID NOs: 538,
539, and 540.
24. The compound of claim 23, wherein said compound comprises the
sequence of SEQ ID NO: 538, 539, or 540.
25. The compound of claim 23, wherein said compound consists of the
sequence of SEQ ID NO: 538, 539, or 540.
26. The compound of claim 1, wherein said contiguous sequence is
complementary to a contiguous sequence present in a nucleic acid
sequence selected from the group consisting of SEQ ID NOs: 538,
539, and 540.
27. The compound of claim 25, wherein said compound comprises a
sequence complementary to SEQ ID NO: 538, 539, or 540.
28. The compound of claim 25, wherein said compound consists of a
sequence complementary to SEQ ID NO: 538, 539, or 540.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is a divisional of U.S. application Ser.
No. 11/975,644, filed Oct. 19, 2007, which claims benefit of U.S.
provisional application Nos. 60/900,801, filed Feb. 7, 2007, and
60/853,410, filed Oct. 20, 2006, each of which is hereby
incorporated by reference.
FIELD OF THE INVENTION
[0002] The present invention provides compounds which comprise
novel nucleobase sequences which correspond to previously unknown
miRNA sequences associated with cancer. The compounds may be used
in therapeutics or diagnostics. The invention provides methods for
treatment of cancer, and methods for the detection and analysis of
non-coding RNAs associated with cancer, such as breast cancer. The
invention furthermore relates to collections of oligonucleotide
probes for detection and analysis of non-coding RNAs associated
with cancer, such as breast cancer.
BACKGROUND OF THE INVENTION
[0003] The present invention relates to the detection and analysis
of target nucleotide sequences associated with cancer. The
invention provides novel microRNAs, oligonucleotide probes which
can detect the novel microRNAs, and methods employing the use of
oligonucleotide probes that are useful for detecting and analysing
target nucleotide sequences associated with cancer.
[0004] MicroRNAs (miRNAs) have rapidly emerged as an important
class of short endogenous RNAs that act as post-transcriptional
regulators of gene expression by base-pairing with their target
mRNAs. The 19-25 nucleotide (nt) mature miRNAs are processed
sequentially from longer hairpin transcripts by the RNAse III
ribonucleases Drosha (Lee, Y., et al., 2003. Nature 425: 415-419.)
and Dicer (Hutvagner, G., et al., 2001. Science 293: 834-838,
Ketting, R. F., et al., 2001. Genes Dev. 15: 2654-2659.). To date
more than 3400 microRNAs have been annotated in vertebrates,
invertebrates and plants according to the miRBase database release
7.1 in October 2005 (Griffiths-Jones, S. 2004. NAR 32 (Database
issue), D109-D111), and many miRNAs that correspond to putative
genes have also been identified. Some miRNAs have multiple loci in
the genome (Reinhart, B. J., et al., 2002. Genes Dev. 16,
1616-1626.) and occasionally, several miRNA genes are arranged in
tandem clusters (Lagos-Quintana, M., et al., 2001. Science 294:
853-858.). Recent bioinformatic predictions combined with array
analyses, small RNA cloning and Northern blot validation indicate
that the total number of miRNAs in vertebrate genomes is
significantly higher than previously estimated and may be as many
as 1000 (Bentwich, I., et al., 2005. Nat. Genet. 37: 766-770,
Berezikov, E., et al., 2005. Cell 120: 21-24, Xie, X., Lu, J., et
al., 2005. Nature 434: 338-345.).
[0005] In a series of publications during recent years, it has
become clear that microRNAs are extensively involved in cancer
pathogenesis, and microRNA has been shown to be differentially
expressed in a number of cancers (Breast cancer: Iorio et al Cancer
Res 2005; 65: 7065. Lung cancer: Yanaihara et al Cell Science 2006;
9: 189-198. Chronic lymphocytic leukaemia (CLL): Galin et al PNAS,
2004 101(32):11755-11760. Colon cancer: Cummins et al PNAS 2006,
103 (10):3687-3692. Prostate cancer: Volinia et al PNAS 2006; 103:
2257). In fact, in a landmark paper Lu et al (Nature 2005;
435:834-838) demonstrated that differential expression of microRNA
in multiple cancers types, and that signatures based on
approximately 200 microRNAs improve classification of poorly
differentiated cancers over mRNA profiles.
[0006] Furthermore, the expected complexity of the "microRNA'nome"
is far smaller than the human transcriptome with the total number
of microRNAs being approximately limited to between 800 to 1000.
Therefore, a microRNA cancer signature can be predicted to include
from 5-20 microRNAs, suggesting that microRNA based theranostics
will be of limited complexity and far more robust than mRNA
profiles.
[0007] Taken together microRNA constitutes a new class of
non-coding RNAs that plays a significant role in determining gene
expression, microRNAs are differentially expressed in human cancers
and a series of recent publication show that microRNA classify
human cancers; in some cases improvement over mRNA classification
is observed.
[0008] The present invention allows for the determination of
microRNA signatures that improve the classification of early
diagnosed cancers. The microRNA signatures--following from the role
of microRNAs in cancer--reveal the true cancerous potential of the
tumor, and enable physicians to select the appropriate treatment.
microRNA based cancer classification may significantly benefit
patient care, because recurrence rate may be improved due to
adequate treatment of traditionally classified low risk patients,
and suitable therapy, such as adjuvant chemotherapy for breast
cancer may be deselected for the large group of patients that do
not benefit from it.
[0009] PCT/DK2005/000838, and U.S. application Ser. No. 11/324,177,
both hereby incorporated by reference, discloses methods for the
detection of microRNAs (miRNAs) using oligonucleotides which
comprise nucleotide analogues, such as locked nucletic acids
(LNAs).
[0010] WO2005/098029, hereby incorporated by reference, discloses a
method using oligonucleotides for the detection, quantification,
monitoring of expression of siRNA and/or miRNA. It is suggested
that the method can be used for determining the differences between
nucleic acid samples from e.g. a cancer patient.
[0011] The Sanger Institute publishes known miRNA sequences in the
miRBase database. To date there are 533 human siRNAs present in the
miRBase database.
[0012] WO2006/015312 discloses sets of genetic markers which can be
correlated with a prognosis of breast cancer.
[0013] Iorio et al, (Cancer Res 2005; 65 (16), pp 7065-7070)
discloses miRNAs whose expression profile is altered between breast
cancer tumors and non tumor cells.
SUMMARY OF THE INVENTION
[0014] The invention provides a compound which comprises a
contiguous sequence of nucleobases of between 8 and 30 nucleobases
in length, wherein the nucleobase sequence of the contiguous
sequence corresponds to a contiguous nucleic acid or nucleobase
sequence present in a nucleobase sequence selected from the group
consisting of: SEQ ID No 1 and/or 2; SEQ ID No 3 and/or 4; SEQ ID
No 5 and/or 6; SEQ ID No 7 and/or 8; SEQ ID No 9 and/or 10; SEQ ID
No 11 and/or 12; SEQ ID No 13 and/or 14; SEQ ID No 15 and/or 16;
SEQ ID No 17 and/or 18; SEQ ID No 19 and/or 20; SEQ ID No 21 and/or
22; SEQ ID No 23 and/or 24; SEQ ID No 25 and/or 26; SEQ ID No 27
and/or 28; SEQ ID No 29 and/or 30; SEQ ID No 31 and/or 32; SEQ ID
No 33 and/or 34; SEQ ID No 35 and/or 36; SEQ ID No 37 and/or 38;
SEQ ID No 39 and/or 40; SEQ ID No 41 and/or 42; SEQ ID No 43 and/or
44; SEQ ID No 45 and/or 46; SEQ ID No 47 and/or 48; SEQ ID No 49
and/or 50; SEQ ID No 51 and/or 52; SEQ ID No 53 and/or 54; SEQ ID
No 55 and/or 56; SEQ ID No 57 and/or 58; SEQ ID No 59 and/or 60;
SEQ ID No 61 and/or 62; SEQ ID No 63 and/or 64; SEQ ID No 65 and/or
66; SEQ ID No 67 and/or 68; SEQ ID No 69 and/or 70; SEQ ID No 71
and/or 72; SEQ ID No 73 and/or 74; SEQ ID No 75 and/or 76; SEQ ID
No 77 and/or 78; SEQ ID No 79 and/or 80; SEQ ID No 81 and/or 82;
SEQ ID No 83 and/or 84; SEQ ID No 85 and/or 86; SEQ ID No 87 and/or
88; SEQ ID No 89 and/or 90; SEQ ID No 91 and/or 92; SEQ ID No 93
and/or 94; SEQ ID No 95 and/or 96; SEQ ID No 97 and/or 98; SEQ ID
No 99 and/or 100; SEQ ID No 101 and/or 102; SEQ ID No 103 and/or
104; SEQ ID No 105 and/or 106; SEQ ID No 107 and/or 108; SEQ ID No
109 and/or 110; SEQ ID No 111 and/or 112; SEQ ID No 113 and/or 114;
SEQ ID No 115 and/or 116; SEQ ID No 117 and/or 118; SEQ ID No 119
and/or 120; SEQ ID No 121 and/or 122; SEQ ID No 123 and/or 124; SEQ
ID No 125 and/or 126; SEQ ID No 127 and/or 128; SEQ ID No 129
and/or 130; SEQ ID No 131 and/or 132; SEQ ID No 133 and/or 134; SEQ
ID No 135 and/or 136; SEQ ID No 137 and/or 138; SEQ ID No 139
and/or 140; SEQ ID No 141 and/or 142; SEQ ID No 143 and/or 144; SEQ
ID No 145 and/or 146; SEQ ID No 147 and/or 148; SEQ ID No 149
and/or 150; SEQ ID No 151 and/or 152; SEQ ID No 153 and/or 154; SEQ
ID No 155 and/or 156; SEQ ID No 157 and/or 158; SEQ ID No 159
and/or 160; SEQ ID No 161 and/or 162; SEQ ID No 163 and/or 164; SEQ
ID No 165 and/or 166; SEQ ID No 167 and/or 168; SEQ ID No 169
and/or 170; SEQ ID No 171 and/or 172; SEQ ID No 173 and/or 174; SEQ
ID No 175 and/or 176; SEQ ID No 177 and/or 178; SEQ ID No 179
and/or 180; SEQ ID No 181 and/or 182; SEQ ID No 183 and/or 184; SEQ
ID No 185 and/or 186; SEQ ID No 187 and/or 188; SEQ ID No 189
and/or 190; SEQ ID No 191 and/or 192; SEQ ID No 193 and/or 194; SEQ
ID No 195 and/or 196; SEQ ID No 197 and/or 198; SEQ ID No 199
and/or 200; SEQ ID No 201 and/or 202; SEQ ID No 203 and/or 204; SEQ
ID No 205 and/or 206; SEQ ID No 207 and/or 208; SEQ ID No 209
and/or 210; SEQ ID No 211 and/or 212; SEQ ID No 213 and/or 214; SEQ
ID No 215 and/or 216; SEQ ID No 217 and/or 218; SEQ ID No 219
and/or 220; SEQ ID No 221 and/or 222; SEQ ID No 223 and/or 224; SEQ
ID No 225 and/or 226; SEQ ID No 227 and/or 228; SEQ ID No 229
and/or 230; SEQ ID No 231 and/or 232; SEQ ID No 233 and/or 234; SEQ
ID No 235 and/or 236; SEQ ID No 237 and/or 238; SEQ ID No 239
and/or 240; SEQ ID No 241 and/or 242; SEQ ID No 243 and/or 244; SEQ
ID No 245 and/or 246; SEQ ID No 247 and/or 248; SEQ ID No 249
and/or 250; SEQ ID No 251 and/or 252; SEQ ID No 253 and/or 254; SEQ
ID No 255 and/or 256; SEQ ID No 257 and/or 258; SEQ ID No 259
and/or 260; SEQ ID No 261 and/or 262; SEQ ID No 263 and/or 264; SEQ
ID No 265 and/or 266; SEQ ID No 267 and/or 268; SEQ ID No 269
and/or 270; SEQ ID No 271 and/or 272; SEQ ID No 273 and/or 274; SEQ
ID No 275 and/or 276; SEQ ID No 277 and/or 278; SEQ ID No 279
and/or 280; SEQ ID No 281 and/or 282; SEQ ID No 283 and/or 284; SEQ
ID No 285 and/or 286; SEQ ID No 287 and/or 288; SEQ ID No 289
and/or 290; SEQ ID No 291 and/or 292; SEQ ID No 293 and/or 294; SEQ
ID No 295 and/or 296; SEQ ID No 297 and/or 298; SEQ ID No 299
and/or 300; SEQ ID No 301 and/or 302; SEQ ID No 303 and/or 304; SEQ
ID No 305 and/or 306; SEQ ID No 307 and/or 308; SEQ ID No 309
and/or 310; SEQ ID No 311 and/or 312; SEQ ID No 313 and/or 314; SEQ
ID No 315 and/or 316; SEQ ID No 317 and/or 318; SEQ ID No 319
and/or 320; SEQ ID No 321 and/or 322; SEQ ID No 323 and/or 324; SEQ
ID No 325 and/or 326; SEQ ID No 327 and/or 328; SEQ ID No 329
and/or 330; SEQ ID No 331 and/or 332; SEQ ID No 333 and/or 334; SEQ
ID No 335 and/or 336; SEQ ID No 337 and/or 338; SEQ ID No 339
and/or 340; SEQ ID No 341 and/or 342; SEQ ID No 343 and/or 344; SEQ
ID No 345 and/or 346; SEQ ID No 347 and/or 348; SEQ ID No 349
and/or 350; SEQ ID No 351 and/or 352; SEQ ID No 353 and/or 354; SEQ
ID No 355 and/or 356; SEQ ID No 357 and/or 358; SEQ ID No 359
and/or 360; SEQ ID No 361 and/or 362; SEQ ID No 363 and/or 364; SEQ
ID No 365 and/or 366; SEQ ID No 367 and/or 368; SEQ ID No 369
and/or 370; SEQ ID No 371 and/or 372; SEQ ID No 373 and/or 374; SEQ
ID No 375 and/or 376; SEQ ID No 377 and/or 378; SEQ ID No 379
and/or 380; SEQ ID No 381 and/or 382; SEQ ID No 383 and/or 384; SEQ
ID No 385 and/or 386; SEQ ID No 387 and/or 388; SEQ ID No 389
and/or 390; SEQ ID No 391 and/or 392; SEQ ID No 393 and/or 394; SEQ
ID No 395 and/or 396; SEQ ID No 397 and/or 398; SEQ ID No 399
and/or 400; SEQ ID No 401 and/or 402; SEQ ID No 403 and/or 404; SEQ
ID No 405 and/or 406 and SEQ ID No 407 and/or 408, SEQ ID NOs 411
and/or 412; SEQ ID NOs 413; 414 and/or 415; SEQ ID NOs 416 and/or
417; SEQ ID NOs 418; 420; 421 and/or 419; SEQ ID NOs 422 and/or
423; SEQ ID NOS 424 and/or 425; SEQ ID NOS 426; 428 and/or 427; SEQ
ID NOS 429; 430; 431; 432 and/or 433; SEQ ID NOS 434 and/or 435;
SEQ ID NOS 436 and/or 437; SEQ ID NOS 438 and/or 439; SEQ ID NOS
440 and/or 441, SEQ ID NOS 442 and/or 443, SEQ ID NOS 444, 445
and/or 446, SEQ ID NOS 447 and/or 448, SEQ ID NOS 449, 451 and/or
450, SEQ ID NOS 452 and/or 453, SEQ ID NOS 454 and/or 455, SEQ ID
NOS 456 and/or 457, SEQ ID NOS 458 and/or 459, SEQ ID NOS 460
and/or 461, SEQ ID NOS 462 and/or 463, SEQ ID NOS 464 and/or 465,
SEQ ID NOS 466, 468 and/or 467, SEQ ID NOS 469, 471, 472 and/or
470, SEQ ID NOS 473 and/or 474, SEQ ID NOS 475, 477 and/or 476, SEQ
ID NOS 478 and/or 479, SEQ ID NOS 480 and/or 481, SEQ ID NOS 482
and/or 483, SEQ ID NOS 484 and/or 485, SEQ ID NOS 486 and/or 487,
SEQ ID NOS 488 and/or 489, SEQ ID NOS 490, 492, 493 494, and/or
491, SEQ ID NOS 495, 497, 498, 499, 501, 496 and/or 500; SEQ ID NOS
502 and/or 503; SEQ ID NOS 504; 505 and/or 506; SEQ ID NOS 507
and/or 508; SEQ ID NOS 509 and/or 510; SEQ ID NOS 511 and/or 512;
SEQ ID NOS 513 and/or 514; SEQ ID NOS 515, 517; and/or 516; SEQ ID
NOS 518 and/or 519; SEQ ID NOS 520 and/or 521; SEQ ID NOS 522
and/or 523; SEQ ID NOS 524 and/or 525; SEQ ID NOS 526 and/or 527;
SEQ ID NOS 528 and/or 529; SEQ ID NOS 530 and/or 531; SEQ ID NOS
532 and/or 533; SEQ ID NOS 534 and/or 535; SEQ ID NOS 536 and/or
537; SEQ ID NOS 538; 540 and/or 539; SEQ ID NOS 541; 543 and/or
542; SEQ ID NOS 544 and/or 545; SEQ ID NOS 546 and/or 547; SEQ ID
NOS 548; 551; 552; 553; and/or 554; SEQ ID NOS 549; 552; 553 and/or
554; SEQ ID NOS 550; 553 and/or 554; SEQ ID NOS 555 and/or 556; SEQ
ID NOS 557 and/or 558; and allelic variants thereof.
[0015] The above sequences and their naturally occurring allelic
variants are referred to as `target nucleotide(s)` or `target
sequence(s)` herein. Preferred groups of target sequences are
according to the preferred groups or individual nucleobase
sequences referred to herein.
[0016] The invention also provides for new molecular markers for
cancer, and the use of such markers in the methods according to the
invention, and for use in the collection of probes and/or kits
according to the invention.
[0017] The invention also provides detection probes which comprise
a continuous sequence of nucleobases according to the compound of
the invention. In this respect, in the above list of sequences,
from SEQ ID 1 to SEQ ID 408, the list is prepared to highlight
pairs of sequences, the first being an odd number (e.g. SEQ ID NO
1), and the second being an even number (e.g. SEQ ID NO 2)--written
as `SEQ ID NO 1 and/or 2`, in the above list. The odd number
sequence in each pair corresponds to the mature miRNA sequence,
whereas the even number corresponds to the pre-mature pre-miRNA
sequence. This pairing of SEQ IDs is particularly important when
considered the case where the compound of the invention is in the
form of a detection probe, where pairs of detection probes
(detection probe pairs) may be used to determine the level of a
specific miRNA sequence present in the sample, compared to the
level of the precursor form of that miRNA. This may provide useful
diagnostic information, for example in cancer derived samples.
[0018] However, for SEQ ID NOs 411-558 the miRNAs and their
corresponding pre-miRNAs, in addition to related miRNA (and/or
related pre-miRNAs) are shown in Table 3. It will be noted that for
some miRNAs there are more than one possible pre-miRNA precursor,
and in some cases, some pre-miRNAs may result in more than one
mature miRNA. Suitably the detection probe pairs may be selected
from one (or more) miRNA and one (or more) of the corresponding
pre-miRNAs.
DESCRIPTION OF INVENTION
Definitions
[0019] For the purposes of the subsequent detailed description of
the invention the following definitions are provided for specific
terms, which are used in the disclosure of the present
invention:
[0020] In the present context "ligand" means something, which
binds. Ligands may comprise biotin and functional groups such as:
aromatic groups (such as benzene, pyridine, naphtalene, anthracene,
and phenanthrene), heteroaromatic groups (such as thiophene, furan,
tetrahydrofuran, pyridine, dioxane, and pyrimidine), carboxylic
acids, carboxylic acid esters, carboxylic acid halides, carboxylic
acid azides, carboxylic acid hydrazides, sulfonic acids, sulfonic
acid esters, sulfonic acid halides, semicarbazides,
thiosemicarbazides, aldehydes, ketones, primary alcohols, secondary
alcohols, tertiary alcohols, phenols, alkyl halides, thiols,
disulphides, primary amines, secondary amines, tertiary amines,
hydrazines, epoxides, maleimides, C.sub.1-C.sub.20 alkyl groups
optionally interrupted or terminated with one or more heteroatoms
such as oxygen atoms, nitrogen atoms, and/or sulphur atoms,
optionally containing aromatic or mono/polyunsaturated
hydrocarbons, polyoxyethylene such as polyethylene glycol,
oligo/polyamides such as poly-6-alanine, polyglycine, polylysine,
peptides, oligo/polysaccharides, oligo/polyphosphates, toxins,
antibiotics, cell poisons, and steroids, and also "affinity
ligands", i.e. functional groups or biomolecules that have a
specific affinity for sites on particular proteins, antibodies,
poly- and oligosaccharides, and other biomolecules.
[0021] The singular form "a", "an" and "the" include plural
references unless the context clearly dictates otherwise. For
example, the term "a cell" includes a plurality of cells, including
mixtures thereof. The term "a nucleic acid molecule" includes a
plurality of nucleic acid molecules.
[0022] "Transcriptome" refers to the complete collection of
transcriptional units of the genome of any species. In addition to
protein-coding mRNAs, it also represents non-coding RNAs, such as
small nucleolar RNAs, siRNAs, microRNAs and antisense RNAs, which
comprise important structural and regulatory roles in the cell.
[0023] A "multi-probe library" or "library of multi-probes"
comprises a plurality of multi-probes, such that the sum of the
probes in the library are able to recognise a major proportion of a
transcriptome, including the most abundant sequences, such that
about 60%, about 70%, about 80%, about 85%, more preferably about
90%, and still more preferably 95%, of the target nucleic acids in
the transcriptome, are detected by the probes.
[0024] "Sample" refers to a sample of cells, or tissue or fluid
isolated from an organism or organisms, including but not limited
to, for example, skin, plasma, serum, spinal fluid, lymph fluid,
synovial fluid, urine, tears, blood cells, organs, tumors, and also
to samples of in vitro cell culture constituents (including but not
limited to conditioned medium resulting from the growth of cells in
cell culture medium, recombinant cells and cell components).
[0025] The terms "Detection probes" or "detection probe" or
"detection probe sequence" refer to an oligonucleotide or
oligonucleotide analogue, which oligonucleotide or oligonucleotide
analogue comprises a recognition sequence complementary to a
nucleotide target, such as an RNA (or DNA) target sequence. It is
preferable that the detection probe(s) are oligonucleotides,
preferably where said recognition sequence is substituted with
high-affinity nucleotide analogues, e.g. LNA, to increase the
sensitivity and specificity of conventional oligonucleotides, such
as DNA oligonucleotides, for hybridization to short target
sequences, e.g. mature miRNAs, stem-loop precursor miRNAs,
pri-miRNAs, siRNAs or other non-coding RNAs as well as miRNA
binding sites in their cognate mRNA targets, mRNAs, mRNA splice
variants, RNA-edited mRNAs, antisense RNAs and small nucleolar RNAs
(snRNA).
[0026] The terms "miRNA" and "microRNA" refer to about 18-25 nt
non-coding RNAs derived from endogenous genes. They are processed
from longer (ca 75 nt) hairpin-like precursors termed pre-miRNAs.
MicroRNAs assemble in complexes termed miRNPs and recognize their
targets by antisense complementarity. If the microRNAs match 100%
their target, i.e. the complementarity is complete, the target mRNA
is cleaved, and the miRNA acts like a siRNA. If the match is
incomplete, i.e. the complementarity is partial, then the
translation of the target mRNA is blocked.
[0027] The terms "Small interfering RNAs" or "siRNAs" refer to
21-25 nt RNAs derived from processing of linear double-stranded
RNA. siRNAs assemble in complexes termed RISC(RNA-induced silencing
complex) and target homologous RNA sequences for endonucleolytic
cleavage. Synthetic siRNAs also recruit RISCs and are capable of
cleaving homologous RNA sequences
[0028] The term "RNA interference" (RNAi) refers to a phenomenon
where double-stranded RNA homologous to a target mRNA leads to
degradation of the targeted mRNA. More broadly defined as
degradation of target mRNAs by homologous siRNAs.
[0029] The terms "microRNA precursor" or "miRNA precursor" or
"pre-miRNA" or "premature miRNA" refer to polynucleotide sequences
(approximately 70 nucleotides in length) that form hairpin-like
structures having a loop region and a stem region. The stem region
includes a duplex cre-ated by the pairing of opposite ends of the
pre-miRNA polynucleotide sequence. The loop region connects the two
halves of the stem region. The pre-miRNAs are transcribed as mono-
or poly-cistronic, long, primary precursor transcripts (pri-miRNAs)
that are then cleaved into individual pre-miRNAs by a nuclear RNAse
III-like enzyme. Subsequently pre-miRNA hairpins are exported to
the cytoplasm where they are processed by a second RNAse III-like
enzyme into miRNAs. The target nucleic acid may be present in a
premature miRNA sequence.
[0030] The fragments from the opposing arm, called the miRNA* (or
"miRNA-star") sequences (Lau et al, Science (2001) 294:858-862) are
found in libraries of cloned miRNAs but typically at much lower
frequency than are the miRNAs. For example, in an effort that
identified over 3400 clones representing 80 C. elegans miRNAs, only
38 clones representing 14 miRNAs* were found. This approximately
100-fold difference in cloning frequency indicates that the
miRNA:miRNA* duplex is generally short lived compared to the miRNA
single strand (Bartel et al, Cell (2004) 116:281-297). The target
nucleic acid may be present in a miRNA* sequence.
[0031] The "miRNA precursor loop sequence" or "loop sequence of the
miRNA precursor" or "loop region" of an miRNA precursor is the
portion of an miRNA precursor that is not present in the stem
region and that is not retained in the mature miRNA (or its
complement) upon cleavage by a RNAse III-like enzyme.
[0032] The "miRNA precursor stem sequence" or "stem sequence of the
miRNA precursor" or "stem region" of an miRNA precursor is the
portion of an miRNA precursor created by the pairing of opposite
ends of the pre-miRNA polynucleotide sequence, and in-cluding the
portion of the miRNA precursor that will be retained in the "mature
miRNA."
[0033] The term "Recognition sequence" refers to a nucleotide
sequence that is complementary to a region within the target
nucleotide sequence essential for sequence-specific hybridization
between the target nucleotide sequence and the recognition
sequence.
[0034] The terms "corresponding to" and "corresponds to" refer to
the comparison between the nucleobase sequence of the compound of
the invention, and the equivalent nucleotide sequence or the
reverse complement thereof. Nucleobases sequences which "correspond
to" a SEQ ID, therefore have between 8 and 30 contiguous
nucleobases which form a sequence which is found with i) either the
one or more of the SEQ ID(s), or ii) the reverse complement
thereof. Nucleotide analogues are compared directly to their
equivalent or corresponding natural nucleotides. Sequences which
form the reverse complement of a SEQ ID are referred to as the
complement sequence of the SEQ ID. In a preferably embodiment, the
term complementary refers to fully or perfectly complementary.
[0035] The terms "homologues", "variants" and "fragments" in the
context of `homologues, variants and fragments thereof` in relation
to detection probe sequences and specific detection probes, refers
to any sequence which has at least 8 consecutive nucleotide
residues (or nucleotide analogues), such as at least 10 consecutive
residues (or nucleotide analogues), such as at least 14 consecutive
nucleotides (or nucleotide analogues), in common with at least one
of the sequences, allowing for no more than 1 mismatch per 8
nucleotides (or nucleotide analogues), preferably with no more than
1 mismatch or no mismatch.
[0036] The term `natural allelic variants` and the term `allelic
variants` encompasses both variants which although have a slightly
different sequence (such as a homologue, fragment or variant),
originate from the same chromosomal position, or the same position
on an allelic chromosome, as the non-coding RNAs, and precursors
thereof herein listed. The term `natural allelic variants` and the
term `allelic variants` also encompasses mature non-coding RNAs
encompasses, which may be differentially processed by the
processing enzymes, as this may lead to variants of the same
microRNAs having different lengths e.g. shortened by 1 or 2
nucleotides, despite originating from the same allelic chromosome
position.
[0037] The term "label" as used herein refers to any atom or
molecule which can be used to provide a detectable (preferably
quantifiable) signal, and which can be attached to a nucleic acid
or protein. Labels may provide signals detectable by fluorescence,
radioactivity, colorimetric, X-ray diffraction or absorption,
magnetism, enzymatic activity, and the like.
[0038] As used herein, the terms "nucleic acid", "polynucleotide"
and "oligonucleotide" refer to primers, probes, oligomer fragments
to be detected, oligomer controls and unlabeled blocking oligomers
and shall be generic to polydeoxyribonucleotides (containing
2-deoxy-D-ribose), to polyribonucleotides (containing D-ribose), to
any other type of polynucleotide which is an N glycoside of a
purine or pyrimidine base, or modified purine or pyrimidine bases,
and in one embodiment, nucleobases (a collective term used to
describe both nucleotides and nucleotide analogues, such as LNA).
There is no intended distinction in length between the term
"nucleic acid", "polynucleotide" and "oligonucleotide", and these
terms will be used interchangeably. These terms refer only to the
primary structure of the molecule. Thus, these terms include
double- and single-stranded DNA, as well as double- and single
stranded RNA. The oligonucleotide is comprised of a sequence of
approximately at least 3 nucleotides, preferably at least about 6
nucleotides, and more preferably at least about 8-30 nucleotides
corresponding to a region of the designated target nucleotide
sequence. "Corresponding" means identical to or complementary to
the designated sequence. The oligonucleotide is not necessarily
physically derived from any existing or natural sequence but may be
generated in any manner, including chemical synthesis, DNA
replication, reverse transcription or a combination thereof.
[0039] The terms "oligonucleotide" or "nucleic acid" intend a
polynucleotide of genomic DNA or RNA, cDNA, semi synthetic, or
synthetic origin which, by virtue of its origin or manipulation:
(1) is not associated with all or a portion of the polynucleotide
with which it is associated in nature; and/or (2) is linked to a
polynucleotide other than that to which it is linked in nature; and
(3) is not found in nature. Because mononucleotides are reacted to
make oligonucleotides in a manner such that the 5'-phosphate of one
mononucleotide pentose ring is attached to the 3' oxygen of its
neighbour in one direction via a phosphodiester linkage, an end of
an oligonucleotide is referred to as the "5' end" if its 5'
phosphate is not linked to the 3' oxygen of a mononucleotide
pentose ring and as the "3' end" if its 3' oxygen is not linked to
a 5' phosphate of a subsequent mononucleotide pentose ring. As used
herein, a nucleic acid sequence, even if internal to a larger
oligonucleotide, also may be said to have a 5' and 3' ends. When
two different, non-overlapping oligonucleotides anneal to different
regions of the same linear complementary nucleic acid sequence, the
3' end of one oligonucleotide points toward the 5' end of the
other; the former may be called the "upstream" oligonucleotide and
the latter the "downstream" oligonucleotide.
[0040] By the term "SBC nucleobases" is meant "Selective Binding
Complementary" nucleobases, i.e. modified nucleobases that can make
stable hydrogen bonds to their complementary nucleobases, but are
unable to make stable hydrogen bonds to other SBC nucleobases. As
an example, the SBC nucleobase A', can make a stable hydrogen
bonded pair with its complementary unmodified nucleobase, T.
Likewise, the SBC nucleobase T' can make a stable hydrogen bonded
pair with its complementary unmodified nucleobase, A. However, the
SBC nucleobases A' and T' will form an unstable hydrogen bonded
pair as compared to the base pairs A'-T and A-T'. Likewise, a SBC
nucleobase of C is designated C' and can make a stable hydrogen
bonded pair with its complementary unmodified nucleobase G, and a
SBC nucleobase of G is designated G' and can make a stable hydrogen
bonded pair with its complementary unmodified nucleobase C, yet C'
and G' will form an unstable hydrogen bonded pair as compared to
the base pairs C'-G and C-G'. A stable hydrogen bonded pair is
obtained when 2 or more hydrogen bonds are formed e.g. the pair
between A' and T, A and T', C and G', and C' and G. An unstable
hydrogen bonded pair is obtained when 1 or no hydrogen bonds is
formed e.g. the pair between A' and T', and C' and G'. Especially
interesting SBC nucleobases are 2,6-diaminopurine (A', also called
D) together with 2-thio-uracil (U', also called .sup.2SU)
(2-thio-4-oxo-pyrimidine) and 2-thio-thymine (T', also called
.sup.2ST) (2-thio-4-oxo-5-methyl-pyrimidine). FIG. 4 in PCT
Publication No. WO 2004/024314 illustrates that the pairs
A-.sup.2ST and D-T have 2 or more than 2 hydrogen bonds whereas the
D-.sup.2ST pair forms a single (unstable) hydrogen bond. Likewise
the SBC nucleobases pyrrolo-[2,3-d]pyrimidine-2(3H)-one (C', also
called PyrroloPyr) and hypoxanthine (G', also called I)
(6-oxo-purine) are shown in FIG. 4 in PCT Publication No. WO
2004/024314 where the pairs PyrroloPyr-G and C--I have 2 hydrogen
bonds each whereas the PyrroloPyr-I pair forms a single hydrogen
bond.
[0041] "SBC LNA oligomer" refers to a "LNA oligomer" containing at
least one LNA monomer where the nucleobase is a "SBC nucleobase".
By "LNA monomer with an SBC nucleobase" is meant a "SBC LNA
monomer". Generally speaking SBC LNA oligomers include oligomers
that besides the SBC LNA monomer(s) contain other modified or
naturally occurring nucleotides or nucleosides. By "SBC monomer" is
meant a non-LNA monomer with a SBC nucleobase. By "isosequential
oligonucleotide" is meant an oligonucleotide with the same sequence
in a Watson-Crick sense as the corresponding modified
oligonucleotide e.g. the sequences agTtcATg is equal to
agTscD.sup.2SUg where s is equal to the SBC DNA monomer 2-thio-t or
2-thio-u, D is equal to the SBC LNA monomer LNA-D and .sup.2SU is
equal to the SBC LNA monomer LNA .sup.2SU.
[0042] The complement of a nucleic acid sequence as used herein
refers to an oligonucleotide which, when aligned with the nucleic
acid sequence such that the 5' end of one sequence is paired with
the 3' end of the other, is in "antiparallel association." Bases
not commonly found in natural nucleic acids may be included in the
nucleic acids of the present invention include, for example,
inosine and 7-deazaguanine. Complementarity may not be perfect;
stable duplexes may contain mismatched base pairs or unmatched
bases. Those skilled in the art of nucleic acid technology can
determine duplex stability empirically considering a number of
variables including, for example, the length of the
oligonucleotide, percent concentration of cytosine and guanine
bases in the oligonucleotide, ionic strength, and incidence of
mismatched base pairs.
[0043] Stability of a nucleic acid duplex is measured by the
melting temperature, or "T.sub.m". The T.sub.m of a particular
nucleic acid duplex under specified conditions is the temperature
at which half of the duplexes have disassociated.
[0044] The term "nucleobase" covers the naturally occurring
nucleobases adenine (A), guanine (G), cytosine (C), thymine (T) and
uracil (U) as well as non-naturally occurring nucleobases such as
xanthine, diaminopurine, 8-oxo-N.sup.6-methyladenine,
7-deazaxanthine, 7-deazaguanine, N.sup.4,N.sup.4-ethanocytosin,
N.sup.6,N.sup.6-ethano-2,6-diaminopurine, 5-methylcytosine,
5-(C.sup.3--C.sup.6)-alkynyl-cytosine, 5-fluorouracil,
5-bromouracil, pseudoisocytosine,
2-hydroxy-5-methyl-4-triazolopyridin, isocytosine, isoguanine,
inosine and the "non-naturally occurring" nucleobases described in
Benner et al., U.S. Pat. No. 5,432,272 and Susan M. Freier and
Karl-Heinz Altmann, Nucleic Acid Research, 25: 4429-4443, 1997. The
term "nucleobase" thus includes not only the known purine and
pyrimidine heterocycles, but also heterocyclic analogues and
tautomers thereof. Further naturally and non naturally occurring
nucleobases include those disclosed in U.S. Pat. No. 3,687,808; in
chapter 15 by Sanghvi, in Antisense Research and Application, Ed.
S. T. Crooke and B. Lebleu, CRC Press, 1993; in Englisch, et al.,
Angewandte Chemie, International Edition, 30: 613-722, 1991 (see,
especially pages 622 and 623, and in the Concise Encyclopedia of
Polymer Science and Engineering, J. I. Kroschwitz Ed., John Wiley
& Sons, pages 858-859, 1990, Cook, Anti-Cancer Drug Design 6:
585-607, 1991, each of which are hereby incorporated by reference
in their entirety).
[0045] The term "nucleosidic base" or "nucleobase analogue" is
further intended to include heterocyclic compounds that can serve
as like nucleosidic bases including certain "universal bases" that
are not nucleosidic bases in the most classical sense but serve as
nucleosidic bases. Especially mentioned as a universal base is
3-nitropyrrole or a 5-nitroindole. Other preferred compounds
include pyrene and pyridyloxazole derivatives, pyrenyl,
pyrenylmethylglycerol derivatives and the like. Other preferred
universal bases include, pyrrole, diazole or triazole derivatives,
including those universal bases known in the art.
[0046] By "oligonucleotide," "oligomer," or "oligo" is meant a
successive chain of monomers (e.g., glycosides of heterocyclic
bases) connected via internucleoside linkages. The linkage between
two successive monomers in the oligo consist of 2 to 4, desirably
3, groups/atoms selected from --CH.sub.2--, --O--, --S--,
--NR.sup.H--, >C.dbd.O, >C.dbd.NR.sup.H, >C.dbd.S,
--Si(R'').sub.2--, --SO--, --S(O).sub.2--, --P(O).sub.2--,
--PO(BH.sub.3)--, --P(O,S)--, --P(S).sub.2--, --PO(R'')--,
--PO(OCH.sub.3)--, and --PO(NHR.sup.H)--, where R.sup.H is selected
from hydrogen and C.sub.1-4-alkyl, and R'' is selected from
C.sub.1-6-alkyl and phenyl. Illustrative examples of such linkages
are --CH.sub.2--CH.sub.2--CH.sub.2--, --CH.sub.2--CO--CH.sub.2--,
--CH.sub.2--CHOH--CH.sub.2--, --O--CH.sub.2--O--,
--O--CH.sub.2--CH.sub.2--, --O--CH.sub.2--CH.dbd. (including
R.sup.5 when used as a linkage to a succeeding monomer),
--CH.sub.2--CH.sub.2--O--, --NR.sup.H--CH.sub.2--CH.sub.2--,
--CH.sub.2--CH.sub.2--NR.sup.H--, --CH.sub.2--NR.sup.H--CH.sub.2--,
--O--CH.sub.2--CH.sub.2--NR.sup.H--, --NR.sup.H--CO--O--,
--NR.sup.H--CO--NR.sup.H--, --NR.sup.H--CS--NR.sup.H--,
--NR.sup.H--C(.dbd.NR.sup.H)--NR.sup.H--,
--NR.sup.H--CO--CH.sub.2--NR.sup.H--, --O--CO--O--,
--O--CO--CH.sub.2--O--, --O--CH.sub.2--CO--O--,
--CH.sub.2--CO--NR.sup.H--, --O--CO--NR.sup.H--,
--NR.sup.H--CO--CH.sub.2--, --O--CH.sub.2--CO--NR.sup.H--,
--O--CH.sub.2--CH.sub.2--NR.sup.H--, --CH.dbd.N--O--,
--CH.sub.2--NR.sup.H--O--, --CH.sub.2--O--N=(including R.sup.5 when
used as a linkage to a succeeding monomer),
--CH.sub.2--O--NR.sup.H--, --CO--NR.sup.H--CH.sub.2--,
--CH.sub.2--NR.sup.H--O--, --CH.sub.2--NR.sup.H--CO--,
--O--NR.sup.H--CH.sub.2--, --O--NR.sup.H--, --O--CH.sub.2--S--,
--S--CH.sub.2--O--, --CH.sub.2--CH.sub.2--S--,
--O--CH.sub.2--CH.sub.2--S--, --S--CH.sub.2--CH.dbd.(including
R.sup.5 when used as a linkage to a succeeding monomer),
--S--CH.sub.2--CH.sub.2--, --S--CH.sub.2--CH.sub.2--O--,
--S--CH.sub.2--CH.sub.2--S--, --CH.sub.2--S--CH.sub.2--,
--CH.sub.2--SO--CH.sub.2--, --CH.sub.2--SO.sub.2--CH.sub.2--,
--O--SO--O--, --O--S(O).sub.2--O--, --O--S(O).sub.2--CH.sub.2--,
--O--S(O).sub.2--NR.sup.H--, --NR.sup.H--S(O).sub.2--CH.sub.2--,
--O--S(O).sub.2--CH.sub.2--, --O--P(O).sub.2--O--,
--O--P(O,S)--O--, --O--P(S).sub.2--O--, --S--P(O).sub.2--O--,
--S--P(O,S)--O--, --S--P(S).sub.2--O--, --O--P(O).sub.2--S--,
--O--P(O,S)--S--, --O--P(S).sub.2--S--, --S--P(O).sub.2--S--,
--S--P(O,S)--S--, --S--P(S).sub.2--S--, --O--PO(R'')--O--,
--O--PO(OCH.sub.3)--O--, --O--PO(OCH.sub.2CH.sub.3)--O--,
--O--PO(OCH.sub.2CH.sub.2S--R)--O--, --O--PO(BH.sub.3)--O--,
--O--PO(NHR.sup.N)--O--, --O--P(O).sub.2--NR.sup.H--,
--NR.sup.H--P(O).sub.2--O--, --O--P(O,NR.sup.H)--O--,
--CH.sub.2--P(O).sub.2--O--, --O--P(O).sub.2--CH.sub.2--, and
--O--Si(R'').sub.2--O--; among which --CH.sub.2--CO--NR.sup.H--,
--CH.sub.2--NR.sup.H--O--, --S--CH.sub.2--O--,
--O--P(O).sub.2--O--, --O--P(O,S)--O--, --O--P(S).sub.2--O--,
--NR.sup.H--P(O).sub.2--O--, --O--P(O,NR.sup.H)--O--,
--O--PO(R'')--O--, --O--PO(CH.sub.3)--O--, and
--O--PO(NHR.sup.N)--O--, where R.sup.H is selected form hydrogen
and C.sub.1-4-alkyl, and R'' is selected from C.sub.1-6-alkyl and
phenyl, are especially desirable. Further illustrative examples are
given in Mesmaeker et. al., Current Opinion in Structural Biology
1995, 5, 343-355 and Susan M. Freier and Karl-Heinz Altmann,
Nucleic Acids Research, 1997, vol 25, pp 4429-4443. The left-hand
side of the internucleoside linkage is bound to the 5-membered ring
as substituent P* at the 3'-position, whereas the right-hand side
is bound to the 5'-position of a preceding monomer.
[0047] By "LNA" or "LNA monomer" (e.g., an LNA nucleoside or LNA
nucleotide) or an LNA oligomer (e.g., an oligonucleotide or nucleic
acid) is meant a nucleoside or nucleotide analogue that includes at
least one LNA monomer. LNA monomers as disclosed in PCT Publication
WO 99/14226 are in general particularly desirable modified nucleic
acids for incorporation into an oligonucleotide of the invention.
Additionally, the nucleic acids may be modified at either the 3'
and/or 5' end by any type of modification known in the art. For
example, either or both ends may be capped with a protecting group,
attached to a flexible linking group, attached to a reactive group
to aid in attachment to the substrate surface, etc. Desirable LNA
monomers and their method of synthesis also are disclosed in U.S.
Pat. No. 6,043,060, U.S. Pat. No. 6,268,490, PCT Publications WO
01/07455, WO 01/00641, WO 98/39352, WO 00/56746, WO 00/56748 and WO
00/66604 as well as in the following papers: Morita et al., Bioorg.
Med. Chem. Lett. 12(1):73-76, 2002; Hakansson et al., Bioorg. Med.
Chem. Lett. 11(7):935-938, 2001; Koshkin et al., J. Org. Chem.
66(25):8504-8512, 2001; Kvaerno et al., J. Org. Chem.
66(16):5498-5503, 2001; Hakansson et al., J. Org. Chem.
65(17):5161-5166, 2000; Kvaerno et al., J. Org. Chem.
65(17):5167-5176, 2000; Pfundheller et al., Nucleosides Nucleotides
18(9):2017-2030, 1999; and Kumar et al., Bioorg. Med. Chem. Lett.
8(16):2219-2222, 1998.
[0048] Preferred LNA monomers, also referred to as "oxy-LNA" are
LNA monomers which include bicyclic compounds as disclosed in PCT
Publication WO 03/020739 wherein the bridge between R.sup.4' and
R.sup.2' as shown in formula (I) below together designate
--CH.sub.2--O-- or --CH.sub.2--CH.sub.2--O--.
[0049] By "LNA modified oligonucleotide" or "LNA substituted
oligonucleotide" is meant a oligonucleotide comprising at least one
LNA monomer of formula (I), described infra, having the below
described illustrative examples of modifications:
##STR00001##
wherein X is selected from --O--, --S--, --N(R.sup.N)--,
--C(R.sup.6R.sup.6*)--, --O--C(R.sup.7R.sup.7*)--,
--C(R.sup.6R.sup.6*)--O--, --S--C(R.sup.7R.sup.7*)--,
--C(R.sup.6R.sup.6*)--S--, --N(R.sup.N*)--C(R.sup.7R.sup.7*)--,
--C(R.sup.6R.sup.6*)--N(R.sup.N*)--, and
--C(R.sup.6R.sup.6*)--C(R.sup.7R.sup.7*).
[0050] B is selected from a modified base as discussed above e.g.
an optionally substituted carbocyclic aryl such as optionally
substituted pyrene or optionally substituted pyrenylmethylglycerol,
or an optionally substituted heteroalicylic or optionally
substituted heteroaromatic such as optionally substituted
pyridyloxazole, optionally substituted pyrrole, optionally
substituted diazole or optionally substituted triazole moieties;
hydrogen, hydroxy, optionally substituted C.sub.1-4-alkoxy,
optionally substituted C.sub.1-4-alkyl, optionally substituted
C.sub.1-4-acyloxy, nucleobases, DNA intercalators, photochemically
active groups, thermochemically active groups, chelating groups,
reporter groups, and ligands.
[0051] P designates the radical position for an internucleoside
linkage to a succeeding monomer, or a 5'-terminal group, such
internucleoside linkage or 5'-terminal group optionally including
the substituent R.sup.5. One of the substituents R.sup.2, R.sup.2*,
R.sup.3, and R.sup.3* is a group P* which designates an
internucleoside linkage to a preceding monomer, or a 2'/3'-terminal
group. The substituents of R.sup.1*, R.sup.4*, R.sup.5, R.sup.5*,
R.sup.6, R.sup.6*, R.sup.7, R.sup.7*, R.sup.N, and the ones of
R.sup.2, R.sup.2*, R.sup.3, and R.sup.3* not designating P* each
designates a biradical comprising about 1-8 groups/atoms selected
from --C(R.sup.aR.sup.b)--, --C(R.sup.a).dbd.C(R.sup.a)--,
--C(R.sup.a).dbd.N--, --C(R.sup.a)--O--, --O--,
--Si(R.sup.a).sub.2--, --C(R.sup.a)--S, --S--, --SO.sub.2--,
--C(R.sup.a)--N(R.sup.b--, --N(R.sup.a)--, and >C=Q, wherein Q
is selected from --O--, --S--, and --N(R.sup.a)--, and R.sup.a and
R.sup.b each is independently selected from hydrogen, optionally
substituted C.sub.1-12-alkyl, optionally substituted
C.sub.2-12-alkenyl, optionally substituted C.sub.2-12-alkynyl,
hydroxy, C.sub.1-12-alkoxy, C.sub.2-12-alkenyloxy, carboxy,
C.sub.1-12-alkoxycarbonyl, C.sub.1-12-alkylcarbonyl, formyl, aryl,
aryloxy-carbonyl, aryloxy, arylcarbonyl, heteroaryl,
hetero-aryloxy-carbonyl, heteroaryloxy, heteroarylcarbonyl, amino,
mono- and di(C.sub.1-6-alkyl)amino, carbamoyl, mono- and
di(C.sub.1-6-alkyl)-amino-carbonyl,
amino-C.sub.1-6-alkyl-aminocarbonyl, mono- and
di(C.sub.1-6-alkyl)amino-C.sub.1-6-alkyl-aminocarbonyl,
C.sub.1-6-alkyl-carbonylamino, carbamido, C.sub.1-6-alkanoyloxy,
sulphono, C.sub.1-6-alkylsulphonyloxy, nitro, azido, sulphanyl,
C.sub.1-6-alkylthio, halogen, DNA intercalators, photochemically
active groups, thermochemically active groups, chelating groups,
reporter groups, and ligands, where aryl and heteroaryl may be
optionally substituted, and where two geminal substituents R.sup.a
and R.sup.b together may designate optionally substituted methylene
(.dbd.CH.sub.2), and wherein two non-geminal or geminal
substituents selected from R.sup.a, R.sup.b, and any of the
substituents R.sup.2, R.sup.2*, R.sup.3, R.sup.3*, R.sup.4*,
R.sup.5, R.sup.5*, R.sup.6 and R.sup.6*, R.sup.7, and R.sup.7*
which are present and not involved in P, P* or the biradical(s)
together may form an associated biradical selected from biradicals
of the same kind as defined before; the pair(s) of non-geminal
substituents thereby forming a mono- or bicyclic entity together
with (i) the atoms to which said non-geminal substituents are bound
and (ii) any intervening atoms.
[0052] Each of the substituents R.sup.1*, R.sup.2, R.sup.2*,
R.sup.3, R.sup.4*, R.sup.5, R.sup.5*, R.sup.6 and R.sup.6*,
R.sup.7, and R.sup.7* which are present and not involved in P, P*
or the biradical(s), is independently selected from hydrogen,
optionally substituted C.sub.1-12-alkyl, optionally substituted
C.sub.2-12-alkenyl, optionally substituted C.sub.2-12-alkynyl,
hydroxy, C.sub.2-12-alkenyloxy, carboxy, C.sub.1-12-alkoxycarbonyl,
C.sub.1-12-alkylcarbonyl, formyl, aryl, aryloxy-carbonyl, aryloxy,
arylcarbonyl, heteroaryl, heteroaryloxy-carbonyl, heteroaryloxy,
heteroarylcarbonyl, amino, mono- and di-(C.sub.1-6-alkyl)amino,
carbamoyl, mono- and di(C.sub.1-6-alkyl)-amino-carbonyl,
amino-C.sub.1-6-alkyl-aminocarbonyl, mono- and
di(C.sub.1-6-alkyl)amino-C.sub.1-6-alkyl-aminocarbonyl,
C.sub.1-6-alkyl-carbonylamino, carbamido, C.sub.1-6-alkanoyloxy,
sulphono, C.sub.1-6-alkylsulphonyloxy, nitro, azido, sulphanyl,
C.sub.1-6-alkylthio, halogen, DNA intercalators, photochemically
active groups, thermochemically active groups, chelating groups,
reporter groups, and ligands, where aryl and heteroaryl may be
optionally substituted, and where two geminal substituents together
may designate oxo, thioxo, imino, or optionally substituted
methylene, or together may form a spiro biradical consisting of a
1-5 carbon atom(s) alkylene chain which is optionally interrupted
and/or terminated by one or more heteroatoms/groups selected from
--O--, --S--, and --(NR.sup.N)-- where R.sup.N is selected from
hydrogen and C.sub.1-4-alkyl, and where two adjacent (non-geminal)
substituents may designate an additional bond resulting in a double
bond; and R.sup.N*, when present and not involved in a biradical,
is selected from hydrogen and C.sub.1-4-alkyl; and basic salts and
acid addition salts thereof.
[0053] Exemplary 5', 3', and/or 2' terminal groups include --H,
--OH, halo (e.g., chloro, fluoro, iodo, or bromo), optionally
substituted aryl, (e.g., phenyl or benzyl), alkyl (e.g., methyl or
ethyl), alkoxy (e.g., methoxy), acyl (e.g. acetyl or benzoyl),
aroyl, aralkyl, hydroxy, hydroxyalkyl, alkoxy, aryloxy, aralkoxy,
nitro, cyano, carboxy, alkoxycarbonyl, aryloxycarbonyl,
aralkoxycarbonyl, acylamino, aroylamino, alkylsulfonyl,
arylsulfonyl, heteroarylsulfonyl, alkylsulfinyl, arylsulfinyl,
heteroarylsulfinyl, alkylthio, arylthio, heteroarylthio,
aralkylthio, heteroaralkylthio, amidino, amino, carbamoyl,
sulfamoyl, alkene, alkyne, protecting groups (e.g., silyl,
4,4'-dimethoxytrityl, monomethoxytrityl, or
trityl(triphenylmethyl)), linkers (e.g., a linker containing an
amine, ethylene glycol, quinone such as anthraquinone), detectable
labels (e.g., radiolabels or fluorescent labels), and biotin.
[0054] It is understood that references herein to a nucleic acid
unit, nucleic acid residue, LNA monomer, or similar term are
inclusive of both individual nucleoside units and nucleotide units
and nucleoside units and nucleotide units within an
oligonucleotide.
[0055] A "modified base" or other similar terms refer to a
composition (e.g., a non-naturally occurring nucleobase or
nucleosidic base), which can pair with a natural base (e.g.,
adenine, guanine, cytosine, uracil, and/or thymine) and/or can pair
with a non-naturally occurring nucleobase or nucleosidic base.
Desirably, the modified base provides a T.sub.m differential of 15,
12, 10, 8, 6, 4, or 2.degree. C. or less as described herein.
Exemplary modified bases are described in EP 1 072 679 and WO
97/12896.
[0056] The term "chemical moiety" refers to a part of a molecule.
"Modified by a chemical moiety" thus refer to a modification of the
standard molecular structure by inclusion of an unusual chemical
structure. The attachment of said structure can be covalent or
non-covalent.
[0057] The term "inclusion of a chemical moiety" in an
oligonucleotide probe thus refers to attachment of a molecular
structure. Such as chemical moiety include but are not limited to
covalently and/or non-covalently bound minor groove binders (MGB)
and/or intercalating nucleic acids (INA) selected from a group
consisting of asymmetric cyanine dyes, DAPI, SYBR Green I, SYBR
Green II, SYBR Gold, PicoGreen, thiazole orange, Hoechst 33342,
Ethidium Bromide, 1-O-(1-pyrenylmethyl)glycerol and Hoechst 33258.
Other chemical moieties include the modified nucleobases,
nucleosidic bases or LNA modified oligonucleotides.
[0058] "Oligonucleotide analogue" refers to a nucleic acid binding
molecule capable of recognizing a particular target nucleotide
sequence. A particular oligonucleotide analogue is peptide nucleic
acid (PNA) in which the sugar phosphate backbone of an
oligonucleotide is replaced by a protein like backbone. In PNA,
nucleobases are attached to the uncharged polyamide backbone
yielding a chimeric pseudopeptide-nucleic acid structure, which is
homomorphous to nucleic acid forms.
[0059] "High affinity nucleotide analogue" or "affinity-enhancing
nucleotide analogue" refers to a non-naturally occurring nucleotide
analogue that increases the "binding affinity" of an
oligonucleotide probe to its complementary recognition sequence
when substituted with at least one such high-affinity nucleotide
analogue.
[0060] As used herein, a probe with an increased "binding affinity"
for a recognition sequence compared to a probe which comprises the
same sequence but does not comprise a stabilizing nucleotide,
refers to a probe for which the association constant (K.sub.a) of
the probe recognition segment is higher than the association
constant of the complementary strands of a double-stranded
molecule. In another preferred embodiment, the association constant
of the probe recognition segment is higher than the dissociation
constant (K.sub.d) of the complementary strand of the recognition
sequence in the target sequence in a double stranded molecule.
[0061] Monomers are referred to as being "complementary" if they
contain nucleobases that can form hydrogen bonds according to
Watson-Crick base-pairing rules (e.g. G with C, A with T or A with
U) or other hydrogen bonding motifs such as for example
diaminopurine with T, 5-methyl C with G, 2-thiothymidine with A,
inosine with C, pseudoisocytosine with G, etc.
[0062] The term "succeeding monomer" relates to the neighbouring
monomer in the 5'-terminal direction and the "preceding monomer"
relates to the neighbouring monomer in the 3'-terminal
direction.
[0063] The term "target nucleic acid" or "target ribonucleic acid"
refers to any relevant nucleic acid of a single specific sequence,
e.g., a biological nucleic acid, e.g., derived from a patient, an
animal (a human or non-human animal), a cell, a tissue, an
organism, etc. In one embodiment, the target nucleic acid is
derived from a patient, e.g., a human patient. In this embodiment,
the invention optionally further includes selecting a treatment,
diagnosing a disease, or diagnosing a genetic predisposition to a
disease, based upon detection of the target nucleic acid. Whilst
the target may be a miRNA, such as the miRNA or pre-miRNA sequences
disclosed herein, such as in the context of the target of a
detection probe, the target is also used in the context as the
target of the miRNA, i.e. the mRNA whose expression is regulated by
the miRNA. Table 11 includes human genes whose mRNAs are targeted
by the miRNAs disclosed herein.
[0064] "Target sequence" refers to a specific nucleic acid sequence
within any target nucleic acid.
[0065] The term "stringent conditions", as used herein, is the
"stringency" which occurs within a range from about
T.sub.m-5.degree. C. (5.degree. C. below the melting temperature
(T.sub.m) of the probe) to about 20.degree. C. to 25.degree. C.
below T.sub.m. As will be understood by those skilled in the art,
the stringency of hybridization may be altered in order to identify
or detect identical or related polynucleotide sequences.
Hybridization techniques are generally described in Nucleic Acid
Hybridization, A Practical Approach, Ed. Hames, B. D. and Higgins,
S. J., IRL Press, 1985; Gall and Pardue, Proc. Natl. Acad. Sci.,
USA 63: 378-383, 1969; and John, et al. Nature 223: 582-587,
1969.
[0066] In one embodiment the term "specifically hybridise" is
determined by whether the oligonucleotide or compound of the
invention hybridises to the target nucleic acid sequence (e.g. a
SEQ selected from SEQ ID NO 1-408) under stringent conditions.
[0067] The terms "contiguous sequence of nucleobases" and
"contiguous nucleobase sequence" are used interchangeably.
DETAILED DESCRIPTION OF THE INVENTION
[0068] New microRNA Markers
[0069] The invention provides 558 new microRNA markers which have
not previously been recognised as miRNAs and/or indicated in
cancer. The 558 miRNAs were identified in cancer tissue and are
considered to be useful indicators of cancer. The present invention
provides a detailed expression analysis of both the pre- and mature
forms of the miRNAs in various cancers and corresponding healthy
tissues, and as such the present invention provides an extensive
collection of miRNA sequences which, when in the form of detection
probes, can be used, individually, or in the form of collections
for cancer diagnostics, and for determining the origin of secondary
cancers.
[0070] The new microRNA markers include mature miRNAs and the
premature (pre-processed) miRNAs from which they originate.
[0071] The odd numbered SEQ IDs from 1 to 407 are novel mature
miRNAs. The even numbers from 2 to 408 are the corresponding
pre-mature miRNAs. For SEQ ID Nos 411-558 the miRNAs and their
corresponding pre-miRNAs, in addition to related miRNA (and/or
related pre-miRNAs) are shown in Table 3.
[0072] The invention provides for a nucleic acid or nucleobase
sequence selected from the group consisting of SEQ ID NO 1-558
and/or a compound which comprises a contiguous sequence of
nucleobases of between 8 and 30 nucleobases in length, wherein the
nucleobase sequence of the contiguous sequence corresponds to a
contiguous nucleic acid sequence present in said SEQ ID selected
from the group consisting of SEQ ID 1-558.
[0073] In one embodiment the contiguous nucleobase sequence (which
may be a nuclei acid sequence) is complementary to the
(corresponding) region of said SEQ ID.
[0074] In one embodiment the sequence of nucleobases of the
contiguous nucleobase sequence (which may be a nuclei acid
sequence) is found within the (corresponding) region of said SEQ
ID.
[0075] As in the above embodiments, and with reference to the list
of embodiments herein and below, the term "corresponding to" may
refer to a complementary sequence (preferably 100% complementary)
or that the sequence is found within said sequence (i.e.
homologous, preferably 100% homologus).
[0076] In one embodiment the SEQ ID is a mature miRNA sequence,
i.e. selected from the odd numbered SEQ IDs present in the group
consisting of SEQ IDs No 1-407 and/or column 1 of table 3.
[0077] In one embodiment the SEQ ID is a mature miRNA sequence,
i.e. selected from the group of miRNAs present in the group
consisting of the mature miRNAs listed in Table 5.
[0078] In one embodiment the SEQ ID is a premature miRNA sequence,
i.e. selected from the group of pre-miRNAs listed in Table 4.
[0079] In one embodiment the SEQ ID is a pre-mature miRNA sequence,
i.e. selected from the even numbered SEQ IDs present in the group
consisting of SEQ IDs No 2-408 and/or column 2 of table 3.
[0080] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 1 or 2 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0081] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 3 or 4 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0082] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 5 or 6 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0083] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 7 or 8 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0084] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 9 or 10 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0085] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 11 or 12 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0086] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 13 or 14 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0087] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 15 or 16 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0088] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 17 or 18 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0089] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 19 or 20 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0090] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 21 or 22 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0091] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 23 or 24 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0092] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 25 or 26 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0093] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 27 or 28 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0094] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 29 or 30 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0095] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 31 or 32 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0096] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 33 or 34 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0097] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 35 or 36 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0098] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 37 or 38 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0099] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 39 or 40 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0100] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 41 or 42 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0101] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 43 or 44 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0102] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 45 or 46 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0103] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 47 or 48 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0104] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 49 or 50 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0105] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 51 or 52 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0106] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 53 or 54 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0107] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 55 or 56 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0108] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 57 or 58 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0109] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 59 or 60 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0110] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 61 or 62 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0111] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 63 or 64 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0112] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 65 or 66 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0113] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 67 or 68 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0114] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 69 or 70 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0115] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 71 or 72 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0116] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 73 or 74 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0117] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 75 or 76 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0118] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 77 or 78 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0119] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 79 or 80 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0120] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 81 or 82 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0121] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 83 or 84 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0122] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 85 or 86 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0123] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 87 or 88 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0124] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 89 or 90 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0125] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 91 or 92 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0126] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 93 or 94 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0127] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 95 or 96 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0128] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 97 or 98 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0129] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 99 or 100 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0130] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 101 or 102 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0131] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 103 or 104 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0132] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 105 or 106 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0133] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 107 or 108 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0134] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 109 or 110 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0135] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 111 or 112 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0136] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 113 or 114 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0137] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 115 or 116 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0138] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 117 or 118 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0139] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 119 or 120 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0140] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 121 or 122 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0141] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 123 or 124 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0142] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 125 or 126 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0143] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 127 or 128 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0144] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 129 or 130 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0145] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 131 or 132 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0146] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 133 or 134 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0147] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 135 or 136 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0148] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 137 or 138 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0149] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 139 or 140 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0150] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 141 or 142 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0151] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 143 or 144 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0152] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 145 or 146 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0153] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 147 or 148 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0154] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 149 or 150 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0155] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 151 or 152 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0156] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 153 or 154 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0157] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 155 or 156 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0158] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 157 or 158 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0159] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 159 or 160 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0160] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 161 or 162 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0161] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 163 or 164 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0162] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 165 or 166 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0163] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 167 or 168 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0164] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 169 or 170 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0165] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 171 or 172 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0166] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 173 or 174 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0167] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 175 or 176 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0168] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 177 or 178 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0169] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 179 or 180 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0170] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 181 or 182 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0171] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 183 or 184 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0172] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 185 or 186 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0173] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 187 or 188 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0174] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 189 or 190 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0175] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 191 or 192 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0176] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 193 or 194 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0177] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 195 or 196 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0178] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 197 or 198 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0179] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 199 or 200 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0180] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 201 or 202 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0181] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 203 or 204 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0182] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 205 or 206 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0183] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 207 or 208 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0184] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 209 or 210 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0185] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 211 or 212 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0186] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 213 or 214 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0187] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 215 or 216 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0188] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 217 or 218 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0189] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 219 or 220 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0190] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 221 or 222 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0191] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 223 or 224 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0192] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 225 or 226 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0193] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 227 or 228 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0194] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 229 or 230 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0195] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 231 or 232 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0196] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 233 or 234 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0197] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 235 or 236 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0198] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 237 or 238 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0199] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 239 or 240 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0200] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 241 or 242 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0201] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 243 or 244 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0202] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 245 or 246 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0203] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 247 or 248 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0204] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 249 or 250 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0205] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 251 or 252 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0206] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 253 or 254 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0207] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 255 or 256 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0208] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 257 or 258 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0209] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 259 or 260 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0210] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 261 or 262 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0211] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 263 or 264 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0212] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 265 or 266 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0213] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 267 or 268 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0214] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 269 or 270 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0215] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 271 or 272 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0216] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 273 or 274 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0217] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 275 or 276 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0218] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 277 or 278 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0219] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 279 or 280 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0220] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 281 or 282 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0221] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 283 or 284 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0222] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 285 or 286 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0223] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 287 or 288 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0224] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 289 or 290 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0225] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 291 or 292 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0226] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 293 or 294 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0227] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 295 or 296 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0228] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 297 or 298 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0229] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 299 or 300 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0230] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 301 or 302 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0231] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 303 or 304 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0232] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 305 or 306 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0233] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 307 or 308 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0234] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 309 or 310 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0235] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 311 or 312 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0236] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 313 or 314 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0237] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 315 or 316 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0238] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 317 or 318 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0239] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 319 or 320 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0240] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 321 or 322 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0241] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 323 or 324 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0242] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 325 or 326 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0243] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 327 or 328 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0244] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 329 or 330 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0245] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 331 or 332 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0246] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 333 or 334 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0247] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 335 or 336 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0248] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 337 or 338 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0249] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 339 or 340 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0250] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 341 or 342 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0251] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 343 or 344 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0252] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 345 or 346 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0253] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 347 or 348 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0254] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 349 or 350 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0255] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 351 or 352 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0256] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 353 or 354 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0257] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 355 or 356 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0258] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 357 or 358 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0259] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 359 or 360 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0260] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 361 or 362 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0261] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 363 or 364 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0262] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 365 or 366 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0263] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 367 or 368 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0264] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 369 or 370 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0265] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 371 or 372 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0266] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 373 or 374 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0267] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 375 or 376 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0268] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 377 or 378 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0269] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 379 or 380 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0270] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 381 or 382 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0271] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 383 or 384 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0272] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 385 or 386 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0273] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 387 or 388 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0274] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 389 or 390 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0275] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 391 or 392 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0276] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 393 or 394 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0277] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 395 or 396 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0278] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 397 or 398 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0279] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 399 or 400 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0280] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 401 or 402 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0281] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 403 or 404 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0282] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 405 or 406 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0283] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 407 or 408 and/or a compound which
comprises a contiguous sequence of nucleobases of between 8 and 30
nucleobases in length, wherein the nucleobase sequence of the
contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0284] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 411 and/or 412, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0285] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 413, 414 and/or 415, and/or a
compound which comprises a contiguous sequence of nucleobases of
between 8 and 30 nucleobases in length, wherein the nucleobase
sequence of the contiguous sequence corresponds to a contiguous
nucleic acid or nucleobases sequence present in said SEQ IDs.
[0286] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 416 and/or 417, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0287] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 418, 420, 421 and/or 419, and/or a
compound which comprises a contiguous sequence of nucleobases of
between 8 and 30 nucleobases in length, wherein the nucleobase
sequence of the contiguous sequence corresponds to a contiguous
nucleic acid or nucleobases sequence present in said SEQ IDs.
[0288] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 422 and/or 423, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0289] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 424 and/or 425, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0290] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 426, 428 and/or 427, and/or a
compound which comprises a contiguous sequence of nucleobases of
between 8 and 30 nucleobases in length, wherein the nucleobase
sequence of the contiguous sequence corresponds to a contiguous
nucleic acid or nucleobases sequence present in said SEQ IDs.
[0291] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 429, 430, 431, 432 and/or 433,
and/or a compound which comprises a contiguous sequence of
nucleobases of between 8 and 30 nucleobases in length, wherein the
nucleobase sequence of the contiguous sequence corresponds to a
contiguous nucleic acid or nucleobases sequence present in said SEQ
IDs.
[0292] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 434 and/or 435, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0293] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 436 and/or 437, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0294] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 438 and/or 439, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0295] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 440 and/or 441, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0296] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 442 and/or 443, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0297] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 444, 445 and/or 446, and/or a
compound which comprises a contiguous sequence of nucleobases of
between 8 and 30 nucleobases in length, wherein the nucleobase
sequence of the contiguous sequence corresponds to a contiguous
nucleic acid or nucleobases sequence present in said SEQ IDs.
[0298] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 447 and/or 448, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0299] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 449, 451 and/or 450, and/or a
compound which comprises a contiguous sequence of nucleobases of
between 8 and 30 nucleobases in length, wherein the nucleobase
sequence of the contiguous sequence corresponds to a contiguous
nucleic acid or nucleobases sequence present in said SEQ IDs.
[0300] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 452 and/or 453, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0301] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 454 and/or 455, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0302] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 456 and/or 457, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0303] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 458 and/or 459, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0304] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 460 and/or 461, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0305] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 462 and/or 463, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0306] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 464 and/or 465, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0307] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 466, 468 and/or 467, and/or a
compound which comprises a contiguous sequence of nucleobases of
between 8 and 30 nucleobases in length, wherein the nucleobase
sequence of the contiguous sequence corresponds to a contiguous
nucleic acid or nucleobases sequence present in said SEQ IDs.
[0308] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 469, 471, 472 and/or 470, and/or a
compound which comprises a contiguous sequence of nucleobases of
between 8 and 30 nucleobases in length, wherein the nucleobase
sequence of the contiguous sequence corresponds to a contiguous
nucleic acid or nucleobases sequence present in said SEQ IDs.
[0309] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 473 and/or 474, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0310] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 475, 477 and/or 476, and/or a
compound which comprises a contiguous sequence of nucleobases of
between 8 and 30 nucleobases in length, wherein the nucleobase
sequence of the contiguous sequence corresponds to a contiguous
nucleic acid or nucleobases sequence present in said SEQ IDs.
[0311] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 478 and/or 479, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0312] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 480 and/or 481, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0313] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 482 and/or 483, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0314] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 484 and/or 485, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0315] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 486 and/or 487, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0316] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 488 and/or 489, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0317] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 490, 492, 493 494, and/or 491,
and/or a compound which comprises a contiguous sequence of
nucleobases of between 8 and 30 nucleobases in length, wherein the
nucleobase sequence of the contiguous sequence corresponds to a
contiguous nucleic acid or nucleobases sequence present in said SEQ
IDs.
[0318] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 495, 497, 498, 499, 501, 496
and/or 500, and/or a compound which comprises a contiguous sequence
of nucleobases of between 8 and 30 nucleobases in length, wherein
the nucleobase sequence of the contiguous sequence corresponds to a
contiguous nucleic acid or nucleobases sequence present in said SEQ
IDs.
[0319] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 502 and/or 503, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0320] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 504, 505 and/or 506, and/or a
compound which comprises a contiguous sequence of nucleobases of
between 8 and 30 nucleobases in length, wherein the nucleobase
sequence of the contiguous sequence corresponds to a contiguous
nucleic acid or nucleobases sequence present in said SEQ IDs.
[0321] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 507 and/or 508, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0322] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 509 and/or 510, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0323] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 511 and/or 512, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0324] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 513 and/or 514, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0325] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 515, 517, and/or 516, and/or a
compound which comprises a contiguous sequence of nucleobases of
between 8 and 30 nucleobases in length, wherein the nucleobase
sequence of the contiguous sequence corresponds to a contiguous
nucleic acid or nucleobases sequence present in said SEQ IDs.
[0326] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 518 and/or 519, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0327] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 520 and/or 521, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0328] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 522 and/or 523, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0329] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 524 and/or 525, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0330] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 526 and/or 527, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0331] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 528 and/or 529, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0332] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 530 and/or 531, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0333] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 532 and/or 533, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0334] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 534 and/or 535, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0335] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 536 and/or 537, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0336] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 538, 540 and/or 539, and/or a
compound which comprises a contiguous sequence of nucleobases of
between 8 and 30 nucleobases in length, wherein the nucleobase
sequence of the contiguous sequence corresponds to a contiguous
nucleic acid or nucleobases sequence present in said SEQ IDs.
[0337] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 541, 543 and/or 542, and/or a
compound which comprises a contiguous sequence of nucleobases of
between 8 and 30 nucleobases in length, wherein the nucleobase
sequence of the contiguous sequence corresponds to a contiguous
nucleic acid or nucleobases sequence present in said SEQ IDs.
[0338] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 544 and/or 545, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0339] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 546 and/or 547, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0340] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 548, 551, 552, 553, and/or 554,
and/or a compound which comprises a contiguous sequence of
nucleobases of between 8 and 30 nucleobases in length, wherein the
nucleobase sequence of the contiguous sequence corresponds to a
contiguous nucleic acid or nucleobases sequence present in said SEQ
IDs.
[0341] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 549, 552, 553 and/or 554, and/or a
compound which comprises a contiguous sequence of nucleobases of
between 8 and 30 nucleobases in length, wherein the nucleobase
sequence of the contiguous sequence corresponds to a contiguous
nucleic acid or nucleobases sequence present in said SEQ IDs.
[0342] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 550, 553 and/or 554, and/or a
compound which comprises a contiguous sequence of nucleobases of
between 8 and 30 nucleobases in length, wherein the nucleobase
sequence of the contiguous sequence corresponds to a contiguous
nucleic acid or nucleobases sequence present in said SEQ IDs.
[0343] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 555 and/or 556, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
[0344] In one embodiment, the invention provides for a nucleic acid
or nucleobase sequence SEQ ID No 557 and/or 558, and/or a compound
which comprises a contiguous sequence of nucleobases of between 8
and 30 nucleobases in length, wherein the nucleobase sequence of
the contiguous sequence corresponds to a contiguous nucleic acid or
nucleobases sequence present in said SEQ IDs.
A Compound
[0345] In a preferred embodiment, in the compound of the invention,
the contiguous sequence of nucleobases is in the form of, or
comprises an oligonucleotide.
[0346] The oligonucleotide may comprise nucleotide nucleobases
and/or nucleotide analogue nucleobases.
[0347] The oligonucleotide (compound) of the invention may, for
example be used in antisense therapy or as a research tool or
diagnostic for a disease.
[0348] The oligonucleotide (compound) of the invention may
comprises at least one nucleotide analogue, such as a LNA.
[0349] The oligonucleotide (compound) of the invention may
comprises between 8 to 30 nucleobases, such as 9, 10, 11, 12, 13,
14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, or 29
nucleobases, such as between 14 to 24 nucleobases, such as between
16 and 22 nucleobases, such as between 8 to 14 nucleobases, such as
between 12 and 20 nucleobases.
[0350] The oligonucleotide (compound) of the invention may comprise
other oligonucleotide analogues in combination with or without LNA.
Such analogues may be selected from the group consisting of
phosphorothioates or 2'-O-protected nucleotides or combinations
thereof.
[0351] Suitable nucleotide analogues include those selected from
the group consisting of Locked Nucleic Acid (LNA) units;
2'-O-alkyl-RNA units, 2'-OMe-RNA units, 2'-amino-DNA units,
2'-fluoro-DNA units, PNA units, HNA units, and INA units.
[0352] In one embodiment, the compound according to the invention
comprises a duplex formed between the contiguous sequence of
nucleobases and a further nucleobase sequence which is
complementary to the contiguous sequence of nucleobases.
[0353] In one embodiment, the compound according to the invention
is in the form of a siRNA or miRNA.
[0354] In one embodiment the compound of the invention is in the
form of a miRNA mimic which can be introduced into a cell to
repress the expression of one or more mRNA target(s) present in the
cell (such as one or more of the mRNA targets listed in Table 11).
miRNA mimics are typically fully complementary to the full length
miRNA sequence.
[0355] The invention therefore provides a method of inducing or
enhancing miRNA mediated repression of a mRNA target in a cell,
said method comprising the step of introducing the compound of the
invention into said cell, so as to repress the mRNA target. miRNA
mimics are compounds comprising a contiguous nucleobase sequence
which are homologous to a corresponding region of one, or more, of
the miRNA sequences provided herein (i.e. sense compounds).
[0356] In one embodiment the compound of the invention is an
anti-miR which can be introduced into a cell to deactivate miRNA
alleviating the miRNA-induced repression of the mRNA target (i.e
de-repression).
[0357] The invention therefore provides a method of reducing or
deactivating a miRNA in a cell, said method comprising the step of
introducing the compound of the invention into said cell, so as to
reduce or deactivate said miRNA in the cell. AntimiR compounds
comprise a contiguous nucleobase sequence which is complementary to
a corresponding region of one, or more, of the miRNA sequences
provided herein (i.e. antisense compounds).
[0358] Numerous examples of the application of miRNA micmics and
antimiRs are known in the art (Guimaraes-Sternberg et al., Leukemia
Research 30(5): 583-595, 2006). For example, WO2007/112754, which
is hereby incorporated by reference, provides suitable designs for
antimiR compounds according to the invention.
Conjugates
[0359] The invention also provides for a conjugate comprising the
compound according to the invention and at least one non-nucleobase
moiety covalently attached thereto. Suitable conjugates for use
with oligonucleotides/oligonucleobases are known in the art, such
as those disclosed in US 2006154888.
Pharmaceutical Composition
[0360] The invention also provides for a pharmaceutical composition
comprising a compound according to the invention, or the conjugate
according to the invention, and at least one pharmaceutically
acceptable diluent, carrier, salt or adjuvant. Suitable
pharmaceutically acceptable diluent, carrier, salt or adjuvant are
known in the art, such as those disclosed in US 2006154888.
[0361] The compound may be formulated as a prodrug, which is
activated once it enters into the cell (such as the prodrug
formulations disclosed in US 2006154888).
First Medical Indication
[0362] The invention also provides for the use of a compound
according to the invention or the conjugate according to the
invention or the pharmaceutical composition according to the
invention as a medicament, such as for the treatment of a disease
or medical condition selected from the group consisting of: cancer,
such as a form of cancer selected from the group consisting of;
breast cancer, adrenal gland cancer, bladder cancer, colon cancer,
Cervical cancer, cancer of the duodenum, cancer of the esophagus,
cancer of the kidney, liver cancer, lung cancer, ovarian cancer,
prostate cancer, rectal cancer, stomach cancer and uterine
cancer.
[0363] In one embodiment, the type of said cancer may be selected
from the group consisting of the following: A solid tumor; ovarian
cancer, breast cancer, non-small cell lung cancer, renal cell
cancer, bladder cancer, esophagus cancer, stomach cancer, prostate
cancer, pancreatic cancer, lung cancer, cervical cancer, colon
cancer, colorectal cancer, renal cell cancer;
[0364] The type of cancer may be selected from the group consisting
of: A carcinoma, such as a carcinoma selected from the group
consisting of ovarian carcinoma, breast carcinoma, non-small cell
lung cancer, renal cell carcinoma, bladder carcinoma, recurrent
superficial bladder cancer, stomach carcinoma, prostatic carcinoma,
pancreatic carcinoma, lung carcinoma, cervical carcinoma, cervical
dysplasia, laryngeal papillomatosis, colon carcinoma, colorectal
carcinoma, carcinoid tumors, renal cell carcinoma, A basal cell
carcinoma; A malignant melanoma, such as a malignant melanoma
selected from the group consisting of superficial spreading
melanoma, nodular melanoma, lentigo maligna melanoma, acral
melagnoma, amelanotic melanoma and desmoplastic melanoma; A
sarcoma, such as a sarcoma selected from the group consisting of
osteosarcoma, Ewing's sarcoma, chondrosarcoma, malignant fibrous
histiocytoma, fibrosarcoma and Kaposi's sarcoma; and a glioma.
[0365] In one embodiment the type of cancer is a teratoma.
Second Medical Indication
[0366] The invention also provides for the use of a compound
according to the invention or the conjugate according to the
invention or the pharmaceutical composition according to the
invention, in the manufacture of a medicament for the treatment of
a disease or medical condition selected from the group consisting
of cancer, such as breast cancer, or one of the above listed forms
of cancer.
Method of Treatment
[0367] The invention also provides for a method of performing
therapy comprising administering the compound according to the
invention or the conjugate according to the invention or the
pharmaceutical composition according to the invention to a patient
in need of therapy, such as a patient suffering from, or
susceptible to be suffering from cancer, such as breast cancer, or
a cancer selected from one of the above listed forms of cancer.
miRNA Seed--Method of Identifying Genes Regulated by miRNAs
[0368] The invention also provides for a method of identifying a
target to an miRNA in an organism, comprising acts of: providing a
conserved miRNA sequence selected from the group consisting of SEQ
ID No 1-558 or natural allelic variants thereof; providing a genome
of an organism; defining at least 6 nucleotides (such as between 6
and 30 nucleotides, such as at least 10, 12, 14 or at least 16
nucleotides) of the conserved miRNA sequence as an miRNA seed;
identifying a conserved UTR of a gene within the genome of the
organism; and identifying the gene as a target of the miRNA by
determining whether the conserved UTR comprises a segment having
perfect complementarity with the miRNA seed. Table 11 provides a
list of human genes whose mRNA are targeted by the miRNAs of the
invention.
Method of Modulating Gene Expression
[0369] A method according of modulating the expression of a gene
associated with a disease or medical condition, comprising
administering the compound according to the invention or the
conjugate according to the invention or the pharmaceutical
composition according to the invention to a cell so as to modulate
the expression of the gene associated with the disease or medical
condition. Table 11 provides a list of possible human genes whose
mRNA are targeted by the miRNAs of the invention. As described
above, the use of miRNA mimics or antimiRs can be used to
(optionally) further repress the mRNA targets, or to silence
(down-regulate) the miRNA, thereby inhibiting the function of the
endogenous miRNA, causing de-repression and increased expression of
the mRNA target.
Detection/Oligonucleotide Probes
Detection Probe and Recognition Sequence
[0370] Typically, an oligonucleotide probe (detection probe) of the
invention includes a plurality of nucleotide analogue monomers and
hybridizes to a miRNA or miRNA precursor. In one embodiment, the
nucleotide analogue is LNA, such as alpha and/or xylo LNA monomers.
In one embodiment, the oligonucleotide probe hybridizes to the loop
sequence of a miRNA precursor, e.g., to 5 nucleotides of the miRNA
precursor loop sequence or to the center of the miRNA precursor
loop sequence. The oligonucleotide probe may or may not also
hybridize to the stem sequence of the miRNA precursor. The
oligonucleotide probe may have a number of nucleotide analogue
monomers corresponding to 20% to 40% of the probe oligonucleotides.
The probes may also have a spacing between nucleotide analogue
monomers such that two of the plurality of nucleotide analogue
monomers are disposed 3 or 4 nucleotides apart, or a combination
thereof. Alternatively, each nucleotide analogue monomer in a probe
may be spaced 3 or 4 nucleotides from the closest nucleotide
analogue monomer. Typically, when nucleotide analogue monomers are
spaced apart, only naturally-occurring nucleotides are disposed
between the nucleotide analogue monomers. Alternatively, two,
three, four, or more nucleotide analogue monomers may be disposed
adjacent to one another. The adjacent nucleotide analogue monomers
may or may not be disposed at the 3' or 5' end of the
oligonucleotide probe or so that one of the nucleotide analogue
monomers hybridizes to the center of the loop sequence of the miRNA
precursor. The probe may include none or at most one mismatched
base, deletion, or addition. Desirably, the probe hybridizes to the
miRNA or precursor thereof under stringent conditions or high
stringency conditions. Desirably, the melting point of the duplex
formed between the probe and the miRNA precursor is at least
1.degree. C. higher, e.g., at least 5.degree. C., than the melting
point of the duplex formed between the miRNA precursor and a
nucleic acid sequence not having a nucleotide analogue monomer, or
any modified backbone. The probe may include at least 70% DNA; at
least 10% nucleotide analogue monomers; and/or at most 30%
nucleotide analogue monomers.
[0371] The probe may further include a 5' or 3' amino group and/or
a 5' or 3' label, e.g., a fluorescent (such as fluorescein) label,
a radioactive label, or a label that is a complex including an
enzyme (such as a complex containing digoxigenin (DIG). The probe
is for example 8 nucleotides to 30 nucleotides long, e.g., 12
nucleotides long or 15 nucleotides long. Other potential
modifications of probes are described herein.
[0372] The probe when hybridized to the miRNA or precursor thereof
may or may not provide a substrate for RNase H. Preferably, the
probes of the invention exhibit increased binding affinity for the
target sequence by at least two-fold, e.g., at least 5-fold or
10-fold, compared to probes of the same sequence without nucleotide
analogue monomers, under the same conditions for hybridization,
e.g., stringent conditions or high stringency conditions.
[0373] The invention features a computer code for a preferred
software program of the invention for the design and selection of
the oligonucleotide probes. The invention provides a method,
system, and computer program embedded in a computer readable medium
("a computer program product") for designing oligonucleotide probes
having at least one stabilizing nucleobase, e.g., such as LNA. The
method includes querying a database of target sequences (listed
herein) and designing oligonucleotide probes which: i) have
sufficient binding stability to bind their respective target
sequence under stringent hybridization conditions, ii) have limited
propensity to self-anneal, and iii) are capable of binding to and
detecting/quantifying at least about 60%, at least about 70%, at
least about 80%, at least about 90% or at least about 95% of their
target sequences in the given database of miRNAs, miRNA precursors,
or other RNA sequences.
[0374] The method further entails calculating stability based on
the assumption that the oligonucleotide probes have at least one
stabilizing nucleotide, such as an LNA monomer. In some cases, the
calculated stability is used to eliminate oligonucleotide probe
candidates with inadequate stability from the database of possible
oligonucleotide probes prior to the query against the database to
identify optimal sequences for the oligonucleotide probe.
[0375] In one embodiment, oligonucleotide probes were designed as
reverse complementary sequences to the loop-region of the
pre-miRNA. A stretch of 25 nucleotides were identified centered
around the loop-region and a capture probe was designed for this
25-mer sequence using the same design rules as for capture probes
for the mature miRNAs. This design process takes into account
predictions of Tm of the capture probe, self-hybridization of the
capture probe to it-self and intra-molecular secondary structures
and the difference between Tm and self-hybridization Tm. Further
criteria to the capture probe design, includes that, in one
embodiment, LNA-residues are not allowed in the 3'-end to enhance
synthesis yield. Inter-probe comparison of capture probes against
different miRNAs ensure that capture probes are designed against
regions of the miRNAs that differ the most from other miRNAs in
order to optimize the discrimination between different miRNAs.
[0376] The compound of the invention may form a detection probe,
such as oligonucleotide probes, including detection probe pairs,
which are independently capable of detecting one of each of the
herein listed (miRNA or pre-miRNA) nucleic acids.
[0377] Therefore the invention also provides for detection
probes.
[0378] Each detection probe comprises a recognition sequence
consisting of nucleobases or equivalent molecular entities.
[0379] The recognition sequence typically comprises of the
contiguous sequence of nucleobases present in the compound of the
invention, although in one embodiment, for use in a detection
probe, the recognition sequence may be as short as 6 nucleotides,
such as 6 or 7 nucleobases.
[0380] The recognition sequence of the diagnostic probe according
to the invention corresponds to the target nucleotide sequence or
sequences as referred to herein.
[0381] The target sequences with odd SEQ IDs (from SEQ ID NO 1 to
407) or in column 1 of table 3, or as shown in Table 5, represent
mature miRNA sequences which are associated with cancer such as
breast cancer. Therefore in one embodiment, the detection probe(s)
according to the invention comprise a contiguous sequence of
nucleobases of at least 6 nucleobases, wherein the contiguous
sequence corresponds (i.e. is homolgous to or complementary) to a
contiguous nucleotide sequence present in the odd numbered SEQ IDs
NO 1 to 407 and/or the SEQ IDs listed in column 1 of table 3, or in
Table 5.
[0382] The target sequences with even SEQ IDs (from SEQ ID NO 2 to
408) or in column 2 of table 3, or as shown in Table 4, represent
pre-mature miRNA sequences which are associated with cancer such as
breast cancer. Therefore in one embodiment, the detection probe(s)
according to the invention comprise a contiguous sequence of
nucleobases of at least 6 nucleobases, wherein the contiguous
sequence corresponds to a contiguous nucleotide sequence present in
the even numbered SEQ IDs NO 2 to 408 and/or the SEQ IDs listed in
column 2 of table 3, or as shown in table 4.
[0383] In one embodiment, the detection probes are capable of
specifically hybridizing to a target sequence selected from the
group consisting of a nucleic acid sequence selected from the group
consisting of SEQ ID No 1-558
[0384] In one embodiment, the detection probes are capable of
specifically hybridizing to a target sequence selected from the
group consisting of a nucleic acid sequence selected from the group
shown in Table 5.
[0385] In one embodiment, the detection probes are capable of
specifically hybridizing to a target sequence selected from the
group consisting of a nucleic acid sequence selected from the group
shown in Table 4.
[0386] In a preferred embodiment, detection probes which are
capable of specifically hybridizing to a target sequence have a
contiguous nucleobase sequence which is complementary to a
corresponding region in the target sequence.
[0387] In one embodiment, the detection probes are capable of
specifically hybridizing to a target sequence selected from the
group consisting of a nucleic acid sequence selected from the group
consisting of SEQ ID No 1-408 (odd and/or even SEQ IDs), and SEQ ID
No 411-558 (i.e. the SEQ IDs listed in columns 1 and/or 2 of Table
3), and allelic variants thereof.
[0388] In one embodiment, the detection probes are capable of
specifically hybridizing to a target sequence selected from the
group consisting of a nucleic acid sequence selected from the group
consisting of the even numbered SEQ IDs present in the group SEQ ID
No 1-408, and/or the SEQ IDs listed in column 2 of table 3, and
allelic variants thereof. These sequences are novel precursor miRNA
sequences which are further processed to form mature non-coding
RNAs.
[0389] In one embodiment, the detection probes are capable of
specifically hybridizing to a target sequence selected from the
group consisting of a nucleic acid sequence selected from the group
consisting of the even numbered SEQ IDs present in the group SEQ ID
No 1-408, and/or the SEQ IDs listed in column 1 of table 3, and
allelic variants thereof. These sequences are novel mature
miRNA.
[0390] In one embodiment, the detection probe or probes are capable
of specifically hybridising to the precursor form of the non-coding
RNA, but are not capable of specifically hybridising to the mature
form of the non-coding RNA. Suitable detection probes are routinely
designed and made utilising the sequence information available,
e.g. by selecting a detection probe which at least partially
hybridises to the loop structure which is cleaved during miRNA
processing. It should be noted that several mature siRNAs may
originate from more than one precursor, hence by designing specific
probes for a particular precursor, highly specific detection probes
for use in the invention may be used.
[0391] It will be understood that whilst in the preferred
embodiment the target sequence are the miRNA or pre-miRNA
precursors themselves, in one embodiment, the target sequence may
be a further nucleotide or nucleobase sequence which retains the
sequence information from the corresponding miRNA/pre-miRNA.
[0392] The detection element of the detection probes according to
the invention may be single or double labeled (e.g. by comprising a
label at each end of the probe, or an internal position). In one
aspect, the detection probe comprises two labels capable of
interacting with each other to produce a signal or to modify a
signal, such that a signal or a change in a signal may be detected
when the probe hybridizes to a target sequence. A particular aspect
is when the two labels comprise a quencher and a reporter
molecule.
[0393] A particular detection aspect of the invention referred to
as a "molecular beacon with a stem region" is when the recognition
segment is flanked by first and second complementary
hairpin-forming sequences which may anneal to form a hairpin. A
reporter label is attached to the end of one complementary sequence
and a quenching moiety is attached to the end of the other
complementary sequence. The stem formed when the first and second
complementary sequences are hybridized (i.e., when the probe
recognition segment is not hybridized to its target) keeps these
two labels in close proximity to each other, causing a signal
produced by the reporter to be quenched by fluorescence resonance
energy transfer (FRET). The proximity of the two labels is reduced
when the probe is hybridized to a target sequence and the change in
proximity produces a change in the interaction between the labels.
Hybridization of the probe thus results in a signal (e.g.
fluorescence) being produced by the reporter molecule, which can be
detected and/or quantified.
[0394] Preferably, the compound of the invention, such as the
detection probes of the invention, are modified in order to
increase the binding affinity of the probes for the target sequence
by at least two-fold compared to probes of the same sequence
without the modification, under the same conditions for
hybridization or stringent hybridization conditions. The preferred
modifications include, but are not limited to, inclusion of
nucleobases, nucleosidic bases or nucleotides that have been
modified by a chemical moiety or replaced by an analogue to
increase the binding affinity. The preferred modifications may also
include attachment of duplex-stabilizing agents e.g., such as
minor-groove-binders (MGB) or intercalating nucleic acids (INA).
Additionally, the preferred modifications may also include addition
of non-discriminatory bases e.g., such as 5-nitroindole, which are
capable of stabilizing duplex formation regardless of the
nucleobase at the opposing position on the target strand. Finally,
multi-probes composed of a non-sugar-phosphate backbone, e.g. such
as PNA, that are capable of binding sequence specifically to a
target sequence are also considered as a modification. All the
different binding affinity-increasing modifications mentioned above
will in the following be referred to as "the stabilizing
modification(s)", and the tagging probes and the detection probes
will in the following also be referred to as "modified
oligonucleotide". More preferably the binding affinity of the
modified oligonucleotide is at least about 3-fold, 4-fold, 5-fold,
or 20-fold higher than the binding of a probe of the same sequence
but without the stabilizing modification(s).
[0395] Most preferably, the stabilizing modification(s) is
inclusion of one or more LNA nucleotide analogs. Probes from 8 to
30 nucleotides according to the invention may comprise from 1 to 8
stabilizing nucleotides, such as LNA nucleotides. When at least two
LNA nucleotides are included, these may be consecutive or separated
by one or more non-LNA nucleotides. In one aspect, LNA nucleotides
are alpha-L-LNA and/or xylo LNA nucleotides as disclosed in PCT
Publications No. WO 2000/66604 and WO 2000/56748.
[0396] In a preferable embodiment, each detection probe preferably
comprises affinity enhancing nucleobase analogues, and wherein the
recognition sequences exhibit a combination of high melting
temperatures and low self-complementarity scores, said melting
temperatures being the melting temperature of the duplex between
the recognition sequence and its complementary DNA or RNA
sequence.
[0397] This design provides for probes which are highly specific
for their target sequences but which at the same time exhibits a
very low risk of self-annealing (as evidenced by a low
self-complementarity score)--self-annealing is, due to the presence
of affinity enhancing nucleobases (such as LNA monomers) a problem
which is more serious than when using conventional
deoxyribonucleotide probes.
[0398] In one embodiment the recognition sequences exhibit a
melting temperature (or a measure of melting temperature)
corresponding to at least 5.degree. C. higher than a melting
temperature or a measure of melting temperature of the
self-complementarity score under conditions where the probe
hybridizes specifically to its complementary target sequence
(alternatively, one can quantify the "risk of self-annealing"
feature by requiring that the melting temperature of the
probe-target duplex must be at least 5.degree. C. higher than the
melting temperature of duplexes between the probes or the probes
internally).
[0399] In a preferred embodiment all of the detection probes
include recognition sequences which exhibit a melting temperature
or a measure of melting temperature corresponding to at least
5.degree. C. higher than a melting temperature or a measure of
melting temperature of the self-complementarity score under
conditions where the probe hybridizes specifically to its
complementary target sequence.
[0400] However, it is preferred that this temperature difference is
higher, such as at least 10.degree. C., such as at least 15, at
least 20, at least 25, at least 30, at least 35, at least 40, at
least 45, and at least 50.degree. C. higher than a melting
temperature or measure of melting temperature of the
self-complementarity score.
[0401] In one embodiment, the affinity-enhancing nucleobase
analogues are regularly spaced between the nucleobases in said
detection probes. One reason for this is that the time needed for
adding each nucleobase or analogue during synthesis of the probes
of the invention is dependent on whether or not a nucleobase
analogue is added. By using the "regular spacing strategy"
considerable production benefits are achieved. Specifically for LNA
nucleobases, the required coupling times for incorporating LNA
amidites during synthesis may exceed that required for
incorporating DNA amidites. Hence, in cases involving simultaneous
parallel synthesis of multiple oligonucleotides on the same
instrument, it is advantageous if the nucleotide analogues such as
LNA are spaced evenly in the same pattern as derived from the
3'-end, to allow reduced cumulative coupling times for the
synthesis. The affinity enhancing nucleobase analogues are
conveniently regularly spaced as every 2.sup.nd, every 3.sup.rd,
every 4.sup.th or every 5.sup.th nucleobase in the recognition
sequence, and preferably as every 3.sup.rd nucleobase.
Alternatively, the affinity enhancing nucleobase analogues may be
spaced at a mixture of, for example every 2.sup.nd, every 3.sup.rd,
every 4.sup.th nucleobase.
[0402] The presence of the affinity enhancing nucleobases in the
recognition sequence preferably confers an increase in the binding
affinity between a probe and its complementary target nucleotide
sequence relative to the binding affinity exhibited by a
corresponding probe, which only include nucleobases. Since LNA
nucleobases/monomers have this ability, it is preferred that the
affinity enhancing nucleobase analogues are LNA nucleobases.
[0403] In some embodiments, the 3' and 5' nucleobases are not
substituted by affinity enhancing nucleobase analogues.
[0404] As detailed herein, one huge advantage of such probes for
use in the method of the invention is their short lengths which
surprisingly provides for high target specificity and advantages in
detecting small RNAs and detecting nucleic acids in samples not
normally suitable for hybridization detection strategies. It is,
however, preferred that the probes comprise a recognition sequence
is at least a 6-mer, such as at least a 7-mer, at least an 8-mer,
at least a 9-mer, at least a 10-mer, at least an 11-mer, at least a
12-mer, at least a 13-mer, at least a 14-mer, at least a 15-mer, at
least a 16-mer, at least a 17-mer, at least an 18-mer, at least a
19-mer, at least a 20-mer, at least a 21-mer, at least a 22-mer, at
least a 23-mer, and at least a 24-mer. On the other hand, the
recognition sequence is preferably at most a 25-mer, such as at
most a 24-mer, at most a 23-mer, at most a 22-mer, at most a
21-mer, at most a 20-mer, at most a 19-mer, at most an 18-mer, at
most a 17-mer, at most a 16-mer, at most a 15-mer, at most a
14-mer, at most a 13-mer, at most a 12-mer, at most an 11-mer, at
most a 10-mer, at most a 9-mer, at most an 8-mer, at most a 7-mer,
and at most a 6-mer.
[0405] The present invention provides oligonucleotide compositions
and probe sequences for the use in detection, isolation,
purification, amplification, identification, quantification, or
capture of miRNAs, their target mRNAs, precursor RNAs, stem-loop
precursor miRNAs, siRNAs, other non-coding RNAs, RNA-edited
transcripts or alternative mRNA splice variants or single stranded
DNA (e.g. viral DNA) characterized in that the probe sequences
contain a number of nucleoside analogues.
[0406] In a preferred embodiment the number of nucleoside analogue
corresponds to from 20 to 40% of the oligonucleotide of the
invention.
[0407] In a preferred embodiment the probe sequences are
substituted with a nucleoside analogue with regular spacing between
the substitutions
[0408] In another preferred embodiment the probe sequences are
substituted with a nucleoside analogue with irregular spacing
between the substitutions
[0409] In a preferred embodiment the nucleoside analogue is
LNA.
[0410] In a further preferred embodiment the detection probe
sequences comprise a photochemically active group, a
thermochemically active group, a chelating group, a reporter group,
or a ligand that facilitates the direct of indirect detection of
the probe or the immobilisation of the oligonucleotide probe onto a
solid support.
[0411] In a further preferred embodiment:
(a) the photochemically active group, the thermochemically active
group, the chelating group, the reporter group, or the ligand
includes a spacer (K), said spacer comprising a chemically
cleavable group; or (b) the photochemically active group, the
thermochemically active group, the chelating group, the reporter
group, or the ligand is attached via the biradical of at least one
of the LNA(s) of the oligonucleotide.
[0412] Methods for defining and preparing probes and probe
collections are disclosed in PCT/DK2005/000838.
[0413] In another aspect the invention features detection probe
sequences containing a ligand, which said ligand means something,
which binds. Such ligand-containing detection probes of the
invention are useful for isolating and/or detecting target RNA
molecules from complex nucleic acid mixtures, such as miRNAs, or
their cognate target mRNAs, for example as shown in Table 11.
[0414] The invention therefore also provides for detection probes,
such as oligonucleotide compositions, which are ligands to the
molecular markers according to the invention.
[0415] In another aspect, the invention features detection probes
whose sequences have been furthermore modified by Selectively
Binding Complementary (SBC) nucleobases, i.e. modified nucleobases
that can make stable hydrogen bonds to their complementary
nucleobases, but are unable to make stable hydrogen bonds to other
SBC nucleobases. Such SBC monomer substitutions are especially
useful when highly self-complementary detection probe sequences are
employed. As an example, the SBC nucleobase A', can make a stable
hydrogen bonded pair with its complementary unmodified nucleobase,
T. Likewise, the SBC nucleobase T' can make a stable hydrogen
bonded pair with its complementary unmodified nucleobase, A.
However, the SBC nucleobases A' and T' will form an unstable
hydrogen bonded pair as compared to the base pairs A'-T and A-T'.
Likewise, a SBC nucleobase of C is designated C' and can make a
stable hydrogen bonded pair with its complementary unmodified
nucleobase G, and a SBC nucleobase of G is designated G' and can
make a stable hydrogen bonded pair with its complementary
unmodified nucleobase C, yet C' and G' will form an unstable
hydrogen bonded pair as compared to the base pairs C'-G and C-G'. A
stable hydrogen bonded pair is obtained when 2 or more hydrogen
bonds are formed e.g. the pair between A' and T, A and T', C and
G', and C' and G. An unstable hydrogen bonded pair is obtained when
1 or no hydrogen bonds is formed e.g. the pair between A' and T',
and C' and G'. Especially interesting SBC nucleobases are
2,6-diaminopurine (A', also called D) together with 2-thio-uracil
(U', also called 2SU) (2-thio-4-oxo-pyrimidine) and 2-thio-thymine
(T', also called 2ST) (2-thio-4-oxo-5-methyl-pyrimidine).
[0416] In another aspect, the detection probe sequences of the
invention are covalently bonded to a solid support by reaction of a
nucleoside phosphoramidite with an activated solid support, and
subsequent reaction of a nucleoside phosphoramide with an activated
nucleotide or nucleic acid bound to the solid support. In some
embodiments, the solid support or the detection probe sequences
bound to the solid support are activated by illumination, a
photogenerated acid, or electric current. In other embodiments the
detection probe sequences contain a spacer, e.g. a randomized
nucleotide sequence or a non-base sequence, such as hexaethylene
glycol, between the reactive group and the recognition sequence.
Such covalently bonded detection probe sequence populations are
highly useful for large-scale detection and expression profiling of
mature miRNAs, stem-loop precursor miRNAs, siRNAs, piRNAs, snRNAs
and other non-coding RNAs.
[0417] The present oligonucleotide compositions and detection probe
sequences of the invention are highly useful and applicable for
detection of individual small RNA molecules in complex mixtures
composed of hundreds of thousands of different nucleic acids, such
as detecting mature miRNAs, their target mRNAs, piRNAs, snRNAs or
siRNAs, by Northern blot analysis or for addressing the
spatiotemporal expression patterns of miRNAs, siRNAs or other
non-coding RNAs as well as mRNAs by in situ hybridization in
whole-mount.
[0418] The oligonucleotide compositions and detection probe
sequences are especially applicable for accurate, highly sensitive
and specific detection and quantitation of microRNAs and other
non-coding RNAs, which are useful as biomarkers for diagnostic
purposes of human diseases, such as cancer, including breast cancer
and other cancers referred to herein, as well as for
antisense-based intervention, targeted against tumorigenic miRNAs
and other non-coding RNAs.
[0419] The detection probes, detection probe pairs, and
oligonucleotide compositions and probe sequences which hybridise to
the molecular markers according to the invention are furthermore
applicable for sensitive and specific detection and quantitation of
microRNAs, which can be used as biomarkers for the identification
of the primary site of metastatic tumors of unknown origin.
Known miRNA/Pre-miRNA Control Sequences
[0420] The detection probes of the invention may be used in
conjunction with other control detection probes against known
sequences which are, for example associated with cancer, or the
characteristic of cancer being investigated (+ve control detection
probes).
[0421] In one embodiment the control detection probes may be a
non-coding RNA, such as a miRNA, siRNA, piRNA or snRNA.
[0422] Suitable target sequences of non coding RNAs may be
identified from the literature, for example the following
publications disclose non coding RNA sequences associates with the
corresponding form of cancer: Breast cancer: Iorio et al Cancer Res
2005; 65: 7065. Lung cancer: Yanaihara et al Cell Science 2006; 9:
189-198. Chronic lymphocytic leukaemia (CLL): Galin et al PNAS,
2004 101(32):11755-11760. Colon cancer: Cummins et al PNAS 2006,
103 (10):3687-3692. Prostate cancer: Volinia et al PNAS 2006; 103:
2257). Cancer: Lu et al (Nature 2005; 435:834-838).
[0423] Preferred +ve control detection probes include detection
probes which specifically hybridise to a noncoding RNA selected
from the group consisting of hsa mIR 21, hsa-Let 7i, hsa miR 101,
hsa miR 145, hsa miR 9, hsa miR122a, hsa miR 128b, hsa miR 149, hsa
miR 125a, hsa miR 143, hsa miR 136, which may be up-regulated in
cancer cells, and hsa-miR 205, which may be down-regulated in
breast cancer cells. The detection probes may be designed against
either the mature miRNA, the pre-miRNA, or both--e.g. may form a
control detection probe pair.
[0424] Detection probes prepared against mRNA and DNA markers
(target nucleic acids) associated with cancer may also be used as
controls, for example markers prepared against the Her-2 gene or
mRNA, which is associated with breast cancer.
Method for the Characterisation of Cancer
[0425] The invention provides a method for the characterisation of
cancer. The data obtained by the method can be used to provide
information on one or more features of cancer.
[0426] The at least one feature of the cancer which is
characterised by the method according to the invention may be
selected from one or more of the following:
[0427] Diagnosis of cancer, the signal data can be used to
determine whether the test sample comprises cells that are
cancerous (i.e. presence or absence of cancer), and/or whether such
cancer is a malignant cancer or a benign cancer.
[0428] The prognosis of the cancer, such as the speed at which the
cancer may develop and or metastasize (i.e. spread from one part of
the body to another) or the life expectancy of the patient with
said cancer (such as less than five years, or greater than five
years). In one embodiment the prognosis may be that the life
expectancy of the patient is less than 5 years, such as less than 4
years, less than 3 years, less than two years, less than 1 year,
less than six months or less than 3 months.
[0429] The origin of said cancer, this may be the cause of the
cancer, or in the case of secondary cancer, the origin of the
primary cancer. The origin may for example be selected from the
following lists of cancer types.
[0430] The type of said cancer, such as a cancer selected from the
group consisting of the following: A solid tumor; ovarian cancer,
breast cancer, non-small cell lung cancer, renal cell cancer,
bladder cancer, oesophagus cancer, stomach cancer, prostate cancer,
pancreatic cancer, lung cancer, cervical cancer, colon cancer,
colorectal cancer, renal cell cancer;
[0431] In one embodiment the type of said cancer is selected from
the group consisting of; breast cancer, adrenal gland cancer,
bladder cancer, colon cancer, Cervical cancer, cancer of the
duodenum, cancer of the esophagus, cancer of the kidney, liver
cancer, lung cancer, ovarian cancer, prostate cancer, rectal
cancer, stomach cancer and uterine cancer.
[0432] The type of cancer may be selected from the group consisting
of: A carcinoma, such as a carcinoma selected from the group
consisting of ovarian carcinoma, breast carcinoma, non-small cell
lung cancer, renal cell carcinoma, bladder carcinoma, recurrent
superficial bladder cancer, stomach carcinoma, prostatic carcinoma,
pancreatic carcinoma, lung carcinoma, cervical carcinoma, cervical
dysplasia, laryngeal papillomatosis, colon carcinoma, colorectal
carcinoma, carcinoid tumors, renal cell carcinoma, A basal cell
carcinoma; A malignant melanoma, such as a malignant melanoma
selected from the group consisting superficial spreading melanoma,
nodular melanoma, lentigo maligna melanoma, acral melagnoma,
amelanotic melanoma and desmoplastic melanoma; A sarcoma, such as a
sarcoma selected from the group consisting of osteosarcoma, Ewing's
sarcoma, chondrosarcoma, malignant fibrous histiocytoma,
fibrosarcoma and Kaposi's sarcoma; and a glioma.
[0433] The use of non-coding RNA markers for determining the origin
of cells is disclosed in U.S. application Ser. No. 11/324,177,
which is hereby incorporated by reference.
[0434] Cancer of unknown primary site is a common clinical entity,
accounting for 2% of all cancer diagnoses in the Surveillance,
Epidemiology, and End Results (SEER) registries between 1973 and
1987 (C. Muir. Cancer of unknown primary site Cancer 1995. 75:
353-356). In spite of the frequency of this syndrome, relatively
little attention has been given to this group of patients, and
systematic study of the entity has lagged behind that of other
areas in oncology. Widespread pessimism concerning the therapy and
prognosis of these patients has been the major reason for the lack
of effort in this area. The patient with carcinoma of unknown
primary site is commonly stereotyped as an elderly, debilitated
individual with metastases at multiple visceral sites. Early
attempts at systemic therapy yielded low response rates and had a
negligible effect on survival, thereby strengthening arguments for
a nihilistic approach to these patients. The heterogeneity of this
group has also made the design of therapeutic studies difficult; it
is well recognized that cancers with different biologies from many
primary sites are represented. In the past 10 years, substantial
improvements have been made in the management and treatment of some
patients with carcinoma of unknown primary site. The identification
of treatable patients within this heterogeneous group has been made
possible by the recognition of several clinical syndromes that
predict chemotherapy responsiveness, and also by the development of
specialized pathologic techniques that can aid in tumor
characterization. Therefore, the optimal management of patients
with cancer of unknown primary site now requires appropriate
clinical and pathologic evaluation to identify treatable subgroups,
followed by the administration of specific therapy. Many patients
with adenocarcinoma of unknown primary site have widespread
metastases and poor performance status at the time of diagnosis.
The outlook for most of these patients is poor, with median
survival of 4 to 6 months. However, subsets of patients with a much
more favorable outlook are contained within this large group, and
optimal initial evaluation enables the identification of these
treatable subsets. In addition, empiric chemotherapy incorporating
newer agents has produced higher response rates and probably
improves the survival of patients with good performance status.
[0435] Fine-needle aspiration biopsy (FNA) provides adequate
amounts of tissue for definitive diagnosis of poorly differentiated
tumors, and identification of the primary source in about one
fourth of cases (C. V. Reyes, K. S. Thompson, J. D. Jensen, and A.
M. Chouelhury. Metastasis of unknown origin: the role of fine
needle aspiration cytology Diagn Cytopathol 1998. 18: 319-322).
[0436] microRNAs have emerged as important non-coding RNAs,
involved in a wide variety of regulatory functions during cell
growth, development and differentiation. Some reports clearly
indicate that microRNA expression may be indicative of cell
differentiation state, which again is an indication of organ or
tissue specification. Therefore a catalogue of mir tissue
expression profiles may serve as the basis for a diagnostic tool
determining the tissue origin of tumors of unknown origin. So,
since it is possible to map miRNA in cells vs. the tissue origin of
cell, the present invention presents a convenient means for
detection of tissue origin of such tumors.
[0437] Hence, the present invention in general relates to a method
for determining tissue origin of tumors comprising probing cells of
the tumor with a collection of probes which is capable of mapping
miRNA to a tissue origin.
[0438] miRNA typing according to the principles of the present
example can be applied to RNA from a variety of normal tissues and
tumor tissues (of known origin) and over time a database is build
up, which consists of miRNA expression profiles from normal and
tumor tissue. When subjecting RNA from a tumor tissue sample, the
resulting miRNA profile can be analysed for its degree of identity
with each of the profiles of the database--the closest matching
profiles are those having the highest likelihood of representing a
tumor having the same origin (but also other characteristics of
clinical significance, such as degree of malignancy, prognosis,
optimum treatment regimen and prediction of treatment success). The
miRNA profile may of course be combined with other tumor origin
determination techniques, cf. e.g. Xiao-Jun Ma et al., Arch Pathol
Lab Med 130, 465-473, which demonstrates molecular classification
of human cancers into 39 tumor classes using a microarray designed
to detect RT-PCR amplified mRNA derived from expression of 92
tumor-related genes. The presently presented technology allows for
an approach which is equivalent safe for the use of a miRNA
detection assay instead of an mRNA detection assay.
[0439] The invention provides a method of characterising a tumor of
unknown origin, such as a metastasis, or putative metastasis,
wherein at least one mature miRNA species is detected in a sample
of RNA from a tumor, (i.e. a first population of target molecules
obtained from at least one test sample) thus providing a miRNA
expression profile from the tumor, and subsequently comparing said
miRNA expression profile with previously established miRNA
expression profiles from normal tissue and/or tumor tissue.
[0440] In one embodiment the tumor may selected from the group
consisting of; breast tumor, adrenal gland tumor, bladder tumor,
colon tumor, Cervical tumor, tumor of the duodenum, tumor of the
esophagus, tumor of the kidney, liver tumor, lung tumor, ovarian
tumor, prostate tumor, rectal tumor, stomach tumor and a uterine
tumor.
[0441] In one embodiment, the tumor may be a breast tumor, or it
may be derived from a breast tumor.
[0442] The RNA may be total RNA isolated from the tumor, or a
purified fraction thereof.
[0443] In one embodiment, the miRNA expression profile from the
tumor and the previously established miRNA expression profiles
provides for an indication of the origin of the tumor, the
patient's prognosis, the optimum treatment regimen of the tumor
and/or a prediction of the outcome of a given anti-tumor
treatment.
[0444] The therapy outcome prediction, such as a prediction of the
responsiveness of the cancer to chemotherapy and/or radiotherapy
and/or the suitability of said cancer to hormone treatment, and
such as the suitability of said cancer for removal by invasive
surgery. In one embodiment, the therapy out come predication may be
the prediction of the suitability of the treatment of the cancer to
combined adjuvant therapy.
[0445] The therapy may be herceptin, which is frequently used for
the treatment of oestrogen receptor positive cancers (such as
breast cancer).
The Patient and Test Sample
[0446] Suitable samples may comprise a wide range of mammalian and
human cells, including protoplasts; or other biological materials,
which may harbour target nucleic acids. The methods are thus
applicable to tissue culture mammalian cells, mammalian cells
(e.g., blood, serum, plasma, reticulocytes, lymphocytes, urine,
bone marrow tissue, cerebrospinal fluid or any product prepared
from blood or lymph) or any type of tissue biopsy (e.g. a muscle
biopsy, a liver biopsy, a kidney biopsy, a bladder biopsy, a bone
biopsy, a cartilage biopsy, a skin biopsy, a pancreas biopsy, a
biopsy of the intestinal tract, a thymus biopsy, a mammae biopsy, a
uterus biopsy, a testicular biopsy, an eye biopsy or a brain
biopsy, e.g., homogenized in lysis buffer), and archival tissue
nucleic acids.
[0447] The test sample is typically obtained from a patient that
has or is suspected of having cancer, such as breast cancer, or who
is suspected of having a high risk of developing cancer. The method
can, therefore be undertaken as a precautionary matter in the
prevention of, or early diagnosis of cancer.
[0448] The patient (or organism) is a mammal, preferably a human
being. The patient may be male or female, although this may depend
on the type of tissue/cancer being investigated (e.g. ovarian
cancer effects only women).
[0449] The test sample is typically obtained from the patient by
biopsy or tissue sampling. When referring to the signal obtained
from a test (or control) sample, it refers to the signal obtained
from the hybridisation using the first (or further) population of
molecules prepared from the test (or control) sample.
The Control Sample
[0450] In one embodiment, the control sample may be obtained from
the same patient at the same time that the test sample is taken. In
one embodiment, the control sample may be a sample taken
previously, e.g. a sample of the same or a different cancer/tumor,
the comparison of which may, for example, provide characterisation
of the source of the new tumor, or progression of the development
of an existing cancer, such as before, during or after
treatment.
[0451] In one embodiment, the control sample may be taken from
healthy tissue, for example tissue taken adjacent to the cancer,
such as within 1 or 2 cm diameter from the external edge of said
cancer. Alternatively the control sample may be taken from an
equivalent position in the patients body, for example in the case
of breast cancer, tissue may be taken from the breast which is not
cancerous.
[0452] In one embodiment, the control sample may also be obtained
from a different patient, e.g. it may be a control sample, or a
collection of control samples, representing different types of
cancer, for example those listed herein (i.e. cancer reference
samples). Comparison of the test sample data with data obtained
from such cancer reference samples may for example allow for the
characterisation of the test cancer to a specific type and/or stage
of cancer.
[0453] In one embodiment, at least one control sample is obtained,
and a second population of nucleic acids from the at least one
control sample is, in addition to the test sample, presented and
hybridised against at least one detection probe.
[0454] The detection probe target for the test and control sample
may be the same, the ratio of the signal obtained between the
control and test sample being indicative of a differential
quantification of the target.
[0455] In one embodiment, the control sample may be obtained from
the same patient as the test sample.
[0456] In one embodiment, the control sample may be obtained from a
non tumorous tissue, such as from tissue adjacent to said putative
tumor, and/or from an equivalent position elsewhere in the
body.
[0457] In one embodiment, the control sample may be obtained from a
non tumorous (or cancerous) tissue selected from the group
consisting; breast tissue, adrenal gland tissue, bladder tissue,
colon tissue, Cervical tissue, tissue of the duodenum, tissue of
the esophagus, tissue of the kidney, liver tissue, lung tissue,
ovarian tissue, prostate tissue, rectal tissue, stomach tissue and
uterine tissue.
[0458] In one embodiment, the control sample (or a further control
sample) may be obtained from a tumorous (or cancerous) tissue
selected from the group consisting; tumorous (or cancerous) breast
tissue, tumorous (or cancerous) adrenal gland tissue, tumorous (or
cancerous) bladder tissue, tumorous (or cancerous) colon tissue,
tumorous (or cancerous) Cervical tissue, tumorous (or cancerous)
tissue of the duodenum, tumorous (or cancerous) tissue of the
esophagus, tumorous (or cancerous) tissue of the kidney, tumorous
(or cancerous) liver tissue, tumorous (or cancerous) lung tissue,
tumorous (or cancerous) ovarian tissue, tumorous (or cancerous)
prostate tissue, tumorous (or cancerous) rectal tissue, tumorous
(or cancerous) stomach tissue and tumorous (or cancerous) uterine
tissue.
[0459] In one embodiment, the control sample may be obtained from a
tumor tissue. In this embodiment, there may be one or more control
samples, e.g. a panel of control samples which represent one or
more tumor types. Thereby allowing comparison of the test sample,
with on or more control samples which have a defined origin. Such
control samples, such as a panel of control samples is particularly
useful when determining the origin of a cancer (e.g. metastasis) of
unknown origin. Such control samples may be selected from one or
more of the following: A solid tumor; ovarian cancer, breast
cancer, non-small cell lung cancer, renal cell cancer, bladder
cancer, esophagus cancer, stomach cancer, prostate cancer,
pancreatic cancer, lung cancer, cervical cancer, colon cancer,
colorectal cancer, renal cell cancer; Such control samples may also
be selected from one or more of the following: The type of cancer
may be selected from the group consisting of: A carcinoma, such as
a carcinoma selected from the group consisting of ovarian
carcinoma, breast carcinoma, non-small cell lung cancer, renal cell
carcinoma, bladder carcinoma, recurrent superficial bladder cancer,
stomach carcinoma, prostatic carcinoma, pancreatic carcinoma, lung
carcinoma, cervical carcinoma, cervical dysplasia, laryngeal
papillomatosis, colon carcinoma, colorectal carcinoma, carcinoid
tumors, renal cell carcinoma, A basal cell carcinoma; A malignant
melanoma, such as a malignant melanoma selected from the group
consisting superficial spreading melanoma, nodular melanoma,
lentigo maligna melanoma, acral melagnoma, amelanotic melanoma and
desmoplastic melanoma; A sarcoma, such as a sarcoma selected from
the group consisting of osteosarcoma, Ewing's sarcoma,
chondrosarcoma, malignant fibrous histiocytoma, fibrosarcoma and
Kaposi's sarcoma; and a glioma.
[0460] Or, such control samples may be selected from one or more of
the following: breast tumor, adrenal gland tumor, bladder tumor,
colon tumor, Cervical tumor, tumor of the duodenum, tumor of the
esophagus, tumor of the kidney, liver tumor, lung tumor, ovarian
tumor, prostate tumor, rectal tumor, stomach tumor and a uterine
tumor.
[0461] In one embodiment, the hybridisation signal obtained from
the test sample is higher than the hybridisation signal obtained
from the control sample.
[0462] In one embodiment, the hybridisation signal obtained from
the test sample is lower than the hybridisation signal obtained
from the control sample.
[0463] In one embodiment, at least two control samples are
obtained, one control sample being obtained from said patient (see
above), and at least one further control sample being obtained from
a previously obtained sample of a cancer, such as a cancer of the
same type as the test sample, or a different cancer such as those
herein listed. The cancer may originate from the same patient or a
different patient.
[0464] In one embodiment, the hybridisation signal obtained from
the at least one further test sample is equivalent to or greater
than the signal obtained from the either the signal obtained from
the first control sample and/or the signal obtained from the test
sample.
[0465] In one embodiment, the hybridisation signal obtained from
the at least one further test sample is less than the signal
obtained from the either the signal obtained from the first control
sample and/or the signal obtained from the test sample.
[0466] In one embodiment, the test and control samples are
hybridised to said at least one detection probe simultaneously,
either in parallel hybridisations or in the same hybridisation
experiment.
[0467] In one embodiment, the test and control sample or samples
are hybridised to said at least one detection probe sequentially,
either in the same hybridisation experiment, or different
hybridisation experiments.
The RNA Fraction
[0468] In one embodiment, the RNA fraction may remain within the
test sample, such as remain in the cells of the biopsy or tissue
sample, for example for in situ hybridisation. The cells may still
be living, or they may be dead. The cells may also be prepared for
in situ hybridisation using methods known in the art, e.g. they may
be treated with an agent to improve permeability of the cells; the
cells may also be fixed or partially fixed.
[0469] The RNA fraction may be isolated from the test sample, such
as a tissue sample.
[0470] The RNA fraction preferably comprises small RNAs such as
those less than 100 bases in length. The RNA fraction preferably
comprises miRNAs.
[0471] In one embodiment the RNA fraction may also comprise other
RNA fractions such as mRNA, and/or in siRNAs and/or piRNAs.
[0472] In one embodiment, the RNA fraction comprises snRNAs.
[0473] The RNA fraction may also comprise other nucleic acids, for
example the RNA fraction may be part of a total nucleic acid
fraction which also comprises DNA, such as genomic and/or
mitochondrial DNA. The RNA fraction may be purified. Care should be
taken during RNA extraction to ensure at least a proportion of the
non encoding RNAs, such as miRNA and siRNAs are retained during the
extraction. Suitably, specific protocols for obtaining RNA
fractions comprising or enriched with small RNAs, such as miRNAs
may be used. The RNA fraction may undergo further purification to
obtain an enriched RNA fraction, for example an RNA fraction
enriched for non-coding RNAs. This can be achieved, for example, by
removing mRNAs by use of affinity purification, e.g. using an
oligodT column. RNA fractions enriched in miRNA and siRNA may be
obtained using. In one embodiment the RNA fraction is not isolated
from the test sample, for example when in situ hybridisation is
performed, the RNA fraction remains in situ in the test sample, and
the detection probes, typically labeled detection probes, are
hybridised to a suitable prepared test sample.
[0474] In one embodiment the RNA fraction is used directly in the
hybridisation with the at least one detection probe.
[0475] The RNA fraction may comprise the target molecule, e.g. the
RNA fraction obtained from a test sample, the presence of the
target molecule within the RNA fraction may indicate a particular
feature of a cancer. Alternatively the RNA fraction may not
comprise the target molecule, e.g. the RNA fraction obtained from a
test sample, the absence of the target (complementary) molecule
within the RNA fraction may indicate a particular feature of a
cancer.
[0476] The RNA fraction comprises non coding RNA such as noncoding
RNA selection from the group consisting of microRNA (miRNA), siRNA,
piRNA and snRNA.
[0477] In one embodiment, prior to (or even during) said
hybridisation, the RNA fraction may be used as a template to
prepare a complement of the RNA present in the fraction, said
compliment may be synthesised by template directed assembly of
nucleoside, nucleotide and/or nucleotide analogue monomers, to
produce, for example an oligonucleotides, such as a DNA
oligonucleotide. The complement may be further copied and
replicated. The compliment may represent the entire template RNA
molecule, or may represent a population of fragments of template
molecules, such as fragments than, preferably in average, retain at
least 8 consecutive nucleoside units of said RNA template, such as
at least 12 of said units or at least 14 of said units. It is
preferred that at least 8 consecutive nucleoside units of said
complementary target, such as at least 12 of said units or at least
14 of said units of said complementary target are retained. When
the complementary target is a precursor RNA, or a molecule derived
therefore, if is preferred that at least part of the loop structure
of the precursor molecule is retained, as this will allow
independent detection over the mature form of the non-coding RNA,
or molecule derived there from.
[0478] Therefore, in one embodiment the RNA fraction itself is not
used in the hybridisation, but a population of molecules, such as
population of oligonucleotides which are derived from said RNA
fraction, and retain sequence information contained within said RNA
fraction, are used. It is envisaged that the population of
molecules derived from said RNA fraction may be further manipulated
or purified prior to the hybridisation step--for example they may
be labeled, or a sub-fraction may be purified there from.
[0479] The target molecule (complementary target) may therefore be
derived from RNA, but may actually comprise an alternative oligo
backbone, for example DNA. The target molecule may, therefore also
be a complement to the original RNA molecule, or part of the
original RNA molecule from which it is derived.
[0480] In one embodiment, the RNA fraction is analysed and the
population of target RNAs and optionally control nucleic acids are
determined. For example the RNA fraction, or a nucleic acid
fraction derived there from may be undergo quantitative analysis
for specific target and control sequences, for example using
oligonucleotide based sequencing, such as oligonucleotide
micro-array hybridization. The data from the quantitative analysis
may then be used in a virtual hybridisation with a detection probe
sequence.
Hybridisation
[0481] Hybridisation refers to the bonding of two complementary
single stranded nucleic acid polymers (such as oligonucleotides),
such as RNA, DNA or polymers comprising or consisting of nucleotide
analogues (such as LNA oligonucleotides). Hybridisation is highly
specific, and may be controlled by regulation of the concentration
of salts and temperature. Hybridisation occurs between
complementary sequences, but may also occur between sequences which
comprise some mismatches. The probes used in the methods of the
present invention may, therefore be 100% complementary to the
target molecule. Alternatively, in one embodiment the detection
probes may comprise one or two mismatches. Typically a single
mismatch will not unduly affect the specificity of binding, however
two or more mismatches per 8 nucleotide/nucleotide residues usually
prevents specific binding of the detection probe to the target
species. The position of the mismatch may also be of importance,
and as such the use of mismatches may be used to determine the
specificity and strength of binding to target RNAs, or to allow
binding to more than one allelic variant of mutation of a target
species.
[0482] In one embodiment, the detection probe consists of no more
than 1 mismatch.
[0483] In one embodiment, the detection probe consists of no more
than 1 mismatch per 8 nucleotide/nucleotide analogue bases.
[0484] In one embodiment, hybridisation may also occur between a
single stranded target molecule, such as a miRNA and a probe which
comprises a complementary surface to the said target molecule, in
this respect, it is the ability of the probe to form the specific
bonding pattern with the target which is important.
[0485] Suitable methods for hybridisation include RNA in-situ
hybridisation, dot blot hybridisation, reverse dot blot
hybridisation, northern blot analysis, RNA protection assays, or
expression profiling by microarrays. Such methods are standard in
the art.
[0486] In one embodiment, the detection probe is capable of binding
to the target non coding RNA sequence under stringent conditions,
or under high stringency conditions.
[0487] Exiqon (Denmark) provide microarrays suitable for use in the
methods of the invention (microRNA Expression Profiling with
miRCURY.TM. LNA Array).
[0488] The detection probe, such as each member of a collection of
detection probes, may be bound (such as conjugated) to a bead.
Luminex (Texas, USA) provides multiplex technology to allow the use
of multiple detection probes to be used in a single hybridisation
experiment. See also Panomics QuantigenePlex.TM..
[0489] Suitable techniques for performing in situ hybridisation are
disclosed in PCT/DK2005/000838
PCR Hybridisation
[0490] Whilst it is recognised that many of the short noncoding
RNAs which are targets for the detection probes are too short to be
detected by amplification by standard PCR, methods of amplifying
such short RNAs are disclosed in WO2005/098029. Therefore, the
hybridisation may occur during PCR, such as RT-PCT or quantitative
PCR (q-PCR).
[0491] However, in one embodiment, the hybridisation step does not
comprise PCR such as RT-PCR or q-pCR.
The Target
[0492] The term the `target` `or complementary target` or `target
nucleic acid` refers to a (typically non-coding) polynucleotide
sequence associated with cancer, preferably an RNA sequence such as
a miRNA, or precursor sequence thereof, or a sequence derived there
from which retains the sequence information present the non-coding
RNA sequence.
[0493] The list of preferred target sequences is provided
herein.
[0494] Preferably the target is a human miRNA or precursor
thereof.
[0495] In one embodiment the target may be one or more of the miRNA
targets i.e. the mRNA targeted by the miRNA (see table 11).
The Signal
[0496] In one embodiment the target is labeled with a signal. In
this respect the population of nucleic acids are labeled with a
signal which can be detected. The hybridisation or the target
molecules to the detection probe, which may be fixed to a solid
surface, and subsequent removal of the remaining nucleic acids from
the population, and therefore allows the determination of the level
of signal from those labeled target which is bound to the detection
probe. This may be appropriate when screening immobilised probes,
such as arrays of detection probes.
[0497] In one embodiment the detection probe is labeled with a
signal. This may be appropriate, for example, when performing in
situ hybridisation and northern blotting, where the population of
nucleic acids are immobilised.
[0498] It is also envisaged that both population of nucleic acids
and detection probes are labeled. For example they may be labeled
with fluorescent probes, such as pairs of FRET probes (Fluorescence
resonance energy transfer), so that when hybridisation occurs, the
FRET pair is formed, which causes a shift in the wavelength of
fluorescent light emitted. It is also envisaged that pairs of
detection probes may be used designed to hybridise to adjacent
regions of the target molecule, and each detection probe carrying
one half of a FRET pair, so that when the probes hybridise to their
respective positions on the target, the FRET pair is formed,
allowing the shift in fluorescence to be detected.
[0499] Therefore, it is also envisaged that neither the population
of nucleic acid molecules or the detection probe need be
immobilised.
[0500] Once the appropriate target RNA sequences have been
selected, probes, such as the preferred LNA substituted detection
probes are preferably chemically synthesized using commercially
available methods and equipment as described in the art
(Tetrahedron 54: 3607-30, 1998). For example, the solid phase
phosphoramidite method can be used to produce short LNA probes
(Caruthers, et al., Cold Spring Harbor Symp. Quant. Biol.
47:411-418, 1982, Adams, et al., J. Am. Chem. Soc. 105: 661
(1983).
[0501] Detection probes, such as LNA-containing-probes, can be
labeled during synthesis. The flexibility of the phosphoramidite
synthesis approach furthermore facilitates the easy production of
detection probes carrying all commercially available linkers,
fluorophores and labelling-molecules available for this standard
chemistry. Detection probes, such as LNA-modified probes, may also
be labeled by enzymatic reactions e.g. by kinasing using T4
polynucleotide kinase and gamma-.sup.32P-ATP or by using terminal
deoxynucleotidyl transferase (TDT) and any given
digoxygenin-conjugated nucleotide triphosphate (dNTP) or
dideoxynucleotide triphosphate (ddNTP).
[0502] Detection probes according to the invention can comprise
single labels or a plurality of labels. In one aspect, the
plurality of labels comprise a pair of labels which interact with
each other either to produce a signal or to produce a change in a
signal when hybridization of the detection probe to a target
sequence occurs.
[0503] In another aspect, the detection probe comprises a
fluorophore moiety and a quencher moiety, positioned in such a way
that the hybridized state of the probe can be distinguished from
the unhybridized state of the probe by an increase in the
fluorescent signal from the nucleotide. In one aspect, the
detection probe comprises, in addition to the recognition element,
first and second complementary sequences, which specifically
hybridize to each other, when the probe is not hybridized to a
recognition sequence in a target molecule, bringing the quencher
molecule in sufficient proximity to said reporter molecule to
quench fluorescence of the reporter molecule. Hybridization of the
target molecule distances the quencher from the reporter molecule
and results in a signal, which is proportional to the amount of
hybridization.
[0504] In the present context, the term "label" means a reporter
group, which is detectable either by itself or as a part of a
detection series. Examples of functional parts of reporter groups
are biotin, digoxigenin, fluorescent groups (groups which are able
to absorb electromagnetic radiation, e.g. light or X-rays, of a
certain wavelength, and which subsequently reemits the energy
absorbed as radiation of longer wavelength; illustrative examples
are DANSYL (5-dimethylamino)-1-naphthalenesulfonyl), DOXYL
(N-oxyl-4,4-dimethyloxazolidine), PROXYL
(N-oxyl-2,2,5,5-tetramethylpyrrolidine), TEMPO
(N-oxyl-2,2,6,6-tetramethylpiperidine), dinitrophenyl, acridines,
coumarins, Cy3 and Cy5 (trademarks for Biological Detection
Systems, Inc.), erythrosine, coumaric acid, umbelliferone, Texas
red, rhodamine, tetramethyl rhodamine, Rox,
7-nitrobenzo-2-oxa-1-diazole (NBD), pyrene, fluorescein, Europium,
Ruthenium, Samarium, and other rare earth metals), radio isotopic
labels, chemiluminescence labels (labels that are detectable via
the emission of light during a chemical reaction), spin labels (a
free radical (e.g. substituted organic nitroxides) or other
paramagnetic probes (e.g. Cu.sup.2+, Mg.sup.2+) bound to a
biological molecule being detectable by the use of electron spin
resonance spectroscopy). Especially interesting examples are
biotin, fluorescein, Texas Red, rhodamine, dinitrophenyl,
digoxigenin, Ruthenium, Europium, Cy5, Cy3, etc.
Control Detection Probes
[0505] It is preferably in the method according to the invention
that in addition to the detection probe for the target in question,
at least one further detection probe is used, where the at least
one further detection probe is capable of hybridising to a control
nucleic acid (control target) present in said population of nucleic
acids (such as the RNA fraction). The control nucleic acid is not
the same as the target in question.
[0506] In one embodiment, the at least one further detection probe
may be derived from or is capable of selectively hybridising with a
molecule selected from the group consisting of: a pre-miRNA
molecule; a pre-siRNA molecule; and a pre-piRNA molecule.
[0507] In another embodiment, the at least one further detection
probe may be derived from or is capable of selectively hybridising
with a molecule selected from the group consisting of a mature
miRNA, a mature siRNA, a mature piRNA and a snRNA.
[0508] A preferred type of detection probe, which may be used with
as a detection probe control and/or as a detection probe, is one
which is capable of hybridising to the loop region of an immature
miRNA, siRNA or piRNA. Recent research has shown that the
processing of pre-microRNAs to mature microRNAs may be controlled
in a cell specific manner (Obernosterer et al). In this respect the
ratio between the immature and mature form can give valuable
information which may be used to characterise the cancer test
sample.
Detection Probes to Precursor Non-Coding RNAs
[0509] The present invention provides for detection probes for the
detection of non coding RNA precursors, such as pre-miRNAs,
pre-siRNAs and pre-piRNAs, and their targets. miRNAs are
transcribed as mono- or poly-cistronic, long, primary precursor
transcripts (pri-miRNAs) that are cleaved into .about.70-nt
precursor hairpins, known as microRNA precursors (pre-miRNAs), by
the nuclear RNase III-like enzyme Drosha (Lee et al., Nature
425:415-419, 2003). MicroRNA precursors (pre-miRNAs) form hairpins
having a loop region and a stem region containing a duplex of the
opposite ends of the RNA strand. Subsequently pre-miRNA hairpins
are exported to the cytoplasm by Exportin-5 (Yi et al., Genes &
Dev., 17:3011-3016, 2003; Bohnsack et al., RNA, 10:185-191, 2004),
where they are processed by a second RNase III-like enzyme, termed
Dicer, into .about.22-nt duplexes (Bernstein et al., Nature
409:363-366, 2001), followed by the asymmetric assembly of one of
the two strands into a functional miRNP or miRISC (Khvorova et al.,
Cell 115:209-216, 2003). miRNAs can recognize regulatory targets
while part of the miRNP complex and inhibit protein translation.
Alternatively, the active RISC complex is guided to degrade the
specific target mRNAs (Lipardi et al., Cell 107:297-307, 2001;
Zhang et al., EMBO J. 21:5875-5885, 2002; Nykanen et al., Cell
107:309-321, 2001). There are several similarities between miRNP
and the RNA-induced silencing complex, RISC, including similar
sizes and both containing RNA helicase and the PPD proteins. It has
therefore been proposed that miRNP and RISC are the same RNP with
multiple functions (Ke et al., Curr. Opin. Chem. Biol. 7:516-523,
2003).
[0510] Most reports in the literature have described the processing
of miRNAs to be complete, suggesting that intermediates like
pri-miRNA and pre-miRNA rarely accumulate in cells and tissues.
However, previous studies describing miRNA profiles of cells and
tissues have only investigated size-fractionated RNAs pools.
Consequently the presence of larger miRNA precursors has been
overlooked.
[0511] Alterations in miRNA biogenesis resulting in different
levels of mature miRNAs and their miRNA precursors could illuminate
the mechanisms underlying many disease processes. For example, the
26 miRNA precursors were equally expressed in non-cancerous and
cancerous colorectal tissues from patients, whereas the expression
of ma-ture human miR143 and miR145 was greatly reduced in cancer
tissues compared with non-cancer tissues, suggesting altered
processing for specific miRNAs in human disease (Michael et al.,
Mol. Cancer Res. 1:882-891, 2003).
[0512] Connections between miRNAs, their precursors, and human
diseases will only strengthen in parallel with the knowledge of
miRNA, their precursors, and the gene networks that they control.
Moreover, the understanding of the regulation of RNA-mediated gene
expression is leading to the development of novel therapeutic
approaches that will be likely to revolutionize the practice of
medicine (Nelson et al., TIBS 28:534-540, 2003).
[0513] siRNAs and piRNAs are considered to undergo a similar
processing from precursor molecules.
[0514] To this end, the invention provides oligonucleotide probes
for precursors of non-coding RNAs, such as miRNA precursors, siRNA
precursors, and piRNA precursors.
[0515] The detection probes for precursors may be a detection probe
that hybridizes to a non-coding RNA precursor molecule, wherein at
least part of said probe hybridizes to a portion of said precursor
not present in the corresponding mature non coding RNA, e.g. the
loop region.
[0516] Such oligonucleotide probes include a sequence complementary
to the desired RNA sequence and preferably a substitution with
nucleotide analogues, preferably high-affinity nucleotide
analogues, e.g., LNA, to increase their sensitivity and specificity
over conventional oligonucleotides, such as DNA oligonucleotides,
for hybridization to the desired RNA sequences.
[0517] An exemplary oligonucleotide probe includes a plurality of
nucleotide analogue monomers and hybridizes to a miRNA precursor.
Desirably, the nucleotide analogue is LNA, wherein the LNA may be
oxy-LNA, preferably beta-D-oxy-LNA, monomers. Desirably, the
oligonucleotide probe will hybridize to part of the loop sequence
of a miRNA precursor, e.g., to 5 nucleotides of the miRNA precursor
loop sequence or to the center of the miRNA precursor loop
sequence. In other embodiments, the oligonucleotide probe will
hybridize to part of the stem sequence of a miRNA precursor.
[0518] The invention also features a method of measuring relative
amounts of non coding RNAs, such as miRNa, piRNA and siRNA, and
their precursors, such as pre-miRNAs, pre-siRNAs and
pre-piRNAs.
[0519] This may be achieved by using a detection probe pair which
comprises of i) a first detection probe that hybridizes to a
non-coding RNA precursor molecule, wherein at least part of said
probe hybridizes to a portion of said precursor not present in the
corresponding mature non-coding RNA, and ii) a further detection
probe that hybridizes to the mature non-coding RNA, but will not
hybridise to the precursor non-coding RNA, e.g. by designing the
detection probe to hybridise to the sequence which flanks the stem
loop splice site of the precursor molecule. The ratio of signal of
hybridisation thereby provides data which can provide said
characterisation of said breast cancer.
[0520] Therefore, the invention further provides for a detection
probe pair which consist of a detection probe which specifically
hybridises to the pre-miRNA sequence and a further detection probe
which specifically hybridises to the mature miRNA sequence. By
designing, for example the pre-miRNA detection probe across the
stem loop region, which is cleaved during the maturation process,
the detection probe which specifically hybridises to the pre-mature
miRNA does not specifically hybridise to the mature miRNA sequence.
Therefore in a preferred embodiment, by comparing the signal
obtained from each member of the detection probe pair it is
possible to determine the comparative population of pre-miRNAs as
compared to the corresponding mature miRNAs in a sample.
[0521] In one embodiment, the comparison is made by contacting a
first probe that hybridizes to the mature noncoding RNA, such as
mature miRNA, with the sample under a first condition that also
allows the corresponding non-coding RNA precursor, such as miRNA
precursor to hybridize; contacting the first probe or a second
probe that hybridizes to mature non-coding RNA with the sample
under a second condition that does not allow corresponding miRNA
precursor to hybridize; comparing the amounts of the probes
hybridized under the two conditions wherein the reduction in amount
hybridized under the second condition compared to the first
condition is indicative of the amount of the miRNA precursor in the
sample.
[0522] Furthermore, the invention features a kit including a probe
of the invention (or a detection probe pair according to the
invention) and packaging and/or labeling indicative of the
non-coding RNA and/or non-coding precursor (e.g. miRNA precursor),
to which the probe (or probe pair) hybridizes and conditions under
which the hybridization occurs. The kit provides for the isolation,
purification, amplification, detection, identification,
quantification, or capture of natural or synthetic nucleic acids.
The probes are preferably immobilized onto a solid support, e.g.,
such as a bead or an array.
[0523] The invention also features a method of treating a disease
or condition in a living organism using any combination of the
probes and methods of the invention.
[0524] The invention further features a method of comparing
relative amounts of miRNA and miRNA precursor in a sample by
contacting the sample with a first probe that hybridizes to miRNA
precursor and a second probe that hybridizes to miRNA; and
detecting the amount of one or more signals indicative of the
relative amounts of miRNA and miRNA precursor.
[0525] The invention also features a method of measuring relative
amounts of miRNA and miRNA precursor in a sample by contacting a
first probe that hybridizes to miRNA with the sample under
conditions that also allow miRNA precursor to hybridize; contacting
the first probe or a second probe that hybridizes to miRNA with the
sample under conditions that do not allow miRNA precursor to
hybridize; comparing the amounts of the probes hybridized under the
two conditions wherein the reduction in amount hybridized under the
second condition compared to the first condition is indicative of
the amount of miRNA precursor in the sample.
[0526] The invention also features methods of using the probes of
the invention as components of Northern blots, in situ
hybridization, arrays, and various forms of PCR analysis including
PCR, RT-PCR, and qPCR.
[0527] Any probe of the invention may be used in performing any
method of the inven-ion. For example, any method of the invention
may involve probes having labels. Furthermore, any method of the
invention may also involve contacting a probe with miRNA precursor
that is endogenously or exogenously produced. Such contacting may
occur in vitro or in vivo, e.g., such as in the body of an animal,
or within or without a cell, which may or may not naturally express
the miRNA precursor.
[0528] Also, primarily with respect to miRNA precursors, nucleotide
analogue containing probes, polynucleotides, and oligonucleotides
are broadly applicable to antisense uses. To this end, the present
invention provides a method for detection and functional analysis
of non-coding antisense RNAs, as well as a method for detecting the
overlapping regions between sense-antisense transcriptional
units.
[0529] The oligonucleotide probes of invention are also useful for
detecting, testing, diagnosing or quantifying miRNA precursors and
their targets implicated in or connected to human disease, e.g.,
analyzing human samples for cancer diagnosis.
[0530] For example, pre-mir-138-2 is ubiquitously expressed, unlike
its mature miRNA derivative. The presence of an unprocessed miRNA
precursor in most tissues of the organism suggests miRNA precursors
as possible diagnostic targets. We envision that miRNA precursor
processing could be a more general feature of the regulation of
miRNA expression and be used to identify underlying disease
processes. One could also imagine that the unprocessed miRNA
precursors might play a different role in the cell, irrespective of
the function of the mature miRNA, providing further insights into
underlying disease processes.
[0531] Imperfect processing of miRNA precursors to mature miRNA as
detected by sample hybridization to oligonucleotide probes may
provide diagnostic or prognostic in-formation. Specifically, the
ratio between levels of mature and precursor transcripts of a given
miRNA may hold prognostic or diagnostic information. Furthermore,
specific spatial expression patterns of mature miRNA compared to
miRNA precursor may likewise hold prognostic or diagnostic
information. In addition, performing in situ hybridization using
mature miRNA and/or miRNA precursor specific oligonucleotide probes
could also detect abnormal expression levels. LNA-containing probes
are particularly well-suited for these purposes.
[0532] The present invention enables discrimination between
different polynucleotide transcripts and detects each variant in a
nucleic acid sample, such as a sample derived from a patient, e.g.,
addressing the spatiotemporal expression patterns by RNA in situ
hybridization. The methods are thus applicable to tissue culture
animal cells, animal cells (e.g., blood, serum, plasma,
reticulocytes, lymphocytes, urine, bone marrow tissue,
cerebrospinal fluid or any product prepared from blood or lymph) or
any type of tissue biopsy (e.g., a muscle biopsy, a liver biopsy, a
kidney biopsy, a bladder biopsy, a bone biopsy, a cartilage biopsy,
a skin biopsy, a pancreas biopsy, a biopsy of the intestinal tract,
a thymus biopsy, a mammae biopsy, a uterus biopsy, a testicular
biopsy, an eye biopsy or a brain biopsy, e.g., homogenized in lysis
buffer), archival tissue nucleic acids such as formalin fixated
paraffin embedded sections of the tissue and the like.
[0533] pre-mir-138-1 and pre-mir-138-2 and their shared mature
miRNA derivative mir-138 differ in their expression levels across
various tissues as detected by oligonucleotide probes. The
differential expression of pre-mir-138-1 and pre-mir-138-2 and
their derived mature miRNA mir-138. pre-mir-138-2 is expressed in
all tissues, and mir-138 is expressed in a tissue-specific manner.
Furthermore, the experiments suggest that an inhibitory factor is
responsible for tissue-specific processing of pre-mir-138-2 into
mir-138 and that this inhibitory factor is specific for certain
miRNA precursors. This inhibitory factor acting on pre-138-2 may be
capable of distinguishing pre-mir-138-1 from pre-mir-138-2 as well.
pre-mir-138-1 and pre-mir-138-2 have different pre-mir sequences,
particularly in the loop region, and thus the inhibitory factor may
be capable of recognizing these sequence differences to achieve
such specificity. It is hypothesized that recognition by an
inhibitory factor is dependent on the differences in the loop
sequence, e.g., the size of the loop sequence, between
pre-mir-138-1 and pre-mir-138-2. It is therefore possible that an
oligonucleotide probe capable of hybridizing specifically to the
sequences that are different between pre-mir-138-1 and
pre-mir-138-2, e.g. in the loop region, could be utilized to block
the inhibitory effect of the inhibitory factor, thereby allowing
the pre-mir-138-2 to be processed.
Signal Data
[0534] The signal data obtained from the hybridisation experiment
may be a quantitative measurement of the level of signal
detected.
[0535] The signal data obtained from the hybridisation experiment
may be a qualitative measurement of the level of signal
detected.
[0536] For example, in the case of non-coding RNAs whose presence
of absence is indicative of the presence/or absence of a feature of
the cancer, the detection of signal, i.e. positive signal data or
negative signal data may be a direct indication of the feature in
question.
[0537] In one embodiment the signal data may be used to obtain a
ratio of the signals obtained between the test sample and a control
sample, or a matrix between the signal between the control sample
and more than one of the controls as herein provided. The ratio or
matrix being indicative of the feature in question.
[0538] The signal data from numerous hybridisations, for example
arrays of a collection of detection probes may provide signals from
hybridisations with several different targets, and it is the
differential pattern of targets which allows for one or more of the
features in question to be determined. Typically, the determination
of previously characterised cancers can provide a dataset which can
subsequently be used for comparison with data obtained from samples
from a patient, thereby allowing determination of the features.
[0539] Therefore, in one embodiment, the method of the invention
comprises the hybridisation of the test sample and one or more
control samples to both i) one or more target detection probes,
such as a collection of detection probes, which may be in the form
as listed above, such as an array such as a micro-array, and ii)
one or more control detection probes, such as [0540] at least one
normalising control probe and at least one mRNA marker control
probe, or [0541] at least one normalising control probe and at
least one DNA marker control probe and optionally at least one mRNA
marker control probe. or [0542] at least one normalising control
probe and at least one immature noncoding RNA, selected from
immature miRNA, immature siRNA and immature piRNA, and optionally
at least one DNA marker control probe and optionally at least one
mRNA marker control probe.
Collection of Probes of the Invention
[0543] In one embodiment a collection of probes according to the
present invention comprises at least 10 detection probes, 15
detection probes, such as at least 20, at least 25, at least 50, at
least 75, at least 100, at least 200, at least 500, at least 1000,
and at least 2000 members.
[0544] The collection of detection probes may comprise a majority
of detection probes to the target as compared to the control
probes.
[0545] In one embodiment, at least 10%, such as at least 20%, such
as at least 30%, such as at least 40%, such as at least 50%, such
at least 60%, such as at least 70%, such as at least 80%, such as
at least 90 or 95% of the detection probes in the collection of
detection probes may be capable of hybridizing to the respective
population of target molecules (as opposed to control-targets).
[0546] The collection of detection probes preferably comprises at
least one control detection probe, and may comprise a collection of
control detection probes.
[0547] In one embodiment, the collection of probes according to the
present invention consists of no more than 500 detection probes,
such as no more than 200 detection probes, such as no more than 100
detection probes, such as no more than 75 detection probes, such as
no more than 50 detection probes, such as no more that 50 detection
probes, such as no more than 25 detection probes, such as no more
than 20 detection probes.
[0548] In one embodiment, the collection of probes according to the
present invention has between 3 and 100 detection probes, such as
between 5 and 50 detection probes, such as between 10 and 25
detection probes.
[0549] In one embodiment, the collection of probes of the invention
is capable of specifically detecting all or substantially all
members of the transcriptome of an organism.
[0550] In another embodiment, the collection of probes is capable
of specifically detecting all small non-coding RNAs of an organism,
such as all miRNAs, piRNAs, snRNAs and/or siRNAs.
[0551] In a preferred embodiment, the collection of probes is
capable of specifically detecting a subset of non-coding RNAs,
preferably a subset which has been selected for their ability to
act as markers for at least one type of cancer, and preferably
appropriate control probes or collection of control probes.
[0552] In one embodiment, the affinity-enhancing nucleobase
analogues are regularly spaced between the nucleobases in at least
80% of the members of said collection, such as in at least 90% or
at least 95% of said collection (in one embodiment, all members of
the collection contains regularly spaced affinity-enhancing
nucleobase analogues).
[0553] In one embodiment of the collection of probes, all members
contain affinity enhancing nucleobase analogues with the same
regular spacing in the recognition sequences.
[0554] Also for production purposes, it is an advantage that a
majority of the probes in a collection are of the same length. In
preferred embodiments, the collection of probes of the invention is
one wherein at least 80% of the members comprise recognition
sequences of the same length, such as at least 90% or at least
95%.
[0555] As discussed above, it is advantageous, in order to avoid
self-annealing, that at least one of the nucleobases in the
recognition sequence is substituted with its corresponding
selectively binding complementary (SBC) nucleobase.
[0556] Typically, the nucleobases in the sequence are selected from
ribonucleotides and deoxyribonucleotides, preferably
deoxyribonucleotides. It is preferred that the recognition sequence
consists of affinity enhancing nucleobase analogues together with
either ribonucleotides or deoxyribonucleotides.
[0557] In certain embodiments, each member of a collection is
covalently bonded to a solid support. Such a solid support may be
selected from a bead, a microarray, a chip, a strip, a
chromatographic matrix, a microtiter plate, a fiber or any other
convenient solid support generally accepted in the art.
[0558] The collection may be so constituted that at least 90% (such
as at least 95%) of the recognition sequences exhibit a melting
temperature or a measure of melting temperature corresponding to at
least 5.degree. C. higher than a melting temperature or a measure
of melting temperature of the self-complementarity score under
conditions where the probe hybridizes specifically to its
complementary target sequence (or that at least the same
percentages of probes exhibit a melting temperature of the
probe-target duplex of at least 5.degree. C. more than the melting
temperature of duplexes between the probes or the probes
internally).
[0559] As also detailed herein, each detection probe in a
collection of the invention may include a detection moiety and/or a
ligand, optionally placed in the recognition sequence but also
placed outside the recognition sequence. The detection probe may
thus include a photochemically active group, a thermochemically
active group, a chelating group, a reporter group, or a ligand that
facilitates the direct of indirect detection of the probe or the
immobilisation of the oligonucleotide probe onto a solid
support.
Methods/Uses of Probes and Probe Collections
[0560] Preferred methods/uses include: Specific isolation,
purification, amplification, detection, identification,
quantification, inhibition or capture of a target nucleotide
sequence in a sample, wherein said target nucleotide sequence is
associated with cancer, such as breast cancer, by contacting said
sample with a member of a collection of probes or a probe defined
herein under conditions that facilitate hybridization between said
member/probe and said target nucleotide sequence. Since the probes
are typically shorter than the complete molecule wherein they form
part, the inventive methods/uses include isolation, purification,
amplification, detection, identification, quantification,
inhibition or capture of a molecule comprising the target
nucleotide sequence.
[0561] Typically, the molecule which is isolated, purified,
amplified, detected, identified, quantified, inhibited or captured
is a small, non-coding RNA, e.g. a miRNA such as a mature miRNA.
Typically the small, non-coding RNA has a length of at most 30
residues, such as at most 29, 28, 27, 26, 25, 24, 23, 22, 21, 20,
19, or 18 residues. The small non-coding RNA typically also has a
length of at least 15 residues, such as at least 16, 17, 18, 19,
20, 21, 22, 23, 24, 25, 26, 27, 28, 29 or 30 residues.
[0562] As detailed in PCT/DK2005/000838, the specific hybridization
between the short probes of the present invention to miRNA and the
fact that miRNA can be mapped to various tissue origins, allows for
an embodiment of the uses/methods of the present invention
comprising identification of the primary site of metastatic tumors
of unknown origin.
[0563] As also detailed in PCT/DK2005/000838, the short, but highly
specific probes of the present invention allows hybridization
assays to be performed on fixated embedded tissue sections, such as
formalin fixated paraffin embedded sections. Hence, an embodiment
of the uses/methods of the present invention are those where the
molecule, which is isolated, purified, amplified, detected,
identified, quantified, inhibited or captured, is DNA (single
stranded such as viral DNA) or RNA present in a fixated, embedded
sample such as a formalin fixated paraffin embedded sample.
[0564] The detection probes herein disclosed may also be used for
detection and assessment of expression patterns for naturally
occurring single stranded nucleic acids such as miRNAs, their
target mRNAs, stem-loop precursor miRNAs, siRNAs, piRNAs, other
non-coding RNAs, RNA-edited transcripts or alternative mRNA splice
variants by RNA in-situ hybridisation, dot blot hybridisation,
reverse dot blot hybridisation, or in Northern blot analysis or
expression profiling by microarrays.
[0565] In one embodiment the hybridisation occurs in in situ
hybridisation of a test sample, such as a biopsy, taken from a
patient during an operation. The use of in situ hybridisation is
preferred when the two dimensional location of the target molecule
is to be used in determining the feature of the cancer. For
example, cancers are often made up of vascular cells, connective
tissue etc as well as cancerous cells, the use of in situ
hybridisation therefore allows a morphological distinction to be
made between hybridisation in non cancer cells and cancer cells
within a sample. Typically the in situ hybridisation is performed
using only a few detection probes, such as between 1 and three
detection probes, such as two detection probes. One or two of the
detection probes may be control probes. The in situ hybridisation
may be performed during or subsequent to a method of therapy such
as surgery for removal or biopsy of a cancer.
[0566] The detection probes herein disclosed may also be used for
antisense-based intervention, targeted against tumorgenic single
stranded nucleic acids such as miRNAs, their target mRNAs,
stem-loop precursor miRNAs, siRNAs, piRNAs, other non-coding RNAs,
RNA-edited transcripts or alternative mRNA splice variants or viral
DNA in vivo in plants or animals, such as human, mouse, rat, by
inhibiting their mode of action, e.g. the binding of mature miRNAs
to their cognate target mRNAs.
[0567] Further embodiments includes the use of the detection probe
as an aptamer in molecular diagnostics or (b) as an aptamer in RNA
mediated catalytic processes or (c) as an aptamer in specific
binding of antibiotics, drugs, amino acids, peptides, structural
proteins, protein receptors, protein enzymes, saccharides,
polysaccharides, biological cofactors, nucleic acids, or
triphosphates or (d) as an aptamer in the separation of enantiomers
from racemic mixtures by stereospecific binding or (e) for
labelling cells or (f) to hybridise to non-protein coding cellular
RNAs, miRNA (preferably) such as tRNA, rRNA, snRNA and scRNA, in
vivo or in-vitro or (g) to hybridise to non-protein coding cellular
RNAs, such as miRNA (preferably) tRNA, rRNA, snRNA and scRNA, in
vivo or in-vitro or (h) in the construction of Taqman probes or
Molecular Beacons.
[0568] The present invention also provides a kit for the isolation,
purification, amplification, detection, identification,
quantification, or capture of nucleic acids, wherein said nucleic
acids are associated with cancer, such as the cancers herein
disclosed, such as breast cancer, where the kit comprises a
reaction body and one or more LNAs as defined herein. The LNAs are
preferably immobilised onto said reactions body (e.g. by using the
immobilising techniques described above).
[0569] For the kits according to the invention, the reaction body
is preferably a solid support material, e.g. selected from
borosilicate glass, soda-lime glass, polystyrene, polycarbonate,
polypropylene, polyethylene, polyethyleneglycol terephthalate,
polyvinylacetate, polyvinylpyrrolidinone, polymethylmethacrylate
and polyvinylchloride, preferably polystyrene and polycarbonate.
The reaction body may be in the form of a specimen tube, a vial, a
slide, a sheet, a film, a bead, a pellet, a disc, a plate, a ring,
a rod, a net, a filter, a tray, a microtitre plate, a stick, or a
multi-bladed stick.
[0570] A written instruction sheet stating the optimal conditions
for use of the kit typically accompanies the kits.
[0571] A preferred embodiment of the invention is a kit for the
characterisation of cancer, such as the cancers listed herein. Such
kits may allow the detection or quantification of target non-coding
RNAs, such as miRNA (preferably), siRNAs, snRNAs, piRNAs,
non-coding antisense transcripts or alternative splice
variants.
[0572] The kit may comprise libraries of detection probes, which
comprise one or more detection probes and optionally one or more
control probes. The kit may also comprise detection probes for
mRNAs (i.e. coding RNAs), and DNA, the presence or absence or level
of which may also contribute to characterising the cancer. It is
preferable that the kit comprises an array comprising a collection
of detection probes, such as an oligonucleotide arrays or
microarray.
[0573] The use of the kit therefore allows detection of non-coding
RNAs which are associated with cancer, and whose level or presence
or absence, may, either alone, or in conjunction with the level or
presence or absence of other non-coding RNAs, and optionally coding
RNAs, provide signal data which can be used to characterise said
cancer.
[0574] In one aspect, the kit comprises in silico protocols for
their use. The detection probes contained within these kits may
have any or all of the characteristics described above. In one
preferred aspect, a plurality of probes comprises at least one
stabilizing nucleotide, such as an LNA nucleotide. In another
aspect, the plurality of probes comprises a nucleotide coupled to
or stably associated with at least one chemical moiety for
increasing the stability of binding of the probe.
[0575] The invention therefore also provides for an array, such as
a microarray which comprises one or more detection probe according
to the invention, such as the collection of detection probes and
optionally one or more control probe, preferably a collection of
control probes. The array or microarray is particularly preferred
for use in the method of the invention.
BRIEF DESCRIPTION OF THE FIGURES
[0576] FIG. 1: Separation of small RNAs from human breast cancer
tissues on a denaturing 12.5% polyacrylamide gel. miRNAs in the
size range of 15-30 nt and 30-100 nt were extracted from the
gel.
[0577] FIG. 2: PAGE analysis (6% PAA) of PCR-amplified cDNAs
obtained from the 15-30 nt (lane 1) and the 30-100 nt (lane 2)
miRNA fraction (100 ng each). The gel was stained with
ethidiumbromide.
[0578] FIG. 3: PAA gel (6%) electrophoresis of the gel purified
cDNA fractions containing miRNA cDNA inserts of 15-30 bp (lane 1,
20 ng) and 30-100 bp (lane 2, 40 ng) and the resulting cDNA pool
(lane 3, 33 ng). The gel was stained with ethidiumbromide.
[0579] FIG. 4: M-A plot showing all miRNA signals before
averaging.
EXAMPLES
[0580] The invention will now be further illustrated with reference
to the following examples. It will be appreciated that what follows
is by way of example only and that modifications to detail may be
made while still falling within the scope of the invention.
[0581] LNA-substituted probes may be prepared according to Example
1 of PCT/DK2005/000838.
Example 1
Identification of Novel miRNAs by 454 Pyrosequencing
[0582] Experimental Methods--Preparation of a cDNA-Pool from Human
Breast Cancer Tissues for 454 Amplicon Sequencing
[0583] Tissue: Five different human breast cancer tissue samples
(about 200 mg in total) were shipped in RNA later on dry ice to
Vertis Biotechnologie AG for RNA purification and
fractionation.
[0584] Preparation and gel purification of miRNAs: Tissues were
ground under liquid nitrogen. RNA species smaller than 200 nt were
enriched with the mirVana miRNA isolation kit (Ambion, Austin,
Tex., USA). The small RNAs were then separated on a denaturing
12.5% polyacrylamide (PAA) gel. As molecular mass standard a
mixture of oligonucleotides was used that range in size between 15
and 100 bases (see FIG. 1). The population of miRNAs with a length
of 15-30 or 30-100 bases was obtained by passive elution of the
RNAs from the gel. The miRNAs were then precipitated with ethanol
and dissolved in water.
[0585] cDNA synthesis: For cDNA synthesis the miRNAs were first
poly(A)-tailed using poly(A) polymerase followed by ligation of a
RNA adapter to the 5''-phosphate of the miRNAs. First-strand cDNA
synthesis was then performed using an oligo(dT)-linker primer and
M-MLV-RNase H-reverse transcriptase. The resulting cDNAs were then
PCR-amplified to about 20 ng/.mu.l in cycle numbers indicated in
Table 1 using Taq polymerase.
TABLE-US-00001 TABLE 1 Number of PCR cycles used for cDNA
amplification, barcode sequences and size fraction of miRNA used
for cDNA synthesis. 5'-TAG Number PCR cycles sequence RNA size cDNA
size 1 20 ATCG 15-30 nt 119-134 bp 2 18 CAGC 30-100 nt 135-205
bp
[0586] The fusion primers used for PCR amplification were designed
for amplicon sequencing according to the instructions of 454 Live
Sciences. Barcode sequences for each cDNA species, which are
attached to the 5'-ends of the cDNAs, are included in Table 1. The
following adapter sequences flank the cDNA inserts:
TABLE-US-00002 5'-end (43 bases): (SEQ ID NO 409)
5'-GCCTCCCTCGCGCCATCAGCTNNNNGACCTTGGCTGTCACTCA-3'. 3'-end (61
bases): (SEQ ID NO 410)
5'-GCCTTGCCAGCCCGCTCAGACGAGACATCGCCCCGCTTTTTTTTTTT
TTTTTTTTTTTTTT-3.
[0587] 454 adapter sequences are underlined.
[0588] The combined length of the flanking sequences is 104 bases.
Therefore, PCR-products containing miRNA cDNAs of 15-30 nt and
30-100 nt must have a total length of 119-134 bp and 135-205 bp.
PAGE analysis of the 2 cDNAs revealed that the cDNAs were of the
expected size (FIG. 2).
[0589] Pool formation: The correct size ranges of 119-134 bp and
135-205 bp of both cDNAs were obtained by separate purification on
6% PAA-gels. For pool formation the purified cDNAs shown in lane 1
(15-30 bp inserts) and 2 (30-100 bp inserts) of FIG. 3 were mixed
in a molar ratio of 3+1. The concentration of the cDNA pool is 11
ng/.mu.l dissolved in 25 .mu.l water. 3 .mu.l of the gel purified
pool was analyzed on a 6% PAA gel (FIG. 3, lane 3).
Example 2
Molecular Classification of Breast Cancer by microRNA
Signatures
[0590] Breast cancer is the most frequent form of cancer among
women worldwide. Currently, treatment and prognosis is based on
clinical and histo-pathological graduation, such as TNM
classification (tumor size, lymph node and distant metastases
status) and estrogen receptor status. To improve both the selection
of therapy and the evaluation of treatment response, more accurate
determinants for prognosis and response, such as molecular tumor
markers, are needed. The primary aim of this study was to study the
expression patterns of microRNAs (miRNAs) in tumors and normal
breast tissue to identify new molecular markers of breast
cancer.
[0591] Biopsies from primary tumors and from the proximal tissue (1
cm from the border zone of tumor) were collected from female
patients (age 55-69) undergoing surgery for invasive ductal
carcinoma. Total-RNA was extracted following the "Fast RNA GREEN"
protocol from Bio101. Assessment of miRNA levels was carried out on
miRCURY.TM. microarrays according to the manufacturers recommended
protocol (Exiqon, Denmark).
[0592] The results from the miRNA analysis revealed numerous
differentially expressed miRNAs, including those reported earlier
to be associated with breast cancer, such as let-7a/d/f,
miR-125a/b, miR-21, miR-32, and miR-136 [1]. In addition, we have
identified several miRNAs that have not previously been connected
with breast cancer.
RNA Extraction
[0593] Before use, all samples were kept at -80.degree. C.
[0594] Two samples--ca. 100 mg of each--were used for RNA
extraction:
PT (primary tumor) 1C (normal adjacent tissue, one cm from the
primary tumor)
[0595] The samples were thawed on ice, and kept in RNAlater.RTM.
(Cat#7020, Ambion) during disruption with a sterile scalpel into
smaller ca. 1 mm wide slices.
[0596] To a FastPrep GREEN (Cat#6040-600, Bio101) tube containing
lysis matrix was added:
500 .mu.L CRSR-GREEN
500 .mu.L PAR
100 .mu.L CIA
[0597] 200 .mu.L tissue
[0598] The tubes were placed in the FastPrep FP120 cell disruptor
(Bio101) and run for 40 seconds at speed 6. This procedure was
repeated twice, before cooling on ice for 5 min. The tubes were
centrifuged at 4.degree. C. and at maximum speed in an Eppendorf
microcentrifuge for 10 min to enable separation into organic and
water phases. The upper phase from each vial was transferred to new
Eppendorf 1.5 mL tubes while avoiding the interphase. 500 .mu.l CIA
was added, vortexed for 10 seconds, and spun at max speed for 2 min
to separate the phases. Again, the top phase was transferred to new
Eppendorf tubes, while the interphase was untouched. 500 .mu.L DIPS
was added, vortexed, and incubated at room temperature for 2 min.
The tubes were centrifuged for 5 min at max speed to pellet the
RNA. The pellet was washed twice with 250 .mu.L SEWS and left at
room temperature for 10 min to air dry. 50 .mu.L SAFE was added to
dissolve the pellet, which was stored at -80.degree. C. until use.
QC of the RNA was performed with the Agilent 2100 BioAnalyser using
the Agilent RNA6000 Nano kit. RNA concentrations were measured in a
Nanoprop ND-1000 spectrophotometer. The PT was only 71 ng/.mu.L, so
it was concentrated in a speedvac for 15 min to 342 ng/.mu.L. The
1C was 230 ng/.mu.L, and was used as is.
RNA Labelling and Hybridization
[0599] Essentially, the instructions detailed in the "miRCURY Array
labelling kit Instruction Manual" were followed:
[0600] All kit reagents were thawed on ice for 15 min, vortexed and
spun down for 10 min. In a 0.6 mL Eppendorf tune, the following
reagents were added:
2.5.times. labelling buffer, 8 .mu.L Fluorescent label, 2 .mu.L 1
.mu.g total-RNA (2.92 .mu.L (PT) and 4.35 .mu.L (1C)) Labeling
enzyme, 2 .mu.L Nuclease-free water to 20 .mu.L (5.08 .mu.L (PT)
and 3.65 .mu.L (1C))
[0601] Each microcentrifuge tube was vortexed and spun for 10
min.
[0602] Incubation at 0.degree. C. for 1 hour was followed by 15 min
at 65.degree. C., then the samples were kept on ice.
[0603] For hybridization, the 12-chamber TECAN HS4800Pro
hybridization station was used.
[0604] 25 .mu.L 2.times. hybridization buffer was added to each
sample, vortexed and spun.
[0605] Incubation at 95.degree. C. for 3 min was followed by
centrifugation for 2 min.
[0606] The hybridization chambers were primed with 1.times.Hyb
buffer.
[0607] 50 .mu.l of the target preparation was injected into the Hyb
station and incubated at 60.degree. C. for 16 hours
(overnight).
[0608] The slides were washed at 60.degree. C. for 1 min with
Buffer A twice, at 23.degree. C. for 1 min with Buffer B twice, at
23.degree. C. for 1 min with Buffer C twice, at 23.degree. C. for
30 sec with Buffer C once.
[0609] The slides were dried for 5 min.
[0610] Scanning was performed in a ScanArray 4000XL (Packard
Bioscience).
Results
[0611] The M-A plot (FIG. 4) shows the Log 2 fold ratio of
tumor/normal (M) as a function of the Log 2
[0612] In this experiment, a total of 86 out of known 398 miRNAs
were found to be differentially expressed between breast cancer and
normal adjacent tissue.
Example 3
LNA-Substituted Detection Probes for Detection of microRNAs
Associated with Breast Cancer in Humans
[0613] LNA nucleotides are depicted by capital letters, DNA
nucleotides by lowercase letters, mC denotes LNA methyl-cytosine.
The detection probes can be used to detect and analyze conserved
vertebrate miRNAs, such as human miRNAs by RNA in situ
hybridization, Northern blot analysis and by silencing using the
probes as miRNA inhibitors. The LNA-modified probes can be
conjugated with a variety of haptens or fluorochromes for miRNA in
situ hybridization using standard methods. 5'-end labeling using T4
polynucleotide kinase and gamma-32P-ATP can be carried out by
standard methods for Northern blot analysis. In addition, the
LNA-modified probe sequences can be used as capture sequences for
expression profiling by LNA oligonucleotide microarrays. Covalent
attachment to the solid surfaces of the capture probes can be
accomplished by incorporating a NH.sub.2--C.sub.6-- or a
NH.sub.2--C.sub.6-hexaethylene glycol monomer or dimer group at the
5'-end or at the 3'-end of the probes during synthesis. As
disclosed in PCT/DK2005/000838,
[0614] It is possible to map miRNA in cells to determine the tissue
origin of these cells, the present invention presents a convenient
means for detection of tissue origin of tumors.
[0615] Hence, the present invention in general relates to a method
for determining tissue origin of breast tumors comprising probing
cells of the tumor with a collection of probes which is capable of
mapping miRNA to a tissue origin.
Example 4
Identification of Further Novel miRNAs
[0616] Further microRNAs and pre-miRNAs were identified using
similar experimental techniques as Examples 1-3. These are
microRNAs and pre-miRNAs are shown in columns 1 and 2 of table 3,
and in some cases they were found to be related to the mircoRNAs
(and pre-miRNAs) identified from the previous examples (as shown in
columns 3 and 4 of table 3). The pre-miRNA sequences and their
corresponding SEQ ID number, pre-miR ID number, are provided in
table 4. The corresponding miRNA sequences and their corresponding
SEQ ID number are provided in table 5.
TABLE-US-00003 TABLE 3 miRNA Pre-miRNA Related Related SEQ ID SEQ
ID miRNAs pre-miRNAs Column 1 Column 2 SEQ IDs SEQ IDs 411 412 413
414 504 505 415 416 417 418 419 147 148 420 279 280 421 447 448 422
423 424 425 426 427 480 481 428 429 433 452 453 430 431 432 434 435
502 503 436 437 438 439 440 441 442 443 444 445 446 447 448 147 148
279 280 418 418 420 421 449 450 513 514 451 452 453 429 433 430 431
432 454 455 456 457 458 459 460 461 462 463 464 465 466 467 468 469
470 95 96 471 472 473 474 475 476 477 478 479 480 481 426 427 428
482 483 484 485 486 487 488 489 490 491 492 493 494 495 496 (429 et
al. 1 (433 et al - 1 497 500 mismatch) mismatch) 498 541 542 499
543 501 502 503 434 435 504 505 506 507 508 509 510 511 512 513 514
449 450 451 530 531 532 533 515 516 517 518 519 228 229 245 246 520
521 239 240 273 274 522 523 524 525 526 527 187 188 528 529 245 246
518 519 530 531 513 514 532 533 532 533 513 514 530 531 534 535 371
372 391 392 536 537 538 539 540 541 542 431 433 543 432 495 496 497
498 499 500 501 544 545 546 547 548 551 552 553 554 549 552 548 551
553 550 554 550 553 548 551 554 549 552 555 556 557 558 175 553
(mismatch) 285 554 339 (mismatch) 548 549 550
TABLE-US-00004 TABLE 4 Pre-miRNA Sequences Seq ID premir_id name
PreMiR sequence 2 9261 premiR_09261
ugaaaucaacgaaucaccuaccuaacuggguuagggcccuggcucc
aucuccuuuaggaaaaccuucuguggggaguggggcuucgacccua
acccaggugggcuguaacacugcuguguuuucuaaggggcagaguu
uucuacuacuuuuccgcuggcccag 4 9252 premiR_09252
aggcagcucggcgggcggcgggcggcauucuggcgcggagcggagc
ggcggcgggcgcagcuagcgggucggccgcggagcggaggugcagc
ucggcuucccccggcaccccucccccucgggcgccagccccacccc uccgccggccgggccgaccc
6 9183 premiR_09183 aucacccucugaucgccgaucaccucugagacccaacuugcucaua
aacaaaacugcccaugucgguccucugcccuggaccugugacacuc
uggacuauuucuguguuuauuuguggccgaguguaacaaccauaua
auaaaucaccucuuccgcuguuuua 8 9128 premiR_09128
auccugccuuggcuggaaaagccagccuuccacccagcgccccuaa
aaugaucggguugacuccaguuuuguuacgaaaggaggccgggcug
cugagaggcucccugaguucccuucguggucgcccgucacauugcc cugcuguauacuuaauaa
10 9242 premiR_09242 cagacaaagaggggcgugagggagcugccugcaggggaagaagccu
uccguaucgagcugggcggucauuacacaggugcgcacagauaugg
augaggaggggcgugagggagccacgugcagggcagcaaguccucu
auaucuugagcuggguggucauu 12 9256 premiR_09256
acccccaaagcucuccugccugcuucugugugauauguuugauauu
ggguuguuuaauuaggaaccaacuaaaugucaaacauauucuuaca
gcagcaggugauucagcaccacccucuuucauacuucaaucucugg
ggcuccugucucuuuuacugaa 14 9147 premiR_09147
auugaguagggcaaauuuuaaauggguauuauuuuucaucuucaaa
caggcagaccuguuauccuaaacuaggugagucagcuuuugguaca
ugugaugauuuucaguguaaccaaugauguaaugauucugccaaau
gaaauauaaugauaucacuguaaaac 16 9111 premiR_09111
uuggcaggaaguuuccauggccaugcagccgcagggucacccugag
ugcuuuucaggguggcagggccuugccucagauggccacaagggca
ccucuccuuggauacuuuaugauucugugacgccagcuacuugguu
ugcuuuuuguauuuuuaugcau 18 9294 premiR_09294
ucuacaaaauuggugguauugguacuguuccuguuggccgagugga
gacugguguucucaaaccuggugugguggucaccuuugcuccaguc
aacauuacaacagaaguaaaaucugucgaaaugcaccaugaagcuu
ugagugaagcucuuccuggggacaaug 20 9192 premiR_09192
auagaaucaucuaaguauucagacacugcuuuccuaggaaauguua
aacuccuugaggcaggcuggcuuccucaccaccuugugcacugcug
cucccacaccacagugacuagcagcacauaaugguuugaauuaaag
cugaaguaaaaaauauccaggucca 22 9245 premiR_09245
uguagcuuaccuccucaaagcaauacacugaaaauguuuagacggg
cucacaucaccccauaaacaaauagguuugguccuagccuuucuau
uagcucuuaguaagauuacacaugcaagcauccccguuccagugag
uucacccucuaaaucaccacgau 24 9237 premiR_09237
auuugagaaggaggcugcugagaugggaaagugcuccuucaaguau
gccugggucuuggauaaacugaaagcugagcgugaacaugguauca
ccauugauaucucuuuguggaaauuugagaccagcaaguacuaugu
gacuaucauugaugccccaggacaca 26 9125 premiR_09125
ugccuaauacugacucagaugcacaauccaguuaacccagaugugu
gagaucuuccgguuugaaagaacuguauuggcaaggcaaaaucaac
cuauuguagaauauauuuauuguauaucagcauggggauuauuaau
auugcuaauaaaaccauuauuuguaaa 28 9234 premiR_09234
gggaaggauaaagggauggcaugguggggguuggaggcguggguuu
uagaaccuaucccuuucuagcccugagcaaugcuugccccagaagg
aguuggggcuaggcccauuccaauccuuccagccuaagauccagac uccaaggcaugccccag 30
9164 premiR_09164 ucaaucuggaggagcucaggguggccggcauucacaagaagguggc
ccggaccaucggcauuucuguggauccgaggaggcggaacaagucc
acggagucccugcaggcgaacgugcagcggcugaaggaguaccgcu
ccaaacucauccucuuccccagga 32 9139 premiR_09139
accccauccaauuuaaucggguguuauuuaauuauacuacuauaau
uguuguauuugcagguuugacuguucucagggaacgcugaagguuc
auaacaguagugauuuguaauugugaggcuugaguguggaauugaa
uuacuucauuagagaguaacc 34 9290 premiR_09290
ugguguggccaaggccaacacugagucgaccugauggagagaagaa
ggcauguguccacuggcuccugaugaccaugcuuuggauguugcca
acaaaauugggaucaucuaaucugaguccagcuugcuaauucuaaa
gguauauauguaucuuuucacca 36 9154 premiR_09154
guuaaaaaaaguaaaaggaacucggcaaaucuuaccccgccuguuu
accaaaaacaucaccucuagcauucucaguauuagaggcaccgccu
gcccagugacaugcguuuaacggccgcgguacccuaacugugcaaa
gguagcauaaucacuuguuccu 38 9269 premiR_09269
gaagaucacuacaacaauuugucugccuccaagguccucugaggca
gcaggcucuggggcuucugcuguccuuuggagggugucuucugggu
agagggaugggaaggaagggacccuuacccccggcucuucuccuga
ccugccaauaaaaauuuauggucca 40 9296 premiR_09296
cacgcuuguggugaaaucaggaauuuuuaggacucuuagcggugga
ucaaaaagaaaaaagaaaacaggacagaguaaaaucuugcuccaaa
gcuuguguucuggcaaauaccgucugugucucgaaugugaaguugu uuccacuccucagagcccac
42 9141 premiR_09141 ccgugaagaggcgggcaugacacagcaagacgagaagacccuaugg
agcuuuaauuuauuaaugcaaacaguaccuaacaaacccacagguc
cuaaacuaccaaaccugcauuaaaaauuucgguuggggcgaccucg
gagcagaacccaaccuccgag 44 9291 premiR_09291
cuuauuuacguuacuggcugaaagccugucugauaggauugacaug
gagcacuaauuaaucaccuagggucuccucauuuacuaaucauauu
gcacaaaacuucccugcugacugaggcucaagggcaaacugugggc
uuccagccagcuuauuuuucauaa 46 9263 premiR_09263
gauucaugaguagcagugacaaaccuaccgagccuggugauagcug
guuguccaagauagaaucuuagacaacucccuauaccagauccucu
aauuaauuuuaauaaaggccuucuauuuauacuagccacaucaagc
cuagccgucuacguacccacagaa 48 9251 premiR_09251
gggcagaagucugagccaguguuucaucaucguucuugcucugccu
cggucuguacaucugugaaaugggacucccucucuguuguggaggc
ccuggggacagcugggaggacuggaggggugguggggagguugugg
uccuuauuagacauucagauacc 50 9300 premiR_09300
ggcaagaacaaguaccuuacgaaagacagcaaaaagggagccaaga
aguggcugauccauuuucuuuuuuucuuuuuucuuuuuuuugagac
agucuugcucuaucccccuggcuggaauacaauggugugaucucag cucacugcaaccuccgccuc
52 9127 premiR_09127 caggccuuucugaaggaguuauucugcuaaaaauggucuuaguugu
cugaaaagccagcucuugaaccucuucacaacaguaucaacacugg
cuucucccgguucauuuuaugcgugcgagaagucagugguaacugc
ugcagggcuuaauacauuagu 54 9304 premiR_09304
ucacucgccaguaaccugucugcaugcaagacugggcagugacaag
cacgaugugcucacugcccaagauuuugcuuugauuuuguuuuacu
gcccaagaucugaacauuuuuugcaaacauagcagcuucucuaccu
cugcugcauugacauauguuugaa 56 9148 premiR_09148
auuuaaaacugugucuuucugugucccugaaauucucacacauggu
acguuuucaaugagcugauuuuguuucuccacucaaugcaguaauu
gagcuucuuugguucagugcaugagugguucagugguucauugggc
auccugguugagggaggggcu 58 9159 premiR_09159
uuuaugguugcuacuguagguuuauaauuuguuuauaauuuggccu
aauuuccaucagccauacuaauauuggauuuuaaaaggaggcaacu
uuuuuucuuuuugaaccaaaggaaugaguuagcuuugaaaacauaa
uuugggauauuauaguaugga 60 9205 premiR_09205
ccugcugggguuggaguucuuaaugaacauacaagugaauacacug
aggcaaaaaaauuaaagcucuccaacugugggguauucauucuguu
cacuguggccaguguggugaucaguacuggccacaccaguggccaa
agagaacugcauucaucaugug 62 9220 premiR_09220
ccucagccccuucagagagcgacuuucaaacucgcgcccgcgucgc
ggcagcaccugggcagccccgcacgccgugcgcgucccgagcccgc
ggggcagcuaccgcucggugaguguccccugauucuccucucuccc
cucuuaucucccugcauuaggcug 64 9182 premiR_09182
cugaaaaguuccagcauauuuugcgaguacucaacaccaacaucga
ugggcagcggaaaauagccuuugccaucacugccauuaagggugug
ggccgaagauaugcucaugugguguugaggaaagcagacacugacc
ucaccaagagggcgggagaacuc 66 9121 premiR_09121
caugcuuuugguuuguuaccaaaauauacaguguggugaagguuga
cugaagaaguccaguguguccaguuaaaacagaaauaaauuaaacu
cuucaucaacaaagaccuguuuuugugacugccuugaguuuuauca
gaauuauuggccuaguaauccuu 68 9276 premiR_09276
augguuacuuauaugggaaggguggguaacaaggauuggacagggu
uagauuagaccccucugaagguaccuuguuuuauaguuguaacuuu
uuuuuuuuuugagauggagucuugcucugucacccaggcuggagug
caguggugcgaucucagcucacug 70 9247 premiR_09247
cacacgauuaacccaagucaauagaagccggcguaaagaguguuuu
agaucacccccuccccaauaaagcuaaaacucaccugaguuguaaa
aaacuccaguugacacaaaauagacuacgaaaguggcuuuaacaua
ucugaacacacaauagcuaagacccaa 72 9198 premiR_09198
ggcggcugccgaagauggcggaggugcaggucccaguccuccaugg
ucgaggccaucuccugggccgccuggcggccaucguggcuaagcag
guaauguugggcuggaaggugguggucguacgcugugagggcauca
acauuucuggcaauuucuaca 74 9123 premiR_09123
gcccaggacuccagcucaugcgccgaauaguagguacaguguucca
augucuuugugguuuguagagaacaaucaacggucggcgaacauca
gugggauaagguaaaauggcugagugaagcauuggacuguaaaucu
aaagacaggggcuaagccucuuuu 76 9146 premiR_09146
ugguguagaggcggagaggagccaagaaacuaaaggugaaaaauac
acuggaacucuggggcaagacaugucuaugguagcugagccaaaca
cguaggauuuccguuuuaagguucacauggaaaagguuauagcuuu
gccuugagauugacucauuaaa 78 9137 premiR_09137
gacuaaaacuauuugaucuuuuaauauuuaauuaaugguuccccgu
ggcguuuuuauagucuguguucuauugugagcaacgagauuuuaau
aagcagguucaggacuuuccauuguguagcaagaugucauugcuuc
caugacacuaauuuggcuuucaua 80 9186 premiR_09186
cacauacucaaggagcccuguuuuacagggcacuggagaacuaauu
aauuugcaaugcagaaagaaugcagugacaucugaaauauuggccu
cggguaucacaggucauuggaaauaguuugugacaaacuggggugg
aggguggggguggggaaggcaacucu 82 9115 premiR_09115
gggggaggccugcgcggaggagcaccgcuuccucccgccgggaggg
ggagucccgggcuccugcguccugucucccuccccggccgucugca
ggagcacgaagggagugccuccucuucgucuccucgguccccguaa
cuucuccccucacuuccuccugg 84 9203 premiR_09203
gucaaaaguccccagagucuucacacaagccgugguguaugaagcu
gcauccucaggaccugggcuugggugguaggaggaauuggugcugg
ucuuucauuuuggauuugacuccagccccacagccucagccacccc
agccaauugucauaggagcugga 86 9241 premiR_09241
gcgcggcggagggggggguggggccuuggggccgcgagugggagcg
ggagcgguucugcggccuccucgggcuucuuggcccugggcggagu
gggauugggugucccggcuguucgcaguggccgcgagugcggccgg
accuggaguaguaccugagccgcu 88 9156 premiR_09156
gagaacaugggucaccagcagggccugagaagagggagaaaauacg
gaaaugugggauuggggucgcugagugcaggcauguaaguuaagug
uuuggggaacagagcagugcuugacugaguguggcuggacgugagu
acugagggggacaaaugagauugaucc 90 9281 premiR_09281
ucgcgggucuguggcgcggggccccgguggucgugucgcguggggg
gcgggugguuggggcguccgguucgccgcgccccgccccggcccca
ccggucccggccgccgcccccgcgcccgcucgcucccucccguccg
cccguccgcggcccguccgucc 92 9124 premiR_09124
uuaggaagucugagaugauaaauauuucaaggucagugaagucuau
caaucauucuccccuuccucaucagcaaugguagauagaaaugucc
uaaacuuuucuaaauccuagugaugaggaugugcugauauucaaca
uaguccuuaaagugaaaacuga 94 9171 premiR_09171
agcuacuuccuuucuucagccucuugcuuucuguucaaaucucagc
uuuuaucacauucuuuucauggagagacaucucaaugucccuuuuc
gccuaggagaagaauguuauugaguggggcucaggauuuaaaccca
ggcagacuaauugguaugugag 96 9155 premiR_09155
cacauacauuggcgaaaagaccaacaagucaugauuguucugaagu
ucccuuuaucauguuguccccuaaucucuacuaccaguaagccuuu
guguuaucuuaggaugaggcaugggugguucaguguuuauaauaag acgagucuaaaauggacaau
98 9142 premiR_09142 uguuuaaugccugaaauccaagucuuccuccaugggaaaauacugu
uauaccaaauaauucuagaugaguaacaaagaucuuuuuaggccuu
cauuuuauguuuuuucuuaacuguuauauuaugauugugacauaga
uuauacuacuacuaauuuuuggaug
100 9166 premiR_09166
cuuacggaaaaggaacagauuguuccuaaaccagaagaggagguug
cccagaagaaaaagguaaauaaguaguugcucgguuuuguuuguga
uaguagaaagauuugugguugcugugaugacuaucuuaggacaccu
uuggaauaacuaugaaagaaaacuau 102 9149 premiR_09149
cuaggagugacucaugcagacucaagcagaaccuuggggcccaggg
cagaagugugacuucaggucucacucagcacucaucagaacacuca
cucaguaugcuuguaauuagaauugagugaucucauggaugaacug
uacugggcuaacuugaagagcaca 104 9118 premiR_09118
uaaaagcugaaaaucucaguuuaaaaaucaaaauguuaacacaaag
cuaagauucaucagagcccacccuauucuaaggaaccacaauaacu
uacucuggccccaguguuaaaacgaucuuucagauuuagagugacu
auguaagaauuuaggauuuccucuuu 106 9201 premiR_09201
ggccccgcggggcccguccgcuccuccagccgcugccucccgggcg
gcgcucgccggcgcggcggcaaagacugagacagcuccgcugcccg
cugaacuccauccucccggcggucgggcggcggcggcugcggucgg ucgcggcagcggcuccgcu
108 9267 premiR_09267
ugccuggucaaaggcuucuaccccagcgacaucgccguggaguggg
agagcagcgggcagccggagaacaacuacaacaccacgccucccau
gcuggacuccgacggcuccuucuuccucuacagcaagcucaccgug
gacaagagcagguggcagcaggg 110 9200 premiR_09200
guggggaggucgaugaaugagugguuaauuaauuuuauuagggggu
uaauuuugcguauuggggucauugguguucuuguaguugaaauaca
acgaugguuuuucauaucauuggucgugguuguaguccgugcgaga auaaugauguaugcuuuguu
112 9193 premiR_09193
auucucaguauuggaucugcacauggaguguuuuucucucuagugg
uuacagaggaugaaugcauauugagauaaagaagugauuuugguuc
caaaaggauuuuaaggaugaugagagaacaguggguacuucauugc
caggucaugucuuugcaagaagaaa 114 9292 premiR_09292
auucgugcugaaaaucucagacucauugaugauagcugccagugac
aggaguaguguugccacuguaagauacgccaucuuuguuaguuacu
cucaucuacucguuucuuguauucugccucuuggucaucuuugauu
cucauuuaucugcaaauuuucuuggua 116 9283 premiR_09283
aucaaacacuuauccuauuaaacacagcauccaucggugaucccag
ugacaaguaauugaauguuaguucuggagucuuuccuggggugaug
gccuggagaagccucucuuuuaaggauuagauucagagguagaggu
aaaugaguguugagcaccaggaagag 118 9271 premiR_09271
gugcccgggagguggacuggggccuggguugugcuggaggccaggc
ugaggcccugccuugguuuggggaggagaucccugcacuccggaac
uccucuguggcccacggaggaucgcucugaacugccucagcguggc
ggccaguggggguagggguggagaga 120 9136 premiR_09136
cuaccugaaguuuuaagagucuuggaaagucaggagugacuucugc
uaaacacggggcuuuccagagucagagaagcuagcaagccuguggu
uuggaccagguacuaaauauuugacaagaguaugccagguguaaug
agcuacugucuauuccccuuuaaagc 122 9284 premiR_09284
cucaccucgauuccucccaggccuggguccagcaccagccuaggaa
gagggugccccaugcugucuagcucuucuucgggauggggggcucc
agguuccuugguauuuugcuuuggccuuuggagccucagucaaaac
ugaggaaaggugucauuuucacau 124 9185 premiR_09185
uaguaaauguaggcaguuucuuuaggguuaaucaucuuucaaaggg
ccuuaggaauguccuucaaacagaauauaaaugucaaagagaauau
cucuuucuguuugaaauuauuugggcagguugaaagaauuugauaa
agggaaauucuauauuuaaucuuuc 126 9270 premiR_09270
uguuguaaacaaacaaguuaagagcaagauucuugccaagagaauu
aaugugcguauugagcacauuaagcacucugagagcugggauagcu
uccugaaauacaugaaggaaaaugaucagaaaaagaaagaagccaa
agagaaagguaccuggguucaacugaa 128 9197 premiR_09197
ccugaagaggaagagaugacuguuggaaagcguuccccucccccau
acggcagaacagcugcggcucccaggggaaagcccccgcaggacag
uccucguggggugugacggcuguugguggagaagguuuggcgcccu auuuucuuaucugccuuucu
130 9225 premiR_09225
gagaggaaaaguccagaccuaggacuaguuauggcaguuggagaga
aagaacaucgggauguuugaaaauaugccauugacuaucuuaacua
cuguaauuuuaucauuuccaacgucaucuaacuggggacuagaaca
aacugugaauucacuuucagcaac 132 9112 premiR_09112
ucuuaaaaguuuuauuaaaggggaggggcaaauauuggcaauuagu
uggcaguggccuguuacgguugggauuggugggguggguuuaggua
auuguuuaguuuaugauugcagauaaacucaugccagagaacuuaa
agucuuagaauggaaaaaguaaagaaa 134 9307 premiR_09307
gaaggaugguuauucccgccuggagaucccacaguagggccuuugg
agugauagacauccccaucucccuccacacccugcccuaccgcccc
ccaacccccaaagcuucaacaaaggcuccuuuuuaaaguuuuccgg
ugcccuuugcucuuuguuugccuu 136 9295 premiR_09295
cuucaaguaugccugggucuugcauaaacugaaagcugagcgugaa
cgugguaucaccauugauaucuccuuguggaaauuugagaccagca
aguacuaugugacuaucauugaugccccaggacucagagacuucau
caaaaacaugauuacagggacauc 138 9120 premiR_09120
uggauuucgcccccgcucccucccggaaacuccuccuggugccugc
gaccguucucacugagcaugugcagacggcggugcgcaugcucugu
ugcgguccgcuucgguuucuguugcgggacccggggugucuccuag
cgcaaccggaacuagccuucugg 140 9153 premiR_09153
cacuguagcauaagcaagggcuuaguuccugaacugaguuacagcu
uuauuuuucuuuugauucagcauguuuuuaaugauccauaaguuaa
aagcugcugguguuuuuauuaaagcugccauuuguuacuaaccagg
cucugugugacuccuaaguggaa 142 9116 premiR_09116
agccacaccccagcagugugcaagggaucagacacaagguugaauc
caucacaaaagcagaaucaccauggcaacugcauccuuugauucuu
gagugugcccagcaaccugagcagaggcgauaguugaagugaacca
aguucuccugagaaauggagggga 144 9312 premiR_09312
aguccgugcgagaauaaugauguaugcuuuguuucuguugagugug
gguuuaguaaugggguuugugggguuuucuucuaagccuucuccua
uuuauggggguuuaguacugauuguuagcgguguggucgggugugu
uauuauucugaauuuugggggagguu 146 9221 premiR_09221
aggaaggggaaacucaaucagcaggacuucagaaagggccuugugu
uuauagcuuugucaaguaaauuuggacgcagcuggagcacaggccc
uguuuguuugcacauaauaaucuuguuuaucacuuuaaaaaauuca guaauaucucagcagucagg
148 9151 premiR_09151
acucccugacagauaucucccucuuccauuucaucaagacccagcu
gagucacugucacugccuaccaaucucgaccggaccucgaccggcu
cgucuguguugccaaucgacucggcguggcgucggucgugguagau
aggcggucaugcauacgaauuuucag 150 9157 premiR_09157
agggacaaugccauauuuauccuucuagcccugacaccucacacaa
ugcagagaacggaagggaguucaauaacugguagcaaagugccaac
uccuugagaauagggccuguguuuagugaguauuuguuaagagaau
gaauaaaugauguacaguugua 152 9305 premiR_09305
agcauuugagggugaugauggauucuguguguuugagagcaacgcc
auugccuacuaugugagcaaugaggagcugcggggaaguacuccag
aggcagcagcccagguggugcagugggugagcuuugcugauuccga
uauagugcccccagccaguaccugg 154 9309 premiR_09309
acaaggauggaagaggcccucgggccugacaacacgcauacgguua
aggcauugccaccuacuucguggcaucuaaccaucguuuuuuuuuu
uuugguguuuuguuuuuuauuuuucuucagacggagucuuauucug
ucgcccagacuggagugcaauggcgcg 156 9239 premiR_09239
uuuuauaaccauagaguggagacagucaguaugaccaccaaaccca
ggagccauauauuaaaauacugauaaauuuaacuauauaaaaaaau
uuuugccgggugcgguggcucacaccugugauucuagcagaaaauc
agaucaggagaucacagaagguc 158 9177 premiR_09177
guccucccacuggccgcacucugugccccauggcccuccugcgccc
cgcccggcguccucucacggccucugucugugcugagcuuggguaa
cucuuguucuuaccuccacagagucuguagaagaggcgacaccagg
gcuuccaaaugaacaaccgaaa 162 9122 premiR_09122
ucauuucugccacagucuuuuuuguugaagcaaguuagcaagcacu
aagcacaucuacaaucaaggagaggggcaggcuuuaccuuuugaag
gaagaaguaugaaaguguaucacugacugaucaaguagagguaagc
aguggaggacacucagaauaccuuuu 164 9275 premiR_09275
acuaauaagcuacaaaacauuuaaaugacuagugucugugugugcu
ugcuaguauuauuauaccaucagaaaguaaaaauggacauacaugu
uaugcauuaaacccacaagagagaaaacuugaggacugauuaauuu
aaguaguaaaugaauccaagaa 166 9259 premiR_09259
ccuaaaauucucaauuaggcuauaaaugcaagaugagcugaaggga
aauaggugauuuccauucuguaguguguauauaugagguuuuauuc
ucaugacaagaaacagacuaugcaaaucucuuuaauuucuggcauu ucagcuuucuagaauua
168 9248 premiR_09248
gggucaauaguacuugccgcaguacucuuaaaacuaggcggcuaug
guauaauacgccucacacucauucucaacccccugacaaaacacau
agccuaccccuuccuuguacuaucccuaugaggcauaauuauaaca
agcuccaucugccuacgacaaa 170 9191 premiR_09191
aaccagaacgugguuugccugaggcuguaacugagagaaagauucu
ggggcuguguuaugaaaauauagacauucucacauaagcccaguuc
aucaccauuuccuccuuuaccuuucagugcaguuucuuuucacauu
aggcuguugguucaaacuuuuggg 172 9172 premiR_09172
guguguguauauauguauacauauauguauguguauguguauauag
agagagagcugagaguuauucuauuuauuccuuuucucuccuaauc
ugaaaauggguguucuguauuuuggguggaagaggcauagaagggg
auguguguugucucuuaagauu 174 9199 premiR_09199
agacagaaaucaggacuaaguccucugcuucaguuucauuguuaac
gggccuuauucugaucucaccugucgcguagcucuaauauucacau
aaacugaaauaaagaaguggaaugaggagcuuugacauucaaauua ugugauguaauuuaucuucc
176 9160 premiR_09160
uauguguaagauagaaugaauauugagcaggaugcuuuaaaaguga
ccaagcagauuugaaaaacauuaaaaauguuggccuucucguccca
guucuucccaaaguugagaaaagcuggguugagaggaugaaaagaa
aaaaaaagaaaaauuuagugga 178 9272 premiR_09272
gaaaauaaaggcaccugaaaagaaacuacuacuuuaacacugcugu
ggaaggccuuugcuuuauaagaaaaauauuauuagcuaugggaaag
uaauguucuuuauguaaagacuuaaaaauagacuaauaguuuacag
aguuauuauauaaaauacgauguga 180 9279 premiR_09279
aaucccuguuugcuucagggcgagaugugugacagagguggcauca
agcucuuacagucccaacccuccaacggaaaugggcgaagaucuca
ggaauggcaucggucacaggaaaucgauaguggcuggcugcuagca
uggccacuuggggcuuaggcag 182 9236 premiR_09236
ggaaguuugggauaguaaaguuuguugccuuugugucuugugucuu
uuuuccuuuucuuccuuucuugggggagauagauagauagacagac
agacagacagacagacacagagagagagagagagagagagagacag auaguguucauggauccugu
184 9143 premiR_09143
guaaaaugaaaucacagugguaugggccucaugggguuaucgaaag
aaugggcugaggucaugugggccaugggcuugguacagugccugag
acauaaugaauacucaguucccugguggucuagugguuagaaaaau aauaauaauaauaauaau
186 9298 premiR_09298
uuuagaaguuucagucgcacacuccuacccgggucggaguuagcuc
aagcgguuaccuccucaugccggacuuucuaucuguccaucucugu
gcugggguucgagacccgcgggugcuuacugacccuuuuaugcaau
aaauucgguauaaucugucacucuga 188 9260 premiR_09260
gggcagccguggggcguggaagccgcgcagaggccaaggcugcggg
guucuucgucgucuacaggcuuucgcggcucaguguggaaaacccg
ccguuuccucgcgccccacguccgacccaggccuccugggcacccu
ucggggaggccgcgaucucgg 190 9228 premiR_09228
guggcgucgcgugugaggcgcgugcagggugagugugaguggacgc
gugagugugugagugugcgcgcuuggagcguguuaggcgagugcgu
gcgcccaccccugcgccccuccucccgcuuacacuuugaucuuauu
ugaucggaucgugaccccagccccgc 192 9219 premiR_09219
gggcuaguguguuuguguuuccauucuaagauugagucuggcaguc
ccuguuuuuuugcauugggguaacugcucuuugauuuuuuuuaauu
gcaguauuugugugauugcaauaauaaaguuugguuugguuuuuac
agucaugcgcagggacgauccuug 194 9244 premiR_09244
agcccagaaccccucucaccccagaaccuuccuugacuucugccag
aguugagcagccggcccucugguaggcgcaugugaguggauguggg
cacauguggcccacuggaucugguggauguggucgcgucuggcccc
cuggaucuggugggugugggc 196 9180 premiR_09180
gugguauuugugauuugguuaaucuguauaaaaauuguaaguagaa
agguuuauauuucaucuuaauucuuuugauguuguaaacguacuuu
uuaaaagauggauuauuugaauguuuauggcaccugacuuguaaaa
aaaaaaaacuacaaaaaaaucc 198 9288 premiR_09288
ugguaagaaacuggaagauggcccuaaauucuugaagucuggugau
gcugccauuguugauacgguuccuggcaagcccuuguguguugaga
gcuucucagacuauccaccugugggucgcuuugcuguucaugaucu
gagacagacaguugucgugggugucau 200 9306 premiR_09306
ccccgcucaccuccucuauccccacaguguacugcugcugcugcuu
ggccaacguuucacugccuggcaucgggggcaccauuccugagucc
aaaccuuucuucuacgugaacguggcugacaucgagagccuggagg
uagagguguccuauguggccug
202 9287 premiR_09287
aguugcgacaagacagaguuggagaauagaggagguucagaguugg
aagaaaugggaguaggugauggcaacaccgaguugucagagugagc
ugaggcaacauccucuacuucuagcucacugaugaaaauauccagg
auagcgggucugggguccagu 204 9258 premiR_09258
cagagugggaccggcagcucccagacuugaggcggaggggccgcgg
gccggagcucccugcagccgcuagccugggaagacuggagugcgcu
gcccaccgagggucugcgccgcgccggccgccccgggccgcuuugu
gcgcgcccgcgcggucuguac 206 9299 premiR_09299
cagaacccaccaaccagaacgugguuugccugaggcuguaacugag
agaaagauucuggggcuguguuaugaaaauauagacauucucacau
aagcccaguucaucaccauuuccuccuuuaccuuucagugcaguuu
cuuuucacauuaggcuguugguucaaa 208 9285 premiR_09285
gcucuuucucuucccucucguuuaguuugccugggagcuugaaagg
agaaagcacggggucgccccaaaccccuucugcuucugcccaucac
aagugccacuaccgccaugggccucacuaucuccucccucuucucc cgacuauuuggcaagaagca
212 9187 premiR_09187
acuuguucccuaaauagggacuuguacgaauugcuacacgaggguu
cagcugucucuuacuuuuaaucagugaaauugaccuaucugugaag
agguggauauaaaaaaauaagacgagaagacccuauggagcuuuaa
uucauuaauacaaauaaaaacucaaac 214 9194 premiR_09194
uggacacagaagaaacgagggagcccgggucuccuccgagugugca
acaagcuggccuggggccccccgaaaggacgcuggagagaagccca
ggaucacccagucuuugcagcagggugcaggcuuggagucccccca
agggcggcuagaaucagguccagg 216 9204 premiR_09204
ugugucuuuuaaacuggaaaaucuucuagcauguuggguuguuaca
gaguauauuuuugucugcagcuguuuguugccccauuccuaagagg
aguuuauccauccugacuuguagcugugugacuucuugcagugccc
ccaccccauccccccgggagag 218 9212 premiR_09212
cagaagugacuuuacuuucucaaguuugauacugaguugacuguuc
ccuuaucccucacccuuccccuucccuuuccuaaggcaauagugca
caacuuagguuauuuuugcuuccgaauuugaaugaaaaacuuaaug
ccauggauuuuuuucuuuugca 220 9303 premiR_09303
cguucuccgcacuccugcuccgcgagggccccuucgaggcggcuga
gacccgagugccggacucccgccgcuggagcggggcucggguucgg
cagccggaaggaggugugcccccggggcgcuugggggcgccugagg
ucccgaggggaggcaagauggga 222 9218 premiR_09218
agaagaaaacaaguuaauuugaagagagucagaaagcgugaguguc
cagagccuacugagcccuggaagucacggauaaaaacaagaaguga
agucaacacucucggugagaaagggagcgguacugacaaacuucua
ccaucccagugugcccgguugcuccc 224 9144 premiR_09144
cccucccggcgcgguuggguggcgccucagcgggugggcagcaugg
ggcggggaggguguccccuccgcgccguuaaaaugaaacucuagug
gcuggaguccgggcagagcuugagggcaguuggugcggucggguug
guucuuacaccccggcgggagc 226 9262 premiR_09262
aagauucaugaguagcagugacaaaccuaccgagccuggugauagc
ugguuguccaagauagaaucuuagacaacucccuauaccagauccu
cuaauuaauuuuaauaaaggccuucuauuuauacuagccacaucaa
gccuagccgucuacguacccacagaau 228 9211 premiR_09211
gccuguugagaaagaccucuggggcccuguuggagauggcuggcag
aaugguucucuugaugagcuucaugauaaagcagacuugccaauaa
uaccaagagagaagacuggcucuacucuccaaaggaguccagggac
agagagucagacagaugacaucagaag 230 9301 premiR_09301
cggcccuguccugcggggguccggucgcggaggcggcggaggggcg
cggggacacuccccaccuccacuguccgcccgucggccccgguggc
cuuuucucgccucgcgcacagcucccccgccgcagggcugagagag agaguggccgucuggugc
232 9231 premiR_09231
agaaauaauaaugcaggguuucuucaaaauaugguucuggacagug
gauuauaguuaccuggagagcuuguguuaaaauaucugaggaugau
uccaaguaccagggcuuauacacaggaauacuugagaaccacugca
cucaagcauuuaaaauuuuccu 234 9240 premiR_09240
uccuagggcuuugacaccaaaaucuacuaugaugagucaggcuagg
cuauaaacuugcaaggacuuagagcccagaaagugacaagcccaac
uagccugccucuuggaggaaaaaagaagaauagcucaaaacacuua agaagguaaggaguccaagc
236 9190 premiR_09190
gggaggguggacaguccuuaacugcucugcagguccaggauguuag
aaaggggcagggacaacaaaugggugaccccaaccucaaccugcug
cuucucucuccaguccccaugugagagcagcagaggcggucuucaa
cauccugccagccccacacagcua 238 9145 premiR_09145
uaggucuguuauuuucacauacacuugguaacucagacuggucuga
auauaaaguagaaauagcuaagaaccauuuguaaugaaugcaacuc
uuauuuguuuuuaaugguguuuuaaggacuuaaggguauuagaacu
gacaacaguuuauucaguuaagc 240 9255 premiR_09255
acccccaaagcucuccugccugcuucugugugauauguuugauauu
ggguuguuuaauuaggaaccaacuaaaugucaaacauauucuuaca
gcagcaggugauucagcaccacccucuuucauacuucaaucucugg
ggcuccugucucuuuuacugaaccuc 242 9131 premiR_09131
auccguuuuggaaccugcgucuggggcuccagucgcugcucuugcu
ggcguccaucgccgccucggacggccgugcauuuucucgucucacg
caguucgaggaggacccuagaaagccaggagcugugauugacagua
gcuguagguuaccagacggcaac 244 9129 premiR_09129
uacauuuuagggugguagagcuacuccuuacuuuaaaugcuaccua
cucacugugacacuguuuaauaaaugguuauugacuagagaaguag
ggaucucugucaccuagcauucuaagucagucagucaucaguuuuu
guagguuaucucagaagcaauag 246 9208 premiR_09208
uucuucuucagcaaacauuaggagaguaucuucucuguuuuggcca
uguguguacucacagccccucacacauggccgaaacagagaaguua
cuuuccuaauauuugccuccuuggagugucucaaguccuggaagca
agagauaauaagcaauuaauauaca 248 9232 premiR_09232
cccgacagaucgacuauguugaucuaacuuuucuaagccaguuucu
gucugauaugccagcuugagcagcuccuuugucccagcuccccugg
gcaucuagcugaugggagcucauuuuucuguuuuuucauuucaggu
uuauuguuggccaaaaccaggcuuu 250 9109 premiR_09109
gggucauggaccagcgccucagugcauuagucauucgcuuuuccuu
acagacaaaucagauaacucuuccccagugauugucaaauguauga
auguaucucuguaaaugugguuuugacaugucacuguuacugaagg
agaguauggaauccccacagga 252 9179 premiR_09179
ggugaggccccgcgcgugugucccggcugcggucggccgcgcucga
gggguccccguggcguccccuuccccgccggccgccuuucucgcgc
cuuccccgucgccccggccucgcccguggucucucgucuucucccg
gcccgcucuuccgaaccgggucgg 254 9308 premiR_09308
gcccagcacccucugucucuuuaugcaaucagugccagguggggag
ggaugcauucuguccaaugacaugcaggcacuuuagagggcuugca
uucauucccaaguccagcggcacacuuuauacauccuuggcugguc auugaggggaacaccggag
256 9249 premiR_09249
accgcaacucccuuucccccacuuuccccaaacgggaggcgcuagc
cauggaacauggcacauccagggcuaccuccucccaaguuacccag
aggucauguguacaagcagcaauucuaacaacagucccucaggcgu
gagcggcauuuuacaguuugcaa 258 9246 premiR_09246
cuuaccuccucaaagcaauacacugaaaauguuuagacgggcucac
aucaccccauaaacaaauagguuugguccuagccuuucuauuagcu
cuuaguaagauuacacaugcaagcauccccguuccagugaguucac ccucuaaaucaccacgauca
260 9226 premiR_09226
cagagcaacuggcuccuggcagcugugcuuguccguuuccugucag
agugaucccagguuuccuccuggcccgucccauggucccuccacag
gagugugagaggaugggggaagcacugugggaagaccaccaaagau
ggcuggacagugggagagagcacguu 262 9282 premiR_09282
caggacggccgccaucuugcgcgcagcuggagucggugccugaggu
ugcagccgagagugugcgccagcccgcggcccagccgaagcucuuu
cccgccgccucuccgcgccucgcccagguucagcuccgccugaccc
uccgcuuggcacgguccccug 264 9184 premiR_09184
guuggcuuuuccagggccagcgugaguggugaggccagcucucuca
gugaccaucagagacaaggccuuggccaguccaggggucuuggggc
uccacuuuucugaauuaugaaauguugaguguuuacccugucaaua
uauauaucauuuauauauuuuuugu 266 9264 premiR_09264
agauucaugaguagcagugacaaaccuaccgagccuggugauagcu
gguuguccaagauagaaucuuagacaacucccuauaccagauccuc
uaauuaauuuuaauaaaggccuucuauuuauacuagccacaucaag ccuagccgucuacguaccca
268 9162 premiR_09162
gaagcccaaguuugaauugggaaggcucauggagcuucauggugaa
ggcaguaguucuggaaaagccacuggggaugagacaggugcuaaag
uuggacgagcugauggauaugaaccaccaguccaagaaucuguuua
aaguucagacuucaaauaguggcaa 270 9227 premiR_09227
cucucagcucugcagcugucugcgguggggggaagguuggggggug
ucuggaggcauguuccccucaccaccccccgugggucucagggagg
ccgggugugaccucaucuuucucauggugcuauccuggugcuauug ggguggggagcucccuccc
272 9206 premiR_09206
auccuuuagcacguuuggauaaaguuggccuucuagguuguggcau
uucaacugguuaugguccugcugugaacacugccaagcuggagccu
ggcucuguuugugccaucuuuggccugggaggauuuggaucggggg uuaccaugggcuguaaagug
274 9254 premiR_09254
ucucuaauuagcuuucccaguauacuucuuagaaaguccaaguguu
caggacuuuuauaccuguuauacuuuggcuugguuuccaugauucu
uacuuuauuagccuaguuuaucaccaauaauacuugacggaaggcu
caguaauuaguuaugaauaugg 276 9134 premiR_09134
cauuaauuagguaauauuuuccucauuucuuuacugcugccauuuu
ccucaccaguauuccagagauggucauagcucauuacucuaccacc
aagaaccuaaaaggaauuagaauacagcagaauuggccucagugaa
gagcuuaaaauuguucuccucgua 278 9289 premiR_09289
ugauauuacucaccauugauaccucuguuuggaaauuugagaccag
caagugacuaucgcuguugccuuaggccacagagacuuuaucaaaa
acgugauuacagggacauaucaggugggcuguguuguccugauuau
ugcugcugguguuggcaacuuug 280 9152 premiR_09152
acucccugacagauaucucccucuuccauuucaucaagacccagcu
gagucacugucacugccuaccaaucucgaccggaccucgaccggcu
cgucuguguugccaaucgacucggcguggcgucggucgugguagau
aggcggucaugcauacgaauuuuc 282 9238 premiR_09238
auauaaaaacauuaggucaagguacagccuaugagguggcaagaaa
ugggcuacauuuucuauauccggcaaaucucacaacaaccuuuaug
aaaucuaagggcucaaggaggauuuaguaguaaaccaagcgcagag
ugcuugguugaauaaggccaugaa 284 9158 premiR_09158
uuucauuuucugugauuauuuuuaaauuagcuucuguguaaacuca
cuaacuuguucccacaugacaauuuauagcaguccaaagauuuuuu
uauagccaugguuguuauaauuuugacagaugcucaaggcuguugu
uugcauuguucuucagaauuucaucuu 286 9168 premiR_09168
cauuacacuccagccugggcaacaagagcaaaacucugucucaaaa
aaaugaaaagaaaagaaaauaccuccauggggccuucucuucccag
uucuuccuggagucggggaaaagcuggguugagaaggugaaaagaa aaaacaaaccuugacugggc
288 9175 premiR_09175
ggaguagcuguaacauuauguggaaagcaagugggagaaucaugaa
aaaaaauaaucccauagauggagaagaauagaaagaaggaaaggag
cauugccuaguguugguuauugaugguuaagcucagcuuuuauuua
uucaauaggccugcagauguagacu 290 9216 premiR_09216
uguaggauuuuuuguuuuuguagcuaacuuauggauugagauguga
ucaaaggcuuuauuaaauuuguacuucagcauaugauggcugcguu
cugcauuucauuccgccauaugccuggaccguucacacuuggguau
cugggcuuagggagcauguaggcuuc 292 9117 premiR_09117
ucuagcucuguuauaaagaaaacauuuaggaaauucucucuuucuc
ucuuucaccuauccuacuuuuuguguguccuuuguaguuuugcacc
aucauuccuaacgaauuuauuuggcauuuggaagauagguuagcaa
aaauuuuacuauauuugaaaggcua 294 9167 premiR_09167
ggucaccaggcugagaaagcaggagaugcuacugcugaggaacugu
cacuugucauuucaagguccacuccuccacccucuggcagcaugag
ucgcucugaaagauuuugaagcugggacaggagagggugagugagg
ugaggccuccgcaugccagguuuuc 296 9126 premiR_09126
auucaucucugguuuucuugccacccucugggaguccccaucccau
uuucauccugagcccaaccaggcccugccauuggccucuugucccu
uggcacacuuguacccacaggugaggggcaggaccugaagguauug
gccuguucaacaaucagucaucaugg 298 9274 premiR_09274
uuuuacauaaguagacacaggugggaaacuguuuagacgggcucac
aucaccccauaaacaaauagguuugguccuagccuuucuauuagcu
cuuaguaagauuacacaugcaagcauccccauuccagugaguucac ccucuaaaucaccacgauga
300 9140 premiR_09140
acuguuuauaagucgguguuguaaaucugaugugaauuuuuguuuc
uuuuuucuuagauuuuugccuuuaugacgacagcuuguuaugguug
caguuugggucuggcuuuacgaagauggcgaccguaacacuccuua
gaaacuggcagucguauguuag 302 9135 premiR_09135
aguaggcaacugaggacugauuucucagggugauuagaaaggaaag
gguggcggccuccuuucauacuucggaaagucuuguucccaucagc
cuuuccucauggugccauaacuggaauggcggcaagguccucuuuc
cugugccugugucuuaaguuucugg 304 9176 premiR_09176
gcgcccccaaaguguccccuccugcugugacuucuagccaagaaga
cauuucucccauggccaagugaucucugauagauccuguaggacca
cugaagucagacaggacaaguugagucagggccuguguguccagug
cgcagcaugcuuggggagugaca 306 9178 premiR_09178
uaacgcaugcgcggggagggcggagcugggcguugccguggcuacu
gggaacgcauuucacgggggcggggcgugguuccggggcggggcgc
ggccgccggaagugcguggccgcccggggccauggcgacacucagc
uucgucuuccugcugcugggg 308 9207 premiR_09207
ucgauggguguucuuuuaaaauacgguucuaagucuaagucuaaca
uucgguguaucuaaccgaauguuaauugauggagacaaggugauac
ggguucagaaaauagaauucagaaaagaaaaggaagaauuggcaaa
auucagaaaucaauuuuaagaaaaau 310 9163 premiR_09163
cccacggauucgccccgccgcgccucuccgcgcguagauuggccgg
agcgaggcgaacgggcccggccuugguagccgccgaccgagcgcug
gcuguccuggaaccuaggcggcgggagcccggggcgccucgcggca cggaagagcggcgagaug
312 9161 premiR_09161
ccuguuccccccaaaacccaaggacacucucaugaucucccggacc
ccugaggucacgugcguggugguggacgugagccaggaagaccccg
agguccaguucaacugguacguggauggcguggaggugcauaaugc
caagacaaagccgcgggaggagc 314 9273 premiR_09273
cuccgugcuaccuaucacaccacguccuaaaguaaggucagcuaaa
uaagcugucaggcccauaccccaaaaauguugguuacauccuuccu
guacuaauuaaccuauuagcucagcuuaucaucuacuuuacuauuu
cuacagguacccuuaucacaaugc 316 9133 premiR_09133
uggaaaucacagcaacccauugaaaacugcccuccccaccagaacg
ugcuacguucuuucuucaugccuaugugugcuccauuccucauuuc
uacuuggcucaagaaaacauuucugcagucaggugagacuuuuaca
aaagaggagaaaaucaaugccuc 318 9210 premiR_09210
cugaucauaguauucugucagauaaugccuaagaaugaccgcugaa
gaacguugacccauuugaguacccggucucagucgucauuuuuaag
uccagugagcauugugguaguuguucuuagauugcaguuucuuaug
uuuugaguuugaaguugauuuuca 320 9113 premiR_09113
aaagucuuagaauggaaaaaguaaagaaauaucaacuuccaaguug
gcaaguaacucccaaugauuuaguuuuuuuccccccaguuugaauu
gggaagcugggggaaguuaaauaugagccacuggguguaccagugc
auuaauuugggcaaggaaagug 322 9268 premiR_09268
cuccaggucaucaucagugugguauuaucuaugagaacuugagcga
cagaguauuucuugaugaauuuauagaucauuugagauguugaguu
acuuuuguuuuguuuucaaauagguagagacuauuaauguaaaaaa
acaagaaaggaaaaugaaaugugc 324 9114 premiR_09114
caccccgugccuuuugaucuagcacagacccuucaccccucaccuc
gaugcagccaguagcuuggauccuugugggcaugauccauaaucgg
uuucaagguaacgauggugucgaggucuuugguggguugaacuaug
uuagaaaaggccauuaauuugcc 326 9189 premiR_09189
cccucuggcaugguucauuagggccaauuaauguggcuggguuauu
ugcaacuuaaacugggggauaaugucgcuugagggagcguuuucgu
uuuaggaaauauuguuuugguuucggguuugaaggcagcugucaaa
aaagcggcauggaaauucauugg 328 9181 premiR_09181
gagcuaguaccuucuccccuuagcaacuuccucauucuaaaauggg
gguggcagaaccauuguuuggcuccaguuguccucagaaagguggc
uuccagaugccagugacugcuggugagugcaggcugcuucaguauu
uccuggccagcugacaagguguua 330 9278 premiR_09278
acauaaaaucuuaucuaugugcagcaugacucucuccaggugacag
aaagggcucuagacagcugagaggaccugaucauguagggagggac
ggggaggggagccaggacccaggagcugcauggcuguaagaggaag
guccuuggaggguaucagcagucuca 332 9188 premiR_09188
ggaaccugcuuggacaagucuucuggcucgaccucgacaugcucca
ucggaugaauuguugguguuagcccugcggccccacgcaccagggu
aagagagacucucgcuuccugcccuggcccgagggaccgacuggcu
gggccugccuucugcccagcucacc 334 9280 premiR_09280
cuucaggaguuggugguguugacugggagugaauugacggaaggga
ccaugggaauuuauauaucauuuugaaacuuaugaaaccuuuuguc
aaaguuucacuuucugacucaggcucaguccaggacauuguucaau uccccugguguaggcauca
336 9233 premiR_09233
ggucaacaaggugagucuggaugaggggcagggaugccaggcaagu
gagcaggucugggagucaggccuugcucaggcccuguucuucuccc
uugcagcuucugucuggccccaaagagaccccugcugcccagagcc
ccaccagaggccccucugacaccaaga 338 9224 premiR_09224
cuguguuguguccugacaccuccaaguucuagggccgucaggacac
gggaggguuuggggacagaguguccuuccucuguccucucauccca
guccugauggccgcuuggugagugucuggugcccugguggccugcc
ccagcucucuucuggcuuucugagcag 340 9130 premiR_09130
uuacaaugguucuaugaggacguggccccacaguaaguugaggagc
acuggguauguaugaauaaaauggcaugacaggccuucucuuucca
guucuucccagaauugggaaaagcuggguugagaggguaagaaaag
aaaaacaaauaaauuuuuuaaa 342 9195 premiR_09195
gcgagacucuuuuuuucuccaggaccugcggagcagccaggcuuca
ugaguuaaaugcagaucugaaccauacccaguugggauugggguac
acacucuacuccucugaaaacuagcuagggguucgaacuuggugag agggagagugggacagagc
344 9213 premiR_09213
aaguucugagaguccaggaggcagaggcugggguuugggggauguc
agagggcaaaucuggggcuuggggggcccaggaagcagagaugaag
guuuuagagucuccagagaacaaaucugguacuuuuaaggcccagg
aagcggaggcuggggucuugggaaa 346 9173 premiR_09173
aauuugaaaauccauauaaagguacguccacauuuauguuauuaug
agugagucauauuggugaagucaggacaauggccugucauuagauc
uuugauucuuguuugcagugaagggagaugugaagaagccauguuc
ucugaacgugcugcuuggaggacu 348 9253 premiR_09253
cgguguuuccggccgccgucgcuguccagggaggcugaggcgagag
guagcuguccggguggggagcccgcacuaccuucuuccucuuccuc
cuccuccuccgggugaggggagcgaagguuggggguccccgagccc auggaccaggaggaggcgga
350 9214 premiR_09214
uuuuuuuuuuaacaccuauauaucacccauugaacggguauuuuac
ugaacacaguacaguagacuguuuaaaacucacauccugguaacuu
ucacuacuugaaauuacaaagugcuuuuguuaauugcauauuuuug cucagccaucuuagaauugu
352 9223 premiR_09223
aagcuggcccuggcuggagauggcuagccccugagacaugcacuuc
ugguuuugaaaugacucugucuguggggcagcagaaacuagagaag
gcaaguggcugccccaccccaaggcgugaccaggaggaacagccug
cagcucacuccaugccacacggg 354 9217 premiR_09217
uuggugguguguauaagaauguuucuugcuaauugaggauguguga
gguuuaaggcugugagcugaucuuugaaaaauaguuuccuguuucu
aaagugacauuacccaguauuugcuuacugcuuugugccuuaucuc
ccgcuuucuuuuuaguauuucug 356 9265 premiR_09265
gaaagagaaagccaagauccacuaccggaagaagaaacagcucaug
aggcuacggaaacaggccgagaagaacguggagaagaaaauugaca
aauacacagagguccucaagacccacggacuccuggucugagccca
auaaagacuguuaauuccucaaa 358 9235 premiR_09235
ugggcgucuacaacggcaagaccuucaaccagguggagaucaagcc
cgagaugaucgaccacuaccugggcgaguucuccaucaccuacaag
cccauaaagcacggcgggcccggcaucggggccagccacuccuccc
gcuucaucccucucaagcaguggcuca 360 9174 premiR_09174
guagcccacauggauagcacaguugucagacaagauuccuucagau
uccgaguugccuaccgguuguuuucguuguuguuguuguuguuuuu
cuuuuucuuuuuuuuuuugaagacagcaauaaccacaguacauauu
acuguaguucucuauaguuuuac 362 9311 premiR_09311
ggcggcgggagaaguagauugaagccaguugauuagggugcuuagc
uguuaacuaaguguuuguggguuuaagucccauuggucuaguaagg
gcuuagcuuaauuaaaguggcugauuugcguucaguugaugcagag
ugggguuuugcaguccuuagcuguug 364 9266 premiR_09266
ucauucugaggucacauaacacauaaaauuaguuucuaugagugua
uaccauuuaaagaauuuuuuuuucaguaaaagggaauauuacaaug
uuggaggagagauaaguuauagggagcuggauuucaaaacgugguc
caagauucaaaaauccuauugauagu 366 9132 premiR_09132
guaagaugguuaucgauaguaguuaaugauggaugaagugcacuga
ggcucuuaaaagauacuuaggauuuuugacuuuacucuguagguuc
uaaaguaaacauauaugagguuuuuaauuucucagauacuauaccu
gcagcucuuuuugcugacucaagau 368 9293 premiR_09293
agcacaaguacacacauaaaaacauuaggucaagguguagcccaug
agguggcacgaaaugggcuacauuuucuauguccagaaaaucucac
aacauccuuuaggaaaucuaagggcucaaggaggauuuagcaguaa
accaagagcagagugcuugguugaa 370 9138 premiR_09138
gucacucaggacagacuucuugaaguagguggcccuucaacugagc
ugaagggaagagaagggcauugcaggcugagggaugaucccagggu
gccccccagaucuaaaguguggagucugucaucgaggugcguagcc
ucuccagaggugucacuguguuccau 372 9169 premiR_09169
cugcaaacagccuuuccacugacgcagugccuugggggcucugcca
agcgaccccuagaauggggauuguggggggucgcucuaggcaccgc
agcacugugcuggggauguugcagcugccugggagugacuucacac
aguccucucugccuccagggucaccc 374 9165 premiR_09165
guggauaugagugaagacggggcaggcaggccacaucucuuagaag
aggaaggugauugccacgucuccuuccuccaugcugauggcaaggc
gugcgggcuguguucucuugcagccagcgucccaugcucgguggcc
ccagaaaagucaguguguaggccu 376 9196 premiR_09196
aggaaacacaagaagaaacgccuggugcagagccccaauuccuacu
ucauggaugugaaaugcccaagaugcuauaaaaucaccacggucuu
uagccaugcacaaacgguaguuuuguguguuggcugcuccacuguc cucugccagccuauaggag
378 9209 premiR_09209
cauagauauguauucagcuugucuucaaauacggccaagcagaaaa
uguuuuauauuuuauaaaucaucuuuugacucuguauuuaaauucu
augauacugaaaauaaaggcauucuggaaaaauacugacugauuuu
ggugcagaaguuuugaguaucaag 380 9310 premiR_09310
aauuaaaguggcugauuugcguucaguugaugcagagugggguuuu
gcaguccuuagcuguugcagaaauuaaguauugcaacuuacugagg
gcuuugaaggcucuuggucuguauuuaaccuaaauuucuauaagau
uauuaguauaaaaggggagauagguag 382 9302 premiR_09302
ccaggagaggaaaaggaaaugggaccucagaaguagaagccucagg
gaaggaguaaaguagaaaucagaagaaaagaagcuucacuugauag
uaauaagguuuuuaacuucaaguaccuucagaaaaugugauuuuga
uaagaggaaagggcaaauuuag 384 9222 premiR_09222
agccugugaccccucgggacugccuggugcaggugguggcagcugg
agggacccaugcagcacccaggucagagcagacccuccccugccgg
ccugcgccagcuggaccugauggcccccuguggcgccuugaccugc
ugggccaggcugcccugggacucuc 386 9150 premiR_09150
ggccccgcauccgcguucgucuaggcgcucuugucaccucgccaug
ccggagccaucgcgggcggcuccggcuucuaaaaagggcuccaaga
aggccauuaccaaggcgcagaagaaggacggcaagaagcgcaagcg
cggccgcaaggagagcuauucuauc 388 9119 premiR_09119
gaaacaucuuaaugcacagccacaaguuacaaugcaacagccugcu
gcucauguacaaggucaggaaccuuugacugcuuccauguugguau
cugcccaugcucaagagcaaaagcaaauguugggugaacggcuguu
uccucuuauucaagccaugcaccuu 390 9257 premiR_09257
ugaaacuaacuagauuuauuggauaucaguacuuugagaguuagaa
augguuacugauugucaucuuuucagugaaggguucuauaguugag
uaaaaaauuugucuaacuuuguaaguauaguuuauauuguagaaau
ugcuuccaauuuuguugguaacuucau 392 9170 premiR_09170
cugcaaacagccuuuccacugacgcagugccuugggggcucugcca
agcgaccccuagaauggggauuguggggggucgcucuaggcaccgc
agcacugugcuggggauguugcagcugccugggagugacuucacac
aguccucucugccuccagggu 394 9277 premiR_09277
cccaggguggcauaggagaaaaaggugcugaaggcacagcuggaaa
ugauggugcaagaguaagugaaaguauuccuuuucuagcugggcua
ggaaccaacauuacaguaucauaugaguuucccugaaccuuccaaa
auaaagucagucauguuuccuuggua 396 9250 premiR_09250
acaugcagggugaaauccucacuauuuuuaugaaacagcaucugau
cuugaacuuuuaugacucaccucagucacuucaccuguauuuuggc
ccugucagaucaugucuuuauuuaaacuuuugauauuuuauucuuu
auauguuuuugcauuaguuuuuauuuu 398 9202 premiR_09202
aaacaauaaaaaacuggcugcuaucgaagcccuaaaugauggugaa
cuccagaaagccauugacuuauucacagaugccaucaagcugaauc
cucacuuggcccuuuuguaugccaagagggccagugucuucgucaa
auuacagaagccaaauacugccau 400 9230 premiR_09230
cugguuauaucaggauaaauucauaaaggguuuuguguguguguuu
guuuuuguuguuguuguuuaggguuuuuuuuuuuuaaacagggucu
ugcuuuguugcccaggaugaaaugcaaucacacacaaucauggcuc
auugcaucacuaucuauguauuca 402 9215 premiR_09215
ucaguuaagccaauacauuuaaaguuuugcaugaggaacacugacu
uuauuaagcauuuucagauguggugguuguauuuuugccccaagaa
guguuuggauaaccacacaaaagcaugaugaaaaggcuucuuguag
ucccauaauuucuugugaacuaauguu 404 9297 premiR_09297
uucugaaggguauauugcauguucauuauuaaucugagugagaggu
uaguugcuauaaaguuacaagauuuuccugaucacuuaaaauuuuc
uuguagucagagauuugacucugaaucgucacuucaaaaauuuguc
uuuucagguacuauguaaagagaacu 406 9286 premiR_09286
cacuacggccucuggccaugcaccaguuguaguuuuguguguuggc
ugcuccacuguugucugccagcccacaggagggaaagugaggcucc
uggaaggacgcuccuucagauggaagcagcacuggaagagccccaa
guugaggugcaugggacacaaacu 408 9110 premiR_09110
auaaugaguaaaaauguucaucuuuaauaaagcaaaaauagagcaa
cccaccaaauaguuaacacuugccuggagagauuuaggaacaccag
uauuccacugagauugcugagagucucaggaaagaaaggucuaacu
uaaauuguauuuuaccauuucugag 412 9313 premiR_09313
gcaaagaaagaagaaucugaggagucugaugaugacaugggcuuug
gucuuuuugacuaaaccucuuuuauaauguguucaauaaaaagcug
aacuuuaaaaaaaagauugggguuuaucauguaauuguuucauuuu
guuguauucuguguuaagauuucuaa 414 9314 premiR_09314
ccggccgcugggcgcacccgucccguucguccccggacguugcucu
cuaccccgggaacgucgagacuggagcgcccgaacugagccaccuu cgcg 415 9315
premiR_09315 ucucggaagcuaagcagggucgggccugguuaguacuuggacggga
gaccgccugggaauaccgggugcuguaggcuuuuucuuuggcuuuu ug 417 9316
premiR_09316 ucuuggaagcuaagcagggucgggccugguuaguacuuggauggga
gaccaccugggaauaccgggugcuguaggcuuuggccgggcguggu gg 419 9317
premiR_09317 ggcguaggggggccggccugcugugaugacauuccaauuaaagcac
guguuagacugcugacgcgggugaugcgaacuggagucugagccug cccgagcggagc 419
9318 premiR_09318 cacugucacugccuaccaaucucgaccggaccucgaccggcucguc
uguguugccaaucgacucggcguggcgucggucgugguagauaggc ggu 419 9319
premiR_09319 cacugucacugccuaccaaucucgaccggaccucgaccggcucguc
uguguugccaaucgacucggcguggcgucggucgugguagauaggc ggu 423 9320
premiR_09320 cacugucacugccuaccaaucucgaccggaccucgaccggcucguc
uguguugccaaucgacucggcguggcgucggucgugguagauaggc ggu 425 9321
premiR_09321 ugcugcagguguuggagagcaguguguguugccuggggacugugug
gacugguaucacccagacagcuugcacugacuccagacccugccgu caug 427 9322
premiR_09322 cgccccgggccgcggcuccugauuguccaaacgcaauucucgaguc
uauggcuccggccgagaguugagucuggacgucccgagccgccgcc cccaa 427 9323
premiR_09323 aguaccaagaaguuaucauuuccauaugacugucauugcuuaaaac
uagcuaguaugagcaggacgguggccauggaagucgaaauucgcua ag 433 9324
premiR_09324 aguaccaagaaguuaucauuuccauaugacugucauugcuuaaaac
uagcuaguaugagcaggacgguggccauggaagucgaaauucgcua ag 433 9325
premiR_09325 gggauugacagauugacagcucuuucucgauucugugggugguggu
gcauggccauucuuaguugguggagugauuugucugguuaauucug auaa 433 9326
premiR_09326 gggauugacagauugacagcucuuucucgauucugugggugguggu
gcauggccauucuuaguugguggagugauuugucugguuaauucug auaa 433 9327
premiR_09327 gggauugacagauugacagcucuuucucgauucugugggugguggu
gcauggccauucuuaguugguggagugauuugucugguuaauucug auaa 435 9328
premiR_09328 gggauugacagauugacagcucuuucucgauucugugggugguggu
gcauggccauucuuaguugguggagugauuugucugguuaauucug auaa 437 9329
premiR_09329 ugcaacugccgucagccauugaugaucguucuucucuccguauugg
ggagugagagggagagaacgcggucugagugguuuuuccuucuuga ug 439 9330
premiR_09330 auacguguaauugugaugaggauggauagcaaggaagccgcuccca
ccugacccucacggccuccguguuaccuguccucuaggugggacgc uc 441 9331
premiR_09331 gggugugagaagccggauccuguggugacccaguggccuaauggau
aaggcaucagccuccggagcuggggauuguggguucgagucccauc ugg 443 9332
premiR_09332 ccgggagcggggaggcggcggcuccagggaccuggcggccgccgau
cggggcugcgaggccccauggcgccgcccccagccccgcuccuggc gccga 445 9333
premiR_09333 gggccgggaaugccgcggcggggacggcgauugguccguaugugug
gugccaccggccgccggcuccgccccggcccccgccccacacgccg cau 446 9334
premiR_09334 ugguggggggagccgcggggaucgccgagggccggucggccgcccc
gggugccgcgcggugccgccggcggcggugaggccccgcgcgugug ucccggcugcgg 448
9335 premiR_09335 ggggaggagacgguuccgggggaccggccgcgacugcggcggcggu
gguggggggagccgcggggaucgccgagggccggucggccgccccg ggugccgcgcgg 450
9336 premiR_09336 acugucacugccuaccaaucucgaccggaccucgaccggcucgucu
guguugccaaucgacucggcguggcgucggucgugguagauaggcg gucaug 453 9338
premiR_09338 aaugcagugugauuucugcccagugcucugaaugucaaagugaaga
aauucagagaagccuggguagccgggcgugguggcucacaccugua aucccagcac 455 9339
premiR_09339 augaauuguugguguuagcccugcggccccacgcaccaggguaaga
gagacucucgcuuccugcccuggcccgagggaccgacuggcugggc cugccuu 457 9340
premiR_09340 cuucuccccagccagagguggagccaagugguccagcgucacucca
gugcucagcuguggcuggaggagcuggccuguggcacagcccugag u 459 9341
premiR_09341 gcacccagaucagugcuuggcaccuagcaagcacucaguaaauauu
uguugagugccugcuaugugccaggcauugugcugagggcuuugug ggga 461 9342
premiR_09342 aguugguagagggcagagggaugagggggaaaguucuauaguccug
agaucuaauuacaggacuauagaacuuucccccucaucccucuacc cuua 463 9343
premiR_09343 gugcugcagguguuggagagcaguguguguugccuggggacugugu
ggacugguaucacccagacagcuugcacugacuccagacccugccg ucaug 465 9344
premiR_09344 gagggcgggagacaaaaucucgcaauucugaccugccuuuggacau
aauugaggcuuuaugaggaagguggggaugcgggaguggcgauccc 465 9345
premiR_09345 ggcguugcuuggcugcaacugccgucagccauugaugaucguucuu
cucuccguauuggggagugagagggagagaacgcggucugaguggu uuuuccuucuuga 465
9346 premiR_09346 ggcguugcuuggcugcaacugccgucagccauugaugaucguucuu
cucuccguauuggggagugagagggagagaacgcggucugaguggu uuuuccuucuuga 465
9347 premiR_09347 ggcguugcuuggcugcaacugccgucagccauugaugaucguucuu
cucuccguauuggggagugagagggagagaacgcggucugaguggu uuuuccuucuuga 465
9348 premiR_09348 ggcguugcuuggcugcaacugccgucagccauugaugaucguucuu
cucuccguauuggggagugagagggagagaacgcggucugaguggu uuuuccuucuuga 467
9349 premiR_09349 ggcguugcuuggcugcaacugccgucagccauugaugaucguucuu
cucuccguauuggggagugagagggagagaacgcggucugaguggu uuuuccuucuuga 467
9350 premiR_09350 ugugagacgaauuuuugagcggguaaaggucgcccucaaggugacc
cgccuacuuugcgggaugccugggaguugcgaucugcccgaccuua uuca 470 9351
premiR_09351 ugugagacgaauuuuugagcggguaaaggucgcccucaaggugacc
cgccuacuuugcgggaugccugggaguugcgaucugcccgaccuua uuca 470 9352
premiR_09352 gugaacggcggcugguggcgaguuccgcugugccagcuuccguugg
cguuugccaucggugcaugggugguucagugguagaauucucgccu g 470 9353
premiR_09353 gugaacggcggcugguggcgaguuccgcugugccagcuuccguugg
cguuugccaucggugcaugggugguucagugguagaauucucgccu g 470 9354
premiR_09354 gugaacggcggcugguggcgaguuccgcugugccagcuuccguugg
cguuugccaucggugcaugggugguucagugguagaauucucgccu g 474 9363
premiR_09363 gugaacggcggcugguggcgaguuccgcugugccagcuuccguugg
cguuugccaucggugcaugggugguucagugguagaauucucgccu gccacgcgggaggc 476
9364 premiR_09364 ccaucaccacugugaaucagagcaacaaaacagcuggaggcagaac
agcacucagcuggagcauuggugguucagugguagaauucucgccu gc 476 9365
premiR_09365 cgaggaaguagugaccugccacuggccaccugcggaaccagaguuc
cccacuggagggccgcguuggugguauaguggugagcauagcugcc uu 479 9367
premiR_09367 cgaggaaguagugaccugccacuggccaccugcggaaccagaguuc
cccacuggagggccgcguuggugguauaguggugagcauagcugcc uuccag 481 9368
premiR_09368 ccgacccgggccugggcuguggcugugacuggcgcugccgugggcg
ccgcagcccucgcgggagccggacgcgguaaugccccagcggcgca gc 483 9369
premiR_09369 guaccaagaaguuaucauuuccauaugacugucauugcuuaaaacu
agcuaguaugagcaggacgguggccauggaagucgaaauucgcuaa gg 483 9370
premiR_09370 cugcugagaguagguggggauguagcucagugguagagcgcaugcu
uugcauguaugaggccccggguucgauccccggcaucuccagugua gu 485 9373
premiR_09373 cugcugagaguagguggggauguagcucagugguagagcgcaugcu
uugcauguaugaggccccggguucgauccccggcaucuccagugua gu 487 9374
premiR_09374 cugcugagaguagguggggauguagcucagugguagagcgcaugcu
uugcauguaugaggccccggguucgauccccggcaucuccagugua gu 489 9375
premiR_09375 cugcugagaguagguggggauguagcucagugguagagcgcaugcu
uugcauguaugaggccccggguucgauccccggcaucuccagugua gu 491 9376
premiR_09376 gccgacagccagcuggaucuccugucccgagcccuggguacugggg
gugccccugaguuggguucccucagaccuggugaucgggcccugga g 491 9377
premiR_09377 cucccucuccucccccgggguucggugcgcggccggggccggaguu
cgcugcaagucggcggaaaguuuggcugcgcggguucccccgaagu ucaggugcg 491 9378
premiR_09378 gucugaucucagaagcuaagcaaggucgggucuaguuaguacuugg
augggagacugccuggaauaccgggugcuguaggcuuuuggccuau cguucccu 491 9379
premiR_09379 agagagccccggggugcagauccuugggagcccuguuagacucugg
auuuuacacuuggagugaacgggcgccaucccgaggcuuugcacag gggcaa 496 9380
premiR_09380 agagagccccggggugcagauccuugggagcccuguuagacucugg
auuuuacacuuggagugaacgggcgccaucccgaggcuuugcacag gggcaa 496 9381
premiR_09381 agagagccccggggugcagauccuugggagcccuguuagacucugg
auuuuacacuuggagugaacgggcgccaucccgaggcuuugcacag gggcaa 496 9382
premiR_09382 agagagccccggggugcagauccuugggagcccuguuagacucugg
auuuuacacuuggagugaacgggcgccaucccgaggcuuugcacag gggcaa 500 9383
premiR_09383 ggauugacagauugauagcucuuucucgauuccgugggugguggug
cauggccguucuuaguugguggagcgauuugucugguuaauuccga uaac 500 9384
premiR_09384 ggauugacagauugauagcucuuucucgauuccgugggugguggug
cauggccguucuuaguugguggagcgauuugucugguuaauuccga
uaac 503 9385 premiR_09385
ggauugacagauugauagcucuuucucgauuccgugggugguggug
cauggccguucuuaguugguggagcgauuugucugguuaauuccga uaac 503 9386
premiR_09386 ggauugacagauugacagcucuuucucgauucugugggugguggug
cauggccauucuuaguugguggagugauuugucugguuaauucuga uaa 503 9387
premiR_09387 ggauugacagauugacagcucuuucucgauucugugggugguggug
cauggccauucuuaguugguggagugauuugucugguuaauucuga uaa 503 9388
premiR_09388 gcaacugccgucagccauugaugaucguucuucucuccguauuggg
gagugagagggagagaacgcggucugagugguuuuuccuucuugau g 503 9389
premiR_09389 gcaacugccgucagccauugaugaucguucuucucuccguauuggg
gagugagagggagagaacgcggucugagugguuuuuccuucuugau g 505 9390
premiR_09390 gcaacugccgucagccauugaugaucguucuucucuccguauuggg
gagugagagggagagaacgcggucugagugguuuuuccuucuugau g 506 9391
premiR_09391 gcaacugccgucagccauugaugaucguucuucucuccguauuggg
gagugagagggagagaacgcggucugagugguuuuuccuucuugau g 508 9392
premiR_09392 gcaacugccgucagccauugaugaucguucuucucuccguauuggg
gagugagagggagagaacgcggucugagugguuuuuccuucuugau g 510 9393
premiR_09393 aucuuggaagcuaagcagggucgggccugguuaguacuuggauggg
agaccaccugggaauaccgggugcuguaggcuuuggccgggcgugg ugg 512 9394
premiR_09394 aucucggaagcuaagcagggucgggccugguuaguacuuggacggg
agaccgccugggaauaccgggugcuguaggcuuuuucuuuggcuuu uug 514 9395
premiR_09395 auccucccuggggcauccuguacugagcugccccgaggcccuucau
gcugcccagcucggggcagcucaguacaggauacucggggugggag uca 516 9396
premiR_09396 acaucaagugacugugcuuggcuguggggcuaccaagaugaagaag
gaaugcuccugcccucgaggagcucacagucuagugggagggaaca augc 516 9397
premiR_09397 ggggcucgcggacccggcccagagggcggcgguggcggcagcuacu
uuucuggucagggcucggacaccggcgcgucucucaagcucgccuc uuc 519 9398
premiR_09398 uaaugcagugugauuucugcccagugcucugaaugucaaagugaag
aaauucagagaagccuggguagccgggcgugguggcucacaccugu aa 521 9399
premiR_09399 guuccuuguguucuuccaguccgcccucugucaccuugcagacggc
uuucucuccgaaugucugcaagugucagaggcgaggaguggcagcu gcau 523 9400
premiR_09400 guuccuuguguucuuccaguccgcccucugucaccuugcagacggc
uuucucuccgaaugucugcaagugucagaggcgaggaguggcagcu gcau 523 9401
premiR_09401 aacauuaggagaguaucuucucuguuuuggccauguguguacucac
agccccucacacauggccgaaacagagaaguuacuuuccuaauauu ugccu 525 9402
premiR_09402 cugccugcuucugugugauauguuugauauuggguuguuuaauuag
gaaccaacuaaaugucaaacauauucuuacagcagcaggugauuca gcacc 527 9403
premiR_09403 ccgggccgcggcuccugauuguccaaacgcaauucucgagucuaug
gcuccggccgagaguugagucuggacgucccgagccgccgccccca aaccuc 529 9404
premiR_09404 ccgggccgcggcuccugauuguccaaacgcaauucucgagucuaug
gcuccggccgagaguugagucuggacgucccgagccgccgccccca aaccuc 531 9405
premiR_09405 guugugcuguccaggugcuggaucagugguucgagucugagccuuu
aaaagccacucuagccacagaugcagugauuggagccaugacaagu cccca 533 9406
premiR_09406 accuaccuaacuggguuagggcccuggcuccaucuccuuuaggaaa
accuucuguggggaguggggcuucgacccuaacccaggugggcugu aac 535 9407
premiR_09407 cauuaggagaguaucuucucuguuuuggccauguguguacucacag
ccccucacacauggccgaaacagagaaguuacuuuccuaauauuug ccucc 537 9408
premiR_09408 uaccucccagcuguguucugcccagugcucugcugacuggacgccu
cuguguccuuagaccaugugcagggccugaugcaggaaagagcuga
uuaaugcagugugauuucugcccagugcucugaaugucaaagugaa 537 9409
premiR_09409 gaaauucagagaagccuggguagccgggcgugguggcucacaccug uaa 539
9410 premiR_09410 accccuagaauggggauuguggggggucgcucuaggcaccgcagca
cugugcuggggauguugcagcugccugggagugacuucacacaguc cucucug 539 9411
premiR_09411 ucguccagcgagggcgcgcuggcccugggcagcguguggcugaagg
ucaccauguucuccuuggccauggggcugcgcggggccagcagguc cacg 542 9412
premiR_09412 ucguccagcgagggcgcgcuggcccugggcagcguguggcugaagg
ucaccauguucuccuuggccauggggcugcgcggggccagcagguc cacg 545 9414
premiR_09414 cagcuccgcagggguuuggggaaacggccgcugagugaggcgucgg
cuguguuucucaccgcggucuuuuccucccacucuuggcugguugg acc 547 9415
premiR_09415 gauugacagauugauagcucuuucucgauuccgugggugguggugc
auggccguucuuaguugguggagcgauuugucugguuaauuccgau aa 551 9416
premiR_09416 gauugacagauugauagcucuuucucgauuccgugggugguggugc
auggccguucuuaguugguggagcgauuugucugguuaauuccgau aac 552 9417
premiR_09417 ccggcggggcgaagcccgcggcugcuggacccacccggccgggaau
agugcuccugguuguuuccggcucgcgugggugugucggcggcggg gcc 553 9418
premiR_09418 ccugagcaaggucacaaagcugggugagaggggcugggauucacau
ucaagcacuguguuccuaggccccuugaccccuggcaucugugggg 554 9419
premiR_09419 cugaaaugucuucaaauguaucaauaagccuucucuucccaguucu
ucuuggagucaggaaaagcuggguugagaggagcagaaaagaaaaa 556 9423
premiR_09423 ucuucuaccuaagaauucugucucuuaggcuuucucuucccagauu
ucccaaaguugggaaaagcuggguugagagggcaaaaggaaaaaaa a
TABLE-US-00005 TABLE 5 miRNA Sequences Seq ID MiR name
mature_sequence 1 miR_1966 CGAGCCUGGUGAUAGCUGGUUGUCCAAG 3 miR_1967
GAGGCUGAGGCGAGAGGU 5 miR_1968 AUUUGUGGCCGAGUGUAACAACC 7 miR_1969
UCCCUUCGUGGUCGCC 9 miR_1970 GCAGGGGAAGAAGCCUUCCGU 11 miR_1971
ACUUUGAGAGUUAGAAAUGGUUACU 13 miR_1972 AACCAAUGAUGUAAUGAUUCUGCC 15
miR_1973 AUGAUUCUGUGACGCCAGCU 17 miR_1974 GAAAGCUGAGCGUGAACGUGGU 19
miR_1975 UAGCAGCACAUAAUGGUUUGAAU 21 miR_1976 UGUUUAGACGGGCUCACAU 23
miR_1977 GGAAAUUUGAGACCAGCAAGUACU 25 miR_1978
AUUGUAUAUCAGCAUGGGGAUUAUU 27 miR_1979 UUGGAGGCGUGGGUU 29 miR_1980
AACGUGCAGCGGCUGAAGGAGU 31 miR_1981 AAUUGUGAGGCUUGAGUGU 33 miR_1982
UGAUAGGAUUGACAUGGAGCAC 35 miR_1983 AACGGCCGCGGUACCCUAAC 37 miR_1984
UCUUGCCAAGAGAAUUAAUGUGCGU 39 miR_1985 AAUCUGAGUGAGAGGUUAGUUGCU 41
miR_1986 AUUAAAAAUUUCGGUUGGG 43 miR_1987 GAUAGCUGCCAGUGACAGGAGUAGU
45 miR_1988 GAGCCUGGUGAUAGCUGG 47 miR_1989 UGGCGCGGAGCGGAGCGG 49
miR_1990 AGGCGGCGGAGGGGCG 51 miR_1991 AUGCGUGCGAGAAGUCAGUGG 53
miR_1992 UGUGUUUGAGAGCAACGCCAUUGCCU 55 miR_1993 GCAUGAGUGGUUCAGUGGU
57 miR_1994 AAGGAAUGAGUUAGCUUUG 59 miR_1995 ACAAGUGAAUACACUGAGGC 61
miR_1996 CUCGCGCCCGCGUCGCGGCAGC 63 miR_1997 GUGGUGUUGAGGAAAGCAGAC
65 miR_1998 GUUUUUGUGACUGCCUUGAGU 67 miR_1999
AAGGCACAGCUGGAAAUGAUGGUG 69 miR_2000 AAACUAGGCGGCUAUGGUAU 71
miR_2001 GUGGUGGUCGUACGCUGUG 73 miR_2002 GCUGAGUGAAGCAUUGGACUGU 75
miR_2003 AGGUUCACAUGGAAAAGGUU 77 miR_2004 CCAUUGUGUAGCAAGAUGUCAU 79
miR_2005 GGAAAUAGUUUGUGACAAACUGGG 81 miR_2006 CUCCUCUUCGUCUCCUCGGUC
83 miR_2007 ACUCCAGCCCCACAGCCUCAG 85 miR_2008
GCCGCGAGUGGGAGCGGGAGCG 87 miR_2009 GCUUGACUGAGUGUGGCUGGACGUG 89
miR_2010 AGUCGGUGCCUGAGGUUGC 91 miR_2011 AGUGAUGAGGAUGUGCUGAU 93
miR_2012 AUUGAGUGGGGCUCAGGAUU 95 miR_2013 GCAUGGGUGGUUCAGUGU 97
miR_2014 AACUGUUAUAUUAUGAUUGUGAC 99 miR_2015
UUGCUGUGAUGACUAUCUUAGGAC 101 miR_2016 AGAAUUGAGUGAUCUCAUGGAU 103
miR_2017 AAAACGAUCUUUCAGAUUUAGAGU 105 miR_2018 GCGGUCGGGCGGCGGCG
107 miR_2019 AUGAGAACUUGAGCGACAGAGU 109 miR_2020 AUUGGUCGUGGUUGUAGU
111 miR_2021 GAUGAGAGAACAGUGGGUACUUC 113 miR_2022
CAAGGUGUAGCCCAUGAGGUGGC 115 miR_2023 AGCACCAGCCUAGGAAGAGGGU 117
miR_2024 ACUUUAACACUGCUGUGGAAGGC 119 miR_2025
AUUUGACAAGAGUAUGCCAGGUGU 121 miR_2026 CUGGGAGCUUGAAAGGAG 123
miR_2027 AUUUGGGCAGGUUGAAAGAAUUU 125 miR_2028
GUGCUGGAGGCCAGGCUGAGGCCC 127 miR_2029 GCUGUUGGUGGAGAAGGU 129
miR_2030 AUGGCAGUUGGAGAGAAAGAAC 131 miR_2031
GCAGAUAAACUCAUGCCAGAGAACU 133 miR_2032 AGUGCCAGGUGGGGAGG 135
miR_2033 GACUCUUAGCGGUGGAUC 137 miR_2034 CUGUUGCGGGACCCGGGGUGU 139
miR_2035 UUAAAGCUGCCAUUUGUUACU 141 miR_2036 GAGCAGAGGCGAUAGUUGAAGU
143 miR_2037 GAUGACAUGGGCUUUGGUCUUUUU 145 miR_2038
AGAAAGGGCCUUGUGUUU 147 miR_2039 ACUCGGCGUGGCGUCGGUCGUGGUAG 149
miR_2040 GUGUUUAGUGAGUAUUUGUU 151 miR_2041 ACUGCUGCUGCUGCUUGGCC 153
miR_2042 UGCAGAGUGGGGUUUUGCAGUCCUU 155 miR_2043
AUGACCACCAAACCCAGGAGC 157 miR_2044 CAGAGUCUGUAGAAGAGGCG 159
miR_2045 CCUUGACUUCUGCCAGAGU 161 miR_2046 AUCACUGACUGAUCAAGUAGAGGU
163 miR_2047 AAGGAUUGGACAGGGUUAGAUU 165 miR_2048
AGGCCAAGGCUGCGGGGUU 167 miR_2049 AACGGGAGGCGCUAGCCAUGG 169 miR_2050
ACCUUUCAGUGCAGUUUCUUUU 171 miR_2051 AUUUUGGGUGGAAGAGGCAU 173
miR_2052 GGAAUGAGGAGCUUUGAC 175 miR_2053 AAAAGCUGGGUUGAGAGGAU 177
miR_2054 AGUAAGGUCAGCUAAAUAAGCU 179 miR_2055 GAAUUGACGGAAGGGAC 181
miR_2056 UUGUGUCUUGUGUCUUUU 183 miR_2057 UCCCUGGUGGUCUAGU 185
miR_2058 UGAGGCUGUAACUGAGAGAAAGAUU 187 miR_2059
UUAGGGCCCUGGCUCCAUCUCCU 189 miR_2060 GAGUGUGAGUGGACGCGUGAGUGU 191
miR_2061 AUUGAGUCUGGCAGUCCCUGUU 193 miR_2062 AAAAUGUUUAGACGGGCUCAC
195 miR_2063 GAAUGUUUAUGGCACCUGAC 197 miR_2064
GGAAAUUUGAGACCAGCAAGU 199 miR_2065 ACAGUAGGGCCUUUGGAGUGAU 201
miR_2066 CUUGAAGUCUGGUGAUGCUGCCAUU 203 miR_2067 AGAUGAGCUGAAGGG 205
miR_2068 AAAAAGGGAGCCAAGAAG 207 miR_2069 AGUUUUGUGUGUUGGCUGCUCC 209
miR_2070 GGUUUAGUGAGCAGAGUU 211 miR_2071 AAGACGAGAAGACCCUAUGGAGCUU
213 miR_2072 AGCAGGGUGCAGGCUUGGAGUC 215 miR_2073
AUGUUGGGUUGUUACAGAGU 217 miR_2074 ACUGAGUUGACUGUUCCCUU 219 miR_2075
ACUGGGCAGUGACAAGCACGAU 221 miR_2076 AGAAAGCGUGAGUGUCCAGAGCCU 223
miR_2077 GCUUGAGGGCAGUUGGUGCGG 225 miR_2078
CCGAGCCUGGUGAUAGCUGGUUGUC 227 miR_2079 UGGAGAUGGCUGGCAGAAUGGUUCU
229 miR_2080 AAGUAGAAGCCUCAGGGAAG 231 miR_2081 AUGGUUCUGGACAGUGGAUU
233 miR_2082 GAUGAGUCAGGCUAGGCU 235 miR_2083 AUGUGAGAGCAGCAGAGGCGGU
237 miR_2084 GUUUUAAGGACUUAAGGGUAU 239 miR_2085
UGAUAUGUUUGAUAUUGGGU 241 miR_2086 AGAAAGCCAGGAGCUGUGAUU 243
miR_2087 AUUCUAAGUCAGUCAGUCAUC 245 miR_2088
UUCUCUGUUUUGGCCAUGUGUGU
247 miR_2090 UUCUAAGCCAGUUUCUGUCUGAU 249 miR_2091
GGUUUUGACAUGUCACUGUU 251 miR_2092 CUCGCCCGUGGUCUCUCGUCUU 253
miR_2093 AACACGCAUACGGUUAAGGCAUUGC 255 miR_2094
UGAAACAGCAUCUGAUCUUGAACUU 257 miR_2095 GCGUAAAGAGUGUUUUAGAUCACCC
259 miR_2096 GUCCGUUUCCUGUCAGAGUGAUCC 261 miR_2097
CCAUCGGUGAUCCCAGUGACAAGU 263 miR_2098 GAAAUGUUGAGUGUUUACCCUGU 265
miR_2099 GAAGAAACAGCUCAUGAGGCU 267 miR_2100 AUGAACCACCAGUCCAAGAAUCU
269 miR_2101 GGAAGGUUGGGGGGUGU 271 miR_2102 UUCUAGGUUGUGGCAUUU 273
miR_2103 UGAUAUGUUUGAUAUUGGGUUGUU 275 miR_2104
AGAAUACAGCAGAAUUGGCCUC 277 miR_2105 CUGAUGGAGAGAAGAAGGCAU 279
miR_2106 ACUCGGCGUGGCGUCGGUCGUG 281 miR_2107 AUGAGGUGGCAAGAAAUGGGCU
283 miR_2108 AAUUUUGACAGAUGCUCAAGGCUGU 285 miR_2109
AAAAGCUGGGUUGAGAAG 287 miR_2110 AUUGAUGGUUAAGCUCAGCUUUU 289
miR_2111 AUGGAUUGAGAUGUGAUCAAAGGC 291 miR_2113
AUUUGGCAUUUGGAAGAUAGGUU 293 miR_2114 ACUGCUGAGGAACUGUCACUUGU 295
miR_2115 AGGUGAGGGGCAGGACCUGAAGGU 297 miR_2116 AGUGUCUGUGUGUGCUUGCU
299 miR_2117 ACGAAGAUGGCGACCGUAAC 301 miR_2118
AACUGGAAUGGCGGCAAGGUCCU 303 miR_2119 AGUUGAGUCAGGGCCUGUGUG 305
miR_2120 GCCGCCCGGGGCCAUGGCG 307 miR_2121 AAGUCUAAGUCUAACAUUCGGUGU
309 miR_2122 CGGCGGGAGCCCGGGG 311 miR_2123 ACGUGGAUGGCGUGGAGGUGC
313 miR_2124 GUUUAGACGGGCUCACAU 315 miR_2125 AUUUCUGCAGUCAGGUGAGAC
317 miR_2126 AAGAAUGACCGCUGAAGAACGU 319 miR_2127
AAAUAUGAGCCACUGGGUGU 321 miR_2128 AAGGUCCUCUGAGGCAGCAGGCU 323
miR_2129 GUCGAGGUCUUUGGUGGGUUG 325 miR_2130 GGUUUCGGGUUUGAAGGCAGC
327 miR_2131 GCUGGUGAGUGCAGGCUGCUUC 329 miR_2132
GACAGAGGUGGCAUCAAGCU 331 miR_2133 CUGCCCUGGCCCGAGGGACCGAC 333
miR_2134 UCGUGUCGCGUGGGGGGCGG 335 miR_2135
GGGAUGCCAGGCAAGUGAGCAGGUC 337 miR_2136 AGGGCCGUCAGGACACGGGAGGGUU
339 miR_2137 GAAAAGCUGGGUUGAGAGGGU 341 miR_2138 AACUAGCUAGGGGUUCG
343 miR_2139 GGGUUUGGGGGAUGUCAGAGGGC 345 miR_2140
UGAAGGGAGAUGUGAAGAAGCC 347 miR_2141 AGAAAGUCCAAGUGUUCAGG 349
miR_2142 UGAACGGGUAUUUUACUG 351 miR_2143 CUGAGACAUGCACUUCUGGUU 353
miR_2144 AAUUGAGGAUGUGUGAGGUUU 355 miR_2145
AGUUUCUAUGAGUGUAUACCAUUU 357 miR_2146 AGGUGGAGAUCAAGCCCGAGAUGAU 359
miR_2147 GAAGACAGCAAUAACCACAGU 361 miR_2148
GUUUCUGUUGAGUGUGGGUUUAGU 363 miR_2149 AUCGCCGUGGAGUGGGAGAGC 365
miR_2150 GGAUGAAGUGCACUGAGGCUCUU 367 miR_2151
UCCUGUUGGCCGAGUGGAGACUGGUGU 369 miR_2152 GUGGAGUCUGUCAUCGAGGUGCGU
371 miR_2153 UUGCAGCUGCCUGGGAGUGACUUC 373 miR_2154
UGCAGCCAGCGUCCCAUGCUCG 375 miR_2155 AGUUUUGUGUGUUGGCU 377 miR_2156
ACGGCCAAGCAGAAAAUGUUUU 379 miR_2157 GAUUAGGGUGCUUAGCUGUUAACU 381
miR_2158 CCUUCGAGGCGGCUGAGACCC 383 miR_2159 AGGUGGUGGCAGCUGGAGGGACC
385 miR_2160 AGAAGAAGGACGGCAAGAAGCGC 387 miR_2161
AAAAGCAAAUGUUGGGUGAACGG 389 miR_2162 GGCGGAGGGGCCGCGGGCC 391
miR_2163 UUGCAGCUGCCUGGGAGUG 393 miR_2164 UCUCUCCAGGUGACAGAAAGGGCU
395 miR_2165 CGUUCUUGCUCUGCCUCGGUC 397 miR_2166
AUGCCAAGAGGGCCAGUGUCUU 399 miR_2167 UUUUGUGUGUGUGUUUGUUUUU 401
miR_2168 AUGAGGAACACUGACUUUAUUAAGC 403 miR_2169
GGGUCGGAGUUAGCUCAAGCGGUU 405 miR_2170 GGAGGUUCAGAGUUGGAAG 407
miR_2171 GAGAGUCUCAGGAAAGAAAGGUC 411 miR_2364 ACCCGUCCCGUUCGUCCCCGG
413 miR_2365 ACCGGGUGCUGUAGGCUUU 416 miR_2366
ACGCGGGUGAUGCGAACUGGAGUCUGAGC 418 miR_2367 ACUCGGCGUGGCGUCGGUCG 420
miR_2368 ACUCGGCGUGGCGUCGGUCGU 421 miR_2369
ACUCGGCGUGGCGUCGGUCGUGGU 422 miR_2370 AGAGCAGUGUGUGUUGCCUGG 424
miR_2371 AGAGUUGAGUCUGGACGUCCCG 426 miR_2372 AGGACGGUGGCCAUGGAAG
428 miR_2373 AGGACGGUGGCCAUGGAAGU 429 miR_2374
AGUUGGUGGAGUGAUUUGUCU 430 miR_2375 AGUUGGUGGAGUGAUUUGUCUG 431
miR_2376 AGUUGGUGGAGUGAUUUGUCUGG 432 miR_2377
AGUUGGUGGAGUGAUUUGUCUGGU 434 miR_2378 AGAACGCGGUCUGAGUGGU 436
miR_2379 AUGAGGAUGGAUAGCAAGG 438 miR_2380 CGGAGCUGGGGAUUGUGGGU 440
miR_2381 CGGCGGCUCCAGGGACCUGGCG 442 miR_2382 CGGCGGGGACGGCGAUUGGU
444 miR_2383 CGGGGAUCGCCGAGGGCCGGUCGGCCGCC 447 miR_2384
CUCGGCGUGGCGUCGGUCGUGGU 449 miR_2385 CUGCCCAGUGCUCUGAAUG 451
miR_2386 CUGCCCAGUGCUCUGAAUGUCAAAGUG 452 miR_2387
CUGCCCUGGCCCGAGGGACCGACU 454 miR_2388 CUGGAGGAGCUGGCCUGU 456
miR_2389 CUUGGCACCUAGCAAGCACUC 458 miR_2390 GACUAUAGAACUUUCCCCCUC
460 miR_2391 GAGAGCAGUGUGUGUUGCCUGG 462 miR_2392 GAGGAAGGUGGGGAUGC
464 miR_2393 GAGUGAGAGGGAGAGAACGCGGUCUGAGUG 466 miR_2394
GAUGCCUGGGAGUUGCGAUCU 468 miR_2395 GAUGCCUGGGAGUUGCGAUCUGC 469
miR_2396 GCAUGGGUGGUUCAGUGG 471 miR_2397 GCAUGGGUGGUUCAGUGGU 472
miR_2398 GCAUGGGUGGUUCAGUGGUAGAAUUCUCGCC 473 miR_2399
GCAUUGGUGGUUCAGUGGU 475 miR_2400 GCGUUGGUGGUAUAGUGGU 477 miR_2402
GCGUUGGUGGUAUAGUGGUGAGC 478 miR_2403 GCUGUGGCUGUGACUGGCG 480
miR_2404 GGACGGUGGCCAUGGAAGU 482 miR_2405 GGGGAUGUAGCUCAGUGGU 484
miR_2406 GGUUCCCUCAGACCUGGU 486 miR_2407 GGAAAGUUUGGCUGCGCGGGUUCCCC
488 miR_2408 GGAAUACCGGGUGCUGUAGGCUUUU
490 miR_2409 GUGAACGGGCGCCAUCCCGAGGC 492 miR_2410
GUGAACGGGCGCCAUCCCGAGGCU 493 miR_2411 GUGAACGGGCGCCAUCCCGAGGCUU 494
miR_2412 GUGAACGGGCGCCAUCCCGAGGCUUU 495 miR_2413
GUUGGUGGAGCGAUUUGUCUG 497 miR_2414 GUUGGUGGAGCGAUUUGUCUGG 498
miR_2415 GUUGGUGGAGCGAUUUGUCUGGU 499 miR_2416 GUUGGUGGAGUGAUUUGUCU
501 miR_2417 GUUGGUGGAGUGAUUUGUCUG 502 miR_2418 GAACGCGGUCUGAGUGGU
504 miR_2419 UACCGGGUGCUGUAGGCUUU 507 miR_2420 UCCUGUACUGAGCUGCCCCG
509 miR_1120 UCGACGAGCUCACAGUCUAGU 511 miR_2422
UCGGACACCGGCGCGUCUCU 513 miR_2423 UCUGCCCAGUGCUCUGAAU 515 miR_2424
UCUGCAAGUGUCAGAGGCGAG 517 miR_2425 UCUGCAAGUGUCAGAGGCGAGG 518
miR_2426 UCUUCUCUGUUUUGGCCAUGUG 520 miR_2427 UGAUAUGUUUGAUAUUGGGUUG
522 miR_2428 UGAUUGUCCAAACGCAAUUCUCG 524 miR_2429
UGCUGGAUCAGUGGUUCGAGUC 526 miR_2430 UUAGGGCCCUGGCUCCAUCU 528
miR_2431 UUCUCUGUUUUGGCCAUGUGUG 530 miR_2432 UUCUGCCCAGUGCUCUG 532
miR_2433 UUCUGCCCAGUGCUCUGAAU 534 miR_2435 UUGGCCAUGGGGCUGCGCGGG
536 miR_2436 UUGGCCAUGGGGCUGCGCGGGG 538 miR_2437
UUGGGGAAACGGCCGCUGAG 540 miR_2438 UUGGGGAAACGGCCGCUGAGU 541
miR_2439 UUGGUGGAGCGAUUUGUCU 543 miR_2440 UUGGUGGAGCGAUUUGUCUG 544
miR_2441 UUUCCGGCUCGCGUGGGUGU 546 miR_2442 AAAGCUGGGUGAGAGGG 548
miR_2443 AAAGCUGGGUUGAGAGG 549 miR_2444 AAAGCUGGGUUGAGAGGG 550
miR_2445 AAAGCUGGGUUGAGAGGGC 555 miR_2446 AACCUUGGAGAGCUGAGC 557
miR_2447 AAGCUGGGUUGAGAGGGC
Example 5
Expression Data
Array Design
[0617] The array contains LNA spiked capture probes for detection
of the microRNAs listed herein
Sample Collection
[0618] Samples were collected and frozen to -80.degree. C., before
treatment with RNAlater ICE (Cat#7030, Ambion) according to
manual.
RNA Extraction
[0619] Samples from RNAlater.RTM. ICE (Ambion) were disrupted and
homogenized using the TissueRuptor (Qiagen) in Trizol reagent
(Invitrogen). Total RNA was extracted from the samples using the
protocol provided with the Trizol reagent.
[0620] Quality Control (QC) of the total RNA samples were performed
with the Agilent 2100 BioAnalyzer using the Agilent RNA6000 Nano
kit. RNA concentrations were measured in a NanoDrop ND-1000
spectrophotometer.
RNA Labelling and Hybridization
[0621] Essentially, the instructions detailed in the "miRCURY.TM.
LNA microRNA, Hy3.TM./Hy5.TM. Power labeling kit" were
followed.
[0622] All kit reagents were thawed on ice for 15 min, vortexed and
spun down for 10 min.
[0623] For hybridization, the 12-chamber TECAN HS4800Pro
hybridization station was used. 25 .mu.L 2.times. hybridization
buffer was added to each sample, vortexed and spun.
[0624] Samples were incubated at 95.degree. C. for 3 min before
injection into the hybridization chamber. The hybridization
chambers were primed with 100 ul of 1.times. Hybridization buffer.
50 .mu.l of the target preparation was injected into the
Hybridization station and incubated at 56.degree. C. for 16 hours
(overnight).
[0625] The slides were washed at 60.degree. C. for 1 min with
Buffer A twice, at 23.degree. C. for 1 min with Buffer B twice, at
23.degree. C. for 1 min with Buffer C twice, at 23.degree. C. for
30 sec with Buffer C once. The slides were dried for 5 min.
[0626] Slides were scanned using the DNA Microarray Scanner
(Agilent) at 100% PMT setting.
Image Analysis and Data Processing
[0627] Image analysis and spot identification was done using
Imagene 7.0.0 software (Biodiscovery). Tumor and normal adjacent
tissue were paired and normalized with Lowess smooth fitting using
Genesight 4.1.6 Lite edition software (Biodiscovery).
[0628] Ratios of tumor/normal were calculated and log 2 transformed
from the Lowess normalized data sets. Raw values (Lowess normalized
values) were further treated and median scaled for comparison
between samples. To illustrate the tissue specificity of the
individually microRNAs, an index was calculated as a ratio between
the median scaled signal and an average of signal across tissues.
This ratio was log 2 transformed and stated as the tissue
specificity index.
Results
[0629] Differential expression of microRNAs between tumor and
normal adjacent tissue can be identified using the log 2 ratios in
Table 6 (ratios.sub.--454miRs) and in Table 7
(ratios.sub.--454premiRs). E.g. seq ID 113 are downregulated in
tumor samples compared to normal adjacent tissue by 2
(-1.61).apprxeq.3 fold. This could indicate a function of this
microRNA in the regulation of normal tissue, which is impaired in
the tumor cells. This observation could lead to the development of
a cancer biomarker useful for identification of special phenotypes
important for treatment of for instance adrenal cancers.
[0630] Table 6 is provided in Parent U.S. application Ser. No.
11/975,644, which is hereby incorporated by reference. table
ratios.sub.--454miRs: Contains ratio data generated on tumor versus
normal adjacent tissues from a variety of tissues.
[0631] Table 7 is provided in Parent U.S. application Ser. No.
11/975,644, which is hereby incorporated by reference.
ratios.sub.--454premiRs: Contains ratio data generated on tumor
versus normal adjacent tissues from a variety of tissues.
[0632] Tissue specific expression of a microRNA can be identified
with Table 8 (tissuespecificity_index.sub.--454miRs) and Table 9
(tissuespecificity_index.sub.--454premiRs). A positive tissue
specificity index value indicates a higher prevalence of this
microRNA in a sample compared to the average, whereas a negative
value indicates lower levels of the specific microRNA. E.g. Seq ID
113 gives negative values for e.g. uterus, stomach, prostate and
colon indicating lower or no expression of this microRNA in these
tissues. This indicates that the expression of this microRNA is
limited to a number of tissues.
[0633] Table 8 is provided in Parent U.S. application Ser. No.
11/975,644, which is hereby incorporated by reference. Tissue
Specificity Index--454miRs: Contains a tissue specificity index
calculated as the log 2 ratio between the lowess normalized
expression value for a sample and the average
[0634] Table 9 is provided in Parent U.S. application Ser. No.
11/975,644, which is hereby incorporated by reference. Tissue
Specificity Index--454premiRs of all samples for the corresponding
microRNA: Contains a tissue specificity index calculated as the log
2 ratio between the lowess normalized expression value for a sample
and the average of all samples for the corresponding
pre-microRNA.
[0635] TABLE 10 LISTS THE DIFFERENT TISSUE SAMPLES USED FOR THE
MICRORNA PROFILING.
TABLE-US-00006 Tissue Sample names Diagnosis Stage Adrenal gland
T/N-30 Phaeochromocytom T/N-30 Technical Phaeochromocytom replicate
Bladder T/N-34 urothel cell carcinoma T/N-46 carcinoma T/N-74
urothel papilloma grade 2 T/N-ambion transitional cel T3aN1MX, G2
carcinoma Colon T/N-47 ND T/N-47 Technical ND replicate Cervix
T/N-ambion Sqaumous cell carcinoma Duodenum T/N-79 Technical
adenocarcinoma replicate T/N-79 Technical adenocarcinoma replicate
Esophagus T/N-ambion infiltrating moderately T2bNXMX, G2
differentiated adenocarcinoma Kidney T/N-48 renal cell carcinoma
T/N-55 renal cell carcinoma T/N-55 Technical renal cell carcinoma
replicate T/N-57 renal cell carcinoma T/N-57 Technical renal cell
carcinoma replicate T/N-60 renal cell carcinoma T/N-61 renal cell
carcinoma T/N-66 oncocytom T/N-73 ND T/N-76 clear cell carcinoma
T/N-81 urothel papilloma grade 2 Liver T/N-ambion hepatoblastoma
Lung T/N-ambion Squamous cell T1N1M0, stage2 carcinoma poorly
differentiated Mammary T/N-32 lobular carcinoma T/N-41 ND T/N-43 ND
T/N-44 ND T/N-45 ND T/N-52 Ductal carcinoma T/N-53 Ductal carcinoma
T/N-ambion invasive ductal T3N1aMX carcinoma Ovary T/N-ambion
papillary cystadenocarcinoma Prostate T/N-ambion Adenocarcinoma
T2bN0M0, gleason score 6 (3 + 3) Rectal T/N-31 ND T/N-50
adenocarcinoma T/N-62 adenocarcinoma T/N-68 adenocarcinoma T/N-69
ND T/N-71 adenocarcinoma T/N-85 ND Stomach T/N-ambion
Adenocarcinoma Uterus T/N-38 endometrial adenocarcinoma T/N-ambion
endometrial T1bNXMXG2 adenocarcinoma
TABLE-US-00007 TABLE 11 SEQ ID NO 538 mRNA comprising miR2437
target region C17orf76 APC2 CUL3 LOC648570 SCRT2 ZFP36L1 HMGB1
SPRY4 C6orf190 ARHGEF12 DLGAP2 LPPR4 SEMA4G ZNF644 HNRPD SRGAP2
CNIH3 AZIN1 DNAJB5 MAPK6 SETD7 ADCY6 HOXA6 TPSD1 CROP BAZ2A DST
MAT2A SFRS1 AP4E1 IGFBP5 TSC22D3 DNAJB12 BNC2 FLJ10154 MBD6 SMARCD1
APH1A KIAA1305 UBTF ELK1 C18orf34 FUBP1 MKLN1 SNURF BRSK1 LHX6 WNK3
JOSD1 C5orf16 G3BP1 MMP2 SRCAP BTRC LOXL3 XKRX MBD2 CALN1 G3BP2
MSI1 STRN4 C12orf48 MBNL3 ZNF207 MYST3 CAMTA1 GABRE NFIB TCF7 C1QL3
NKX2-2 ZNF219 ROBO2 CASKIN1 GDI1 PCDH17 TGFBR1 C1orf108 PLCH1
ZNF618 SLC4A10 CD24 GJA5 PRDM12 THRA CYFIP2 PSD3 USP12 CENTD2 HCCS
PUM1 TSGA14 DBN1 PSMD2 YES1 CHD3 IGF2BP1 RBM14 UBE2R2 DCHS1 PVRL2
ZFHX4 CPEB4 IGSF21 RHOC VTI1A ELAVL3 RAB6B PHC2 CRSP2 IL1RAPL1
RSBN1 WDR68 FBS1 RBM25 AKAP11 CTF8 KIAA0174 SCRT1 XPO7 FHL5 SLC35A2
SEQ ID NO 540 mRNA comprising miR2438 target region C17orf76 APC2
CUL3 LOC648570 SCRT2 ZFP36L1 HMGB1 SPRY4 C6orf190 ARHGEF12 DLGAP2
LPPR4 SEMA4G ZNF644 HNRPD SRGAP2 CNIH3 AZIN1 DNAJB5 MAPK6 SETD7
ADCY6 HOXA6 TPSD1 CROP BAZ2A DST MAT2A SFRS1 AP4E1 IGFBP5 TSC22D3
DNAJB12 BNC2 FLJ10154 MBD6 SMARCD1 APH1A KIAA1305 UBTF ELK1
C18orf34 FUBP1 MKLN1 SNURF BRSK1 LHX6 WNK3 JOSD1 C5orf16 G3BP1 MMP2
SRCAP BTRC LOXL3 XKRX MBD2 CALN1 G3BP2 MSI1 STRN4 C12orf48 MBNL3
ZNF207 MYST3 CAMTA1 GABRE NFIB TCF7 C1QL3 NKX2-2 ZNF219 ROBO2
CASKIN1 GDI1 PCDH17 TGFBR1 C1orf108 PLCH1 ZNF618 SLC4A10 CD24 GJA5
PRDM12 THRA CYFIP2 PSD3 USP12 CENTD2 HCCS PUM1 TSGA14 DBN1 PSMD2
YES1 CHD3 IGF2BP1 RBM14 UBE2R2 DCHS1 PVRL2 ZFHX4 CPEB4 IGSF21 RHOC
VTI1A ELAVL3 RAB6B PHC2 CRSP2 IL1RAPL1 RSBN1 WDR68 FBS1 RBM25
AKAP11 CTF8 KIAA0174 SCRT1 XPO7 FHL5 SLC35A2
[0636] The remainder of Table 11 is provided is provided in Parent
U.S. application Ser. No. 11/975,644, which is hereby incorporated
by reference
[0637] TABLE 12--A SUMMARY OF EXPRESSION ANALYSIS IS PROVIDED IN
PARENT U.S. application Ser. No. 11/975,644, WHICH IS HEREBY
INCORPORATED BY REFERENCE (T=Tumor, N=Normal, ratio T/N of values
for each tissue type is provided.).
TABLE-US-00008 TABLE 13 Seq ID MiR name BC NAT T7826 T7732 538
miR_2437 0 2 0 0 540 miR_2438 1 1 0 1 The table represents the
microRNA cloning frequency from four separate samples - methodology
as described above. BC = Breast cancer tissue from 5 BC patients
(described under "Example 1) E: NAT, normal adjacent breast tissue
from the same patients F: T7826, an ovarian teratoma cell line, Hs
38.T (ATCC No. CRL-7826) G: T7732, a sacrococcygeal teratoma cell
line, TE76.T (ATCC No. CRL-7732) The cloning frequencies were not
normalized The total counts from each of the four samples were: BC:
93.078 NAT: 119.148 T7826: 66.970 T7732: 66.585
[0638] The remainder of Table 13 is provided in Parent U.S.
application Ser. No. 11/975,644, which is hereby incorporated by
reference
Sequence CWU 1
1
558128RNAHomo sapiens 1cgagccuggu gauagcuggu uguccaag 282165RNAHomo
sapiens 2aagauucaug aguagcagug acaaaccuac cgagccuggu gauagcuggu
uguccaagau 60agaaucuuag acaacucccu auaccagauc cucuaauuaa uuuuaauaaa
ggccuucuau 120uuauacuagc cacaucaagc cuagccgucu acguacccac agaau
165318RNAHomo sapiens 3gaggcugagg cgagaggu 184158RNAHomo sapiens
4cgguguuucc ggccgccguc gcuguccagg gaggcugagg cgagagguag cuguccgggu
60ggggagcccg cacuaccuuc uuccucuucc uccuccuccu ccgggugagg ggagcgaagg
120uugggggucc ccgagcccau ggaccaggag gaggcgga 158523RNAHomo sapiens
5auuuguggcc gaguguaaca acc 236163RNAHomo sapiens 6aucacccucu
gaucgccgau caccucugag acccaacuug cucauaaaca aaacugccca 60ugucgguccu
cugcccugga ccugugacac ucuggacuau uucuguguuu auuuguggcc
120gaguguaaca accauauaau aaaucaccuc uuccgcuguu uua 163716RNAHomo
sapiens 7ucccuucgug gucgcc 168156RNAHomo sapiens 8auccugccuu
ggcuggaaaa gccagccuuc cacccagcgc cccuaaaaug aucggguuga 60cuccaguuuu
guuacgaaag gaggccgggc ugcugagagg cucccugagu ucccuucgug
120gucgcccguc acauugcccu gcuguauacu uaauaa 156921RNAHomo sapiens
9gcaggggaag aagccuuccg u 2110161RNAHomo sapiens 10cagacaaaga
ggggcgugag ggagcugccu gcaggggaag aagccuuccg uaucgagcug 60ggcggucauu
acacaggugc gcacagauau ggaugaggag gggcgugagg gagccacgug
120cagggcagca aguccucuau aucuugagcu ggguggucau u 1611125RNAHomo
sapiens 11acuuugagag uuagaaaugg uuacu 2512165RNAHomo sapiens
12ugaaacuaac uagauuuauu ggauaucagu acuuugagag uuagaaaugg uuacugauug
60ucaucuuuuc agugaagggu ucuauaguug aguaaaaaau uugucuaacu uuguaaguau
120aguuuauauu guagaaauug cuuccaauuu uguugguaac uucau 1651324RNAHomo
sapiens 13aaccaaugau guaaugauuc ugcc 2414164RNAHomo sapiens
14auugaguagg gcaaauuuua aauggguauu auuuuucauc uucaaacagg cagaccuguu
60auccuaaacu aggugaguca gcuuuuggua caugugauga uuuucagugu aaccaaugau
120guaaugauuc ugccaaauga aauauaauga uaucacugua aaac 1641520RNAHomo
sapiens 15augauucugu gacgccagcu 2016160RNAHomo sapiens 16uuggcaggaa
guuuccaugg ccaugcagcc gcagggucac ccugagugcu uuucagggug 60gcagggccuu
gccucagaug gccacaaggg caccucuccu uggauacuuu augauucugu
120gacgccagcu acuugguuug cuuuuuguau uuuuaugcau 1601722RNAHomo
sapiens 17gaaagcugag cgugaacgug gu 2218162RNAHomo sapiens
18cuucaaguau gccugggucu ugcauaaacu gaaagcugag cgugaacgug guaucaccau
60ugauaucucc uuguggaaau uugagaccag caaguacuau gugacuauca uugaugcccc
120aggacucaga gacuucauca aaaacaugau uacagggaca uc 1621923RNAHomo
sapiens 19uagcagcaca uaaugguuug aau 2320163RNAHomo sapiens
20auagaaucau cuaaguauuc agacacugcu uuccuaggaa auguuaaacu ccuugaggca
60ggcuggcuuc cucaccaccu ugugcacugc ugcucccaca ccacagugac uagcagcaca
120uaaugguuug aauuaaagcu gaaguaaaaa auauccaggu cca 1632119RNAHomo
sapiens 21uguuuagacg ggcucacau 1922158RNAHomo sapiens 22cuuaccuccu
caaagcaaua cacugaaaau guuuagacgg gcucacauca ccccauaaac 60aaauagguuu
gguccuagcc uuucuauuag cucuuaguaa gauuacacau gcaagcaucc
120ccguuccagu gaguucaccc ucuaaaucac cacgauca 1582324RNAHomo sapiens
23ggaaauuuga gaccagcaag uacu 2424164RNAHomo sapiens 24auuugagaag
gaggcugcug agaugggaaa gugcuccuuc aaguaugccu gggucuugga 60uaaacugaaa
gcugagcgug aacaugguau caccauugau aucucuuugu ggaaauuuga
120gaccagcaag uacuauguga cuaucauuga ugccccagga caca 1642525RNAHomo
sapiens 25auuguauauc agcaugggga uuauu 2526165RNAHomo sapiens
26ugccuaauac ugacucagau gcacaaucca guuaacccag augugugaga ucuuccgguu
60ugaaagaacu guauuggcaa ggcaaaauca accuauugua gaauauauuu auuguauauc
120agcaugggga uuauuaauau ugcuaauaaa accauuauuu guaaa 1652715RNAHomo
sapiens 27uuggaggcgu ggguu 1528155RNAHomo sapiens 28gggaaggaua
aagggauggc augguggggg uuggaggcgu ggguuuuaga accuaucccu 60uucuagcccu
gagcaaugcu ugccccagaa ggaguugggg cuaggcccau uccaauccuu
120ccagccuaag auccagacuc caaggcaugc cccag 1552922RNAHomo sapiens
29aacgugcagc ggcugaagga gu 2230162RNAHomo sapiens 30ucaaucugga
ggagcucagg guggccggca uucacaagaa gguggcccgg accaucggca 60uuucugugga
uccgaggagg cggaacaagu ccacggaguc ccugcaggcg aacgugcagc
120ggcugaagga guaccgcucc aaacucaucc ucuuccccag ga 1623119RNAHomo
sapiens 31aauugugagg cuugagugu 1932159RNAHomo sapiens 32accccaucca
auuuaaucgg guguuauuua auuauacuac uauaauuguu guauuugcag 60guuugacugu
ucucagggaa cgcugaaggu ucauaacagu agugauuugu aauugugagg
120cuugagugug gaauugaauu acuucauuag agaguaacc 1593322RNAHomo
sapiens 33ugauaggauu gacauggagc ac 2234162RNAHomo sapiens
34cuuauuuacg uuacuggcug aaagccuguc ugauaggauu gacauggagc acuaauuaau
60caccuagggu cuccucauuu acuaaucaua uugcacaaaa cuucccugcu gacugaggcu
120caagggcaaa cugugggcuu ccagccagcu uauuuuucau aa 1623520RNAHomo
sapiens 35aacggccgcg guacccuaac 2036160RNAHomo sapiens 36guuaaaaaaa
guaaaaggaa cucggcaaau cuuaccccgc cuguuuacca aaaacaucac 60cucuagcauu
cucaguauua gaggcaccgc cugcccagug acaugcguuu aacggccgcg
120guacccuaac ugugcaaagg uagcauaauc acuuguuccu 1603725RNAHomo
sapiens 37ucuugccaag agaauuaaug ugcgu 2538165RNAHomo sapiens
38uguuguaaac aaacaaguua agagcaagau ucuugccaag agaauuaaug ugcguauuga
60gcacauuaag cacucugaga gcugggauag cuuccugaaa uacaugaagg aaaaugauca
120gaaaaagaaa gaagccaaag agaaagguac cuggguucaa cugaa 1653924RNAHomo
sapiens 39aaucugagug agagguuagu ugcu 2440164RNAHomo sapiens
40uucugaaggg uauauugcau guucauuauu aaucugagug agagguuagu ugcuauaaag
60uuacaagauu uuccugauca cuuaaaauuu ucuuguaguc agagauuuga cucugaaucg
120ucacuucaaa aauuugucuu uucagguacu auguaaagag aacu 1644119RNAHomo
sapiens 41auuaaaaauu ucgguuggg 1942159RNAHomo sapiens 42ccgugaagag
gcgggcauga cacagcaaga cgagaagacc cuauggagcu uuaauuuauu 60aaugcaaaca
guaccuaaca aacccacagg uccuaaacua ccaaaccugc auuaaaaauu
120ucgguugggg cgaccucgga gcagaaccca accuccgag 1594325RNAHomo
sapiens 43gauagcugcc agugacagga guagu 2544165RNAHomo sapiens
44auucgugcug aaaaucucag acucauugau gauagcugcc agugacagga guaguguugc
60cacuguaaga uacgccaucu uuguuaguua cucucaucua cucguuucuu guauucugcc
120ucuuggucau cuuugauucu cauuuaucug caaauuuucu uggua 1654518RNAHomo
sapiens 45gagccuggug auagcugg 1846158RNAHomo sapiens 46agauucauga
guagcaguga caaaccuacc gagccuggug auagcugguu guccaagaua 60gaaucuuaga
caacucccua uaccagaucc ucuaauuaau uuuaauaaag gccuucuauu
120uauacuagcc acaucaagcc uagccgucua cguaccca 1584718RNAHomo sapiens
47uggcgcggag cggagcgg 1848158RNAHomo sapiens 48aggcagcucg
gcgggcggcg ggcggcauuc uggcgcggag cggagcggcg gcgggcgcag 60cuagcggguc
ggccgcggag cggaggugca gcucggcuuc ccccggcacc ccucccccuc
120gggcgccagc cccaccccuc cgccggccgg gccgaccc 1584916RNAHomo sapiens
49aggcggcgga ggggcg 1650156RNAHomo sapiens 50cggcccuguc cugcgggggu
ccggucgcgg aggcggcgga ggggcgcggg gacacucccc 60accuccacug uccgcccguc
ggccccggug gccuuuucuc gccucgcgca cagcuccccc 120gccgcagggc
ugagagagag aguggccguc uggugc 1565121RNAHomo sapiens 51augcgugcga
gaagucagug g 2152159RNAHomo sapiens 52caggccuuuc ugaaggaguu
auucugcuaa aaauggucuu aguugucuga aaagccagcu 60cuugaaccuc uucacaacag
uaucaacacu ggcuucuccc gguucauuuu augcgugcga 120gaagucagug
guaacugcug cagggcuuaa uacauuagu 1595326RNAHomo sapiens 53uguguuugag
agcaacgcca uugccu 2654163RNAHomo sapiens 54agcauuugag ggugaugaug
gauucugugu guuugagagc aacgccauug ccuacuaugu 60gagcaaugag gagcugcggg
gaaguacucc agaggcagca gcccaggugg ugcagugggu 120gagcuuugcu
gauuccgaua uagugccccc agccaguacc ugg 1635519RNAHomo sapiens
55gcaugagugg uucaguggu 1956159RNAHomo sapiens 56auuuaaaacu
gugucuuucu gugucccuga aauucucaca caugguacgu uuucaaugag 60cugauuuugu
uucuccacuc aaugcaguaa uugagcuucu uugguucagu gcaugagugg
120uucagugguu cauugggcau ccugguugag ggaggggcu 1595719RNAHomo
sapiens 57aaggaaugag uuagcuuug 1958159RNAHomo sapiens 58uuuaugguug
cuacuguagg uuuauaauuu guuuauaauu uggccuaauu uccaucagcc 60auacuaauau
uggauuuuaa aaggaggcaa cuuuuuuucu uuuugaacca aaggaaugag
120uuagcuuuga aaacauaauu ugggauauua uaguaugga 1595920RNAHomo
sapiens 59acaagugaau acacugaggc 2060160RNAHomo sapiens 60ccugcugggg
uuggaguucu uaaugaacau acaagugaau acacugaggc aaaaaaauua 60aagcucucca
acuguggggu auucauucug uucacugugg ccaguguggu gaucaguacu
120ggccacacca guggccaaag agaacugcau ucaucaugug 1606122RNAHomo
sapiens 61cucgcgcccg cgucgcggca gc 2262162RNAHomo sapiens
62ccucagcccc uucagagagc gacuuucaaa cucgcgcccg cgucgcggca gcaccugggc
60agccccgcac gccgugcgcg ucccgagccc gcggggcagc uaccgcucgg ugaguguccc
120cugauucucc ucucuccccu cuuaucuccc ugcauuaggc ug 1626321RNAHomo
sapiens 63gugguguuga ggaaagcaga c 2164161RNAHomo sapiens
64cugaaaaguu ccagcauauu uugcgaguac ucaacaccaa caucgauggg cagcggaaaa
60uagccuuugc caucacugcc auuaagggug ugggccgaag auaugcucau gugguguuga
120ggaaagcaga cacugaccuc accaagaggg cgggagaacu c 1616521RNAHomo
sapiens 65guuuuuguga cugccuugag u 2166161RNAHomo sapiens
66caugcuuuug guuuguuacc aaaauauaca guguggugaa gguugacuga agaaguccag
60uguguccagu uaaaacagaa auaaauuaaa cucuucauca acaaagaccu guuuuuguga
120cugccuugag uuuuaucaga auuauuggcc uaguaauccu u 1616724RNAHomo
sapiens 67aaggcacagc uggaaaugau ggug 2468164RNAHomo sapiens
68cccagggugg cauaggagaa aaaggugcug aaggcacagc uggaaaugau ggugcaagag
60uaagugaaag uauuccuuuu cuagcugggc uaggaaccaa cauuacagua ucauaugagu
120uucccugaac cuuccaaaau aaagucaguc auguuuccuu ggua 1646920RNAHomo
sapiens 69aaacuaggcg gcuaugguau 2070160RNAHomo sapiens 70gggucaauag
uacuugccgc aguacucuua aaacuaggcg gcuaugguau aauacgccuc 60acacucauuc
ucaacccccu gacaaaacac auagccuacc ccuuccuugu acuaucccua
120ugaggcauaa uuauaacaag cuccaucugc cuacgacaaa 1607119RNAHomo
sapiens 71gugguggucg uacgcugug 1972159RNAHomo sapiens 72ggcggcugcc
gaagauggcg gaggugcagg ucccaguccu ccauggucga ggccaucucc 60ugggccgccu
ggcggccauc guggcuaagc agguaauguu gggcuggaag gugguggucg
120uacgcuguga gggcaucaac auuucuggca auuucuaca 1597322RNAHomo
sapiens 73gcugagugaa gcauuggacu gu 2274162RNAHomo sapiens
74gcccaggacu ccagcucaug cgccgaauag uagguacagu guuccaaugu cuuugugguu
60uguagagaac aaucaacggu cggcgaacau cagugggaua agguaaaaug gcugagugaa
120gcauuggacu guaaaucuaa agacaggggc uaagccucuu uu 1627520RNAHomo
sapiens 75agguucacau ggaaaagguu 2076160RNAHomo sapiens 76ugguguagag
gcggagagga gccaagaaac uaaaggugaa aaauacacug gaacucuggg 60gcaagacaug
ucuaugguag cugagccaaa cacguaggau uuccguuuua agguucacau
120ggaaaagguu auagcuuugc cuugagauug acucauuaaa 1607722RNAHomo
sapiens 77ccauugugua gcaagauguc au 2278162RNAHomo sapiens
78gacuaaaacu auuugaucuu uuaauauuua auuaaugguu ccccguggcg uuuuuauagu
60cuguguucua uugugagcaa cgagauuuua auaagcaggu ucaggacuuu ccauugugua
120gcaagauguc auugcuucca ugacacuaau uuggcuuuca ua 1627924RNAHomo
sapiens 79ggaaauaguu ugugacaaac uggg 2480164RNAHomo sapiens
80cacauacuca aggagcccug uuuuacaggg cacuggagaa cuaauuaauu ugcaaugcag
60aaagaaugca gugacaucug aaauauuggc cucggguauc acaggucauu ggaaauaguu
120ugugacaaac ugggguggag ggugggggug gggaaggcaa cucu 1648121RNAHomo
sapiens 81cuccucuucg ucuccucggu c 2182161RNAHomo sapiens
82gggggaggcc ugcgcggagg agcaccgcuu ccucccgccg ggagggggag ucccgggcuc
60cugcguccug ucucccuccc cggccgucug caggagcacg aagggagugc cuccucuucg
120ucuccucggu ccccguaacu ucuccccuca cuuccuccug g 1618321RNAHomo
sapiens 83acuccagccc cacagccuca g 2184161RNAHomo sapiens
84gucaaaaguc cccagagucu ucacacaagc cgugguguau gaagcugcau ccucaggacc
60ugggcuuggg ugguaggagg aauuggugcu ggucuuucau uuuggauuug acuccagccc
120cacagccuca gccaccccag ccaauuguca uaggagcugg a 1618522RNAHomo
sapiens 85gccgcgagug ggagcgggag cg 2286162RNAHomo sapiens
86gcgcggcgga ggggggggug gggccuuggg gccgcgagug ggagcgggag cgguucugcg
60gccuccucgg gcuucuuggc ccugggcgga gugggauugg gugucccggc uguucgcagu
120ggccgcgagu gcggccggac cuggaguagu accugagccg cu 1628725RNAHomo
sapiens 87gcuugacuga guguggcugg acgug 2588165RNAHomo sapiens
88gagaacaugg gucaccagca gggccugaga agagggagaa aauacggaaa ugugggauug
60gggucgcuga gugcaggcau guaaguuaag uguuugggga acagagcagu gcuugacuga
120guguggcugg acgugaguac ugagggggac aaaugagauu gaucc 1658919RNAHomo
sapiens 89agucggugcc ugagguugc 1990159RNAHomo sapiens 90caggacggcc
gccaucuugc gcgcagcugg agucggugcc ugagguugca gccgagagug 60ugcgccagcc
cgcggcccag ccgaagcucu uucccgccgc cucuccgcgc cucgcccagg
120uucagcuccg ccugacccuc cgcuuggcac gguccccug 1599120RNAHomo
sapiens 91agugaugagg augugcugau 2092160RNAHomo sapiens 92uuaggaaguc
ugagaugaua aauauuucaa ggucagugaa gucuaucaau cauucucccc 60uuccucauca
gcaaugguag auagaaaugu ccuaaacuuu ucuaaauccu agugaugagg
120augugcugau auucaacaua guccuuaaag ugaaaacuga 1609320RNAHomo
sapiens 93auugaguggg gcucaggauu 2094160RNAHomo sapiens 94agcuacuucc
uuucuucagc cucuugcuuu cuguucaaau cucagcuuuu aucacauucu 60uuucauggag
agacaucuca augucccuuu ucgccuagga gaagaauguu auugaguggg
120gcucaggauu uaaacccagg cagacuaauu gguaugugag 1609518RNAHomo
sapiens 95gcaugggugg uucagugu 1896158RNAHomo sapiens 96cacauacauu
ggcgaaaaga ccaacaaguc augauuguuc ugaaguuccc uuuaucaugu 60uguccccuaa
ucucuacuac caguaagccu uuguguuauc uuaggaugag gcaugggugg
120uucaguguuu auaauaagac gagucuaaaa uggacaau 1589723RNAHomo sapiens
97aacuguuaua uuaugauugu gac 2398163RNAHomo sapiens 98uguuuaaugc
cugaaaucca agucuuccuc caugggaaaa uacuguuaua ccaaauaauu 60cuagaugagu
aacaaagauc uuuuuaggcc uucauuuuau guuuuuucuu aacuguuaua
120uuaugauugu gacauagauu auacuacuac uaauuuuugg aug 1639924RNAHomo
sapiens 99uugcugugau gacuaucuua ggac 24100164RNAHomo sapiens
100cuuacggaaa aggaacagau uguuccuaaa ccagaagagg agguugccca
gaagaaaaag 60guaaauaagu aguugcucgg uuuuguuugu gauaguagaa agauuugugg
uugcugugau 120gacuaucuua ggacaccuuu ggaauaacua ugaaagaaaa cuau
16410122RNAHomo sapiens 101agaauugagu gaucucaugg au 22102162RNAHomo
sapiens 102cuaggaguga cucaugcaga cucaagcaga accuuggggc ccagggcaga
agugugacuu 60caggucucac ucagcacuca ucagaacacu cacucaguau gcuuguaauu
agaauugagu 120gaucucaugg augaacugua cugggcuaac uugaagagca ca
16210324RNAHomo sapiens 103aaaacgaucu uucagauuua gagu
24104164RNAHomo sapiens 104uaaaagcuga aaaucucagu uuaaaaauca
aaauguuaac acaaagcuaa gauucaucag 60agcccacccu auucuaagga accacaauaa
cuuacucugg ccccaguguu aaaacgaucu 120uucagauuua gagugacuau
guaagaauuu aggauuuccu cuuu 16410517RNAHomo sapiens 105gcggucgggc
ggcggcg 17106157RNAHomo sapiens 106ggccccgcgg ggcccguccg cuccuccagc
cgcugccucc cgggcggcgc ucgccggcgc 60ggcggcaaag acugagacag cuccgcugcc
cgcugaacuc cauccucccg gcggucgggc 120ggcggcggcu gcggucgguc
gcggcagcgg cuccgcu 15710722RNAHomo sapiens 107augagaacuu gagcgacaga
gu 22108162RNAHomo sapiens 108cuccagguca ucaucagugu gguauuaucu
augagaacuu gagcgacaga guauuucuug 60augaauuuau agaucauuug agauguugag
uuacuuuugu uuuguuuuca aauagguaga 120gacuauuaau guaaaaaaac
aagaaaggaa aaugaaaugu gc 16210918RNAHomo sapiens 109auuggucgug
guuguagu 18110158RNAHomo sapiens 110guggggaggu cgaugaauga
gugguuaauu aauuuuauua ggggguuaau uuugcguauu 60ggggucauug guguucuugu
aguugaaaua caacgauggu uuuucauauc auuggucgug 120guuguagucc
gugcgagaau aaugauguau gcuuuguu 15811123RNAHomo sapiens
111gaugagagaa caguggguac uuc 23112163RNAHomo sapiens 112auucucagua
uuggaucugc acauggagug uuuuucucuc uagugguuac agaggaugaa 60ugcauauuga
gauaaagaag ugauuuuggu uccaaaagga uuuuaaggau gaugagagaa
120caguggguac uucauugcca ggucaugucu uugcaagaag aaa 16311323RNAHomo
sapiens 113caagguguag cccaugaggu ggc 23114163RNAHomo sapiens
114agcacaagua cacacauaaa aacauuaggu caagguguag cccaugaggu
ggcacgaaau 60gggcuacauu uucuaugucc agaaaaucuc acaacauccu uuaggaaauc
uaagggcuca 120aggaggauuu agcaguaaac caagagcaga gugcuugguu gaa
16311522RNAHomo sapiens 115agcaccagcc uaggaagagg gu 22116162RNAHomo
sapiens 116cucaccucga uuccucccag gccugggucc agcaccagcc uaggaagagg
gugccccaug 60cugucuagcu cuucuucggg auggggggcu ccagguuccu ugguauuuug
cuuuggccuu 120uggagccuca gucaaaacug aggaaaggug ucauuuucac au
16211723RNAHomo sapiens 117acuuuaacac ugcuguggaa ggc
23118163RNAHomo sapiens 118gaaaauaaag gcaccugaaa agaaacuacu
acuuuaacac ugcuguggaa ggccuuugcu 60uuauaagaaa aauauuauua gcuaugggaa
aguaauguuc uuuauguaaa gacuuaaaaa 120uagacuaaua guuuacagag
uuauuauaua aaauacgaug uga 16311924RNAHomo sapiens 119auuugacaag
aguaugccag gugu 24120164RNAHomo sapiens 120cuaccugaag uuuuaagagu
cuuggaaagu caggagugac uucugcuaaa cacggggcuu 60uccagaguca gagaagcuag
caagccugug guuuggacca gguacuaaau auuugacaag 120aguaugccag
guguaaugag cuacugucua uuccccuuua aagc 16412118RNAHomo sapiens
121cugggagcuu gaaaggag 18122158RNAHomo sapiens 122gcucuuucuc
uucccucucg uuuaguuugc cugggagcuu gaaaggagaa agcacggggu 60cgccccaaac
cccuucugcu ucugcccauc acaagugcca cuaccgccau gggccucacu
120aucuccuccc ucuucucccg acuauuuggc aagaagca 15812323RNAHomo
sapiens 123auuugggcag guugaaagaa uuu 23124163RNAHomo sapiens
124uaguaaaugu aggcaguuuc uuuaggguua aucaucuuuc aaagggccuu
aggaaugucc 60uucaaacaga auauaaaugu caaagagaau aucucuuucu guuugaaauu
auuugggcag 120guugaaagaa uuugauaaag ggaaauucua uauuuaaucu uuc
16312524RNAHomo sapiens 125gugcuggagg ccaggcugag gccc
24126164RNAHomo sapiens 126gugcccggga gguggacugg ggccuggguu
gugcuggagg ccaggcugag gcccugccuu 60gguuugggga ggagaucccu gcacuccgga
acuccucugu ggcccacgga ggaucgcucu 120gaacugccuc agcguggcgg
ccaguggggg uaggggugga gaga 16412718RNAHomo sapiens 127gcuguuggug
gagaaggu 18128158RNAHomo sapiens 128ccugaagagg aagagaugac
uguuggaaag cguuccccuc ccccauacgg cagaacagcu 60gcggcuccca ggggaaagcc
cccgcaggac aguccucgug gggugugacg gcuguuggug 120gagaagguuu
ggcgcccuau uuucuuaucu gccuuucu 15812922RNAHomo sapiens
129auggcaguug gagagaaaga ac 22130162RNAHomo sapiens 130gagaggaaaa
guccagaccu aggacuaguu auggcaguug gagagaaaga acaucgggau 60guuugaaaau
augccauuga cuaucuuaac uacuguaauu uuaucauuuc caacgucauc
120uaacugggga cuagaacaaa cugugaauuc acuuucagca ac 16213125RNAHomo
sapiens 131gcagauaaac ucaugccaga gaacu 25132165RNAHomo sapiens
132ucuuaaaagu uuuauuaaag gggaggggca aauauuggca auuaguuggc
aguggccugu 60uacgguuggg auuggugggg uggguuuagg uaauuguuua guuuaugauu
gcagauaaac 120ucaugccaga gaacuuaaag ucuuagaaug gaaaaaguaa agaaa
16513317RNAHomo sapiens 133agugccaggu ggggagg 17134157RNAHomo
sapiens 134gcccagcacc cucugucucu uuaugcaauc agugccaggu ggggagggau
gcauucuguc 60caaugacaug caggcacuuu agagggcuug cauucauucc caaguccagc
ggcacacuuu 120auacauccuu ggcuggucau ugaggggaac accggag
15713518RNAHomo sapiens 135gacucuuagc gguggauc 18136158RNAHomo
sapiens 136cacgcuugug gugaaaucag gaauuuuuag gacucuuagc gguggaucaa
aaagaaaaaa 60gaaaacagga cagaguaaaa ucuugcucca aagcuugugu ucuggcaaau
accgucugug 120ucucgaaugu gaaguuguuu ccacuccuca gagcccac
15813721RNAHomo sapiens 137cuguugcggg acccggggug u 21138161RNAHomo
sapiens 138uggauuucgc ccccgcuccc ucccggaaac uccuccuggu gccugcgacc
guucucacug 60agcaugugca gacggcggug cgcaugcucu guugcggucc gcuucgguuu
cuguugcggg 120acccggggug ucuccuagcg caaccggaac uagccuucug g
16113921RNAHomo sapiens 139uuaaagcugc cauuuguuac u 21140161RNAHomo
sapiens 140cacuguagca uaagcaaggg cuuaguuccu gaacugaguu acagcuuuau
uuuucuuuug 60auucagcaug uuuuuaauga uccauaaguu aaaagcugcu gguguuuuua
uuaaagcugc 120cauuuguuac uaaccaggcu cugugugacu ccuaagugga a
16114122RNAHomo sapiens 141gagcagaggc gauaguugaa gu 22142162RNAHomo
sapiens 142agccacaccc cagcagugug caagggauca gacacaaggu ugaauccauc
acaaaagcag 60aaucaccaug gcaacugcau ccuuugauuc uugagugugc ccagcaaccu
gagcagaggc 120gauaguugaa gugaaccaag uucuccugag aaauggaggg ga
16214324RNAHomo sapiens 143gaugacaugg gcuuuggucu uuuu
24144164RNAHomo sapiens 144gcaaagaaag aagaaucuga ggagucugau
gaugacaugg gcuuuggucu uuuugacuaa 60accucuuuua uaauguguuc aauaaaaagc
ugaacuuuaa aaaaaagauu gggguuuauc 120auguaauugu uucauuuugu
uguauucugu guuaagauuu cuaa 16414518RNAHomo sapiens 145agaaagggcc
uuguguuu 18146158RNAHomo sapiens 146aggaagggga aacucaauca
gcaggacuuc agaaagggcc uuguguuuau agcuuuguca 60aguaaauuug gacgcagcug
gagcacaggc ccuguuuguu ugcacauaau aaucuuguuu 120aucacuuuaa
aaaauucagu aauaucucag cagucagg 15814726RNAHomo sapiens
147acucggcgug gcgucggucg ugguag 26148164RNAHomo sapiens
148acucccugac agauaucucc cucuuccauu ucaucaagac ccagcugagu
cacugucacu 60gccuaccaau cucgaccgga ccucgaccgg cucgucugug uugccaaucg
acucggcgug 120gcgucggucg ugguagauag gcggucaugc auacgaauuu ucag
16414920RNAHomo sapiens 149guguuuagug aguauuuguu 20150160RNAHomo
sapiens 150agggacaaug ccauauuuau ccuucuagcc cugacaccuc acacaaugca
gagaacggaa 60gggaguucaa uaacugguag caaagugcca acuccuugag aauagggccu
guguuuagug 120aguauuuguu aagagaauga auaaaugaug uacaguugua
16015120RNAHomo sapiens 151acugcugcug cugcuuggcc 20152160RNAHomo
sapiens 152ccccgcucac cuccucuauc cccacagugu acugcugcug cugcuuggcc
aacguuucac 60ugccuggcau cgggggcacc auuccugagu ccaaaccuuu cuucuacgug
aacguggcug 120acaucgagag ccuggaggua gagguguccu auguggccug
16015325RNAHomo sapiens 153ugcagagugg gguuuugcag uccuu
25154165RNAHomo sapiens 154aauuaaagug gcugauuugc guucaguuga
ugcagagugg gguuuugcag uccuuagcug 60uugcagaaau uaaguauugc aacuuacuga
gggcuuugaa ggcucuuggu cuguauuuaa 120ccuaaauuuc uauaagauua
uuaguauaaa aggggagaua gguag 16515521RNAHomo sapiens 155augaccacca
aacccaggag c 21156161RNAHomo sapiens 156uuuuauaacc auagagugga
gacagucagu augaccacca aacccaggag ccauauauua 60aaauacugau aaauuuaacu
auauaaaaaa auuuuugccg ggugcggugg cucacaccug 120ugauucuagc
agaaaaucag aucaggagau cacagaaggu c 16115720RNAHomo sapiens
157cagagucugu agaagaggcg 20158160RNAHomo sapiens 158guccucccac
uggccgcacu cugugcccca uggcccuccu gcgccccgcc cggcguccuc 60ucacggccuc
ugucugugcu gagcuugggu aacucuuguu cuuaccucca cagagucugu
120agaagaggcg acaccagggc uuccaaauga acaaccgaaa 16015919RNAHomo
sapiens 159ccuugacuuc ugccagagu 19160159RNAHomo sapiens
160agcccagaac cccucucacc ccagaaccuu ccuugacuuc ugccagaguu
gagcagccgg 60cccucuggua ggcgcaugug aguggaugug ggcacaugug gcccacugga
ucugguggau 120guggucgcgu cuggcccccu ggaucuggug ggugugggc
15916124RNAHomo sapiens 161aucacugacu gaucaaguag aggu
24162164RNAHomo sapiens 162ucauuucugc cacagucuuu uuuguugaag
caaguuagca agcacuaagc acaucuacaa 60ucaaggagag gggcaggcuu uaccuuuuga
aggaagaagu augaaagugu aucacugacu 120gaucaaguag agguaagcag
uggaggacac ucagaauacc uuuu 16416322RNAHomo sapiens 163aaggauugga
caggguuaga uu 22164162RNAHomo sapiens 164augguuacuu auaugggaag
gguggguaac aaggauugga caggguuaga uuagaccccu 60cugaagguac cuuguuuuau
aguuguaacu uuuuuuuuuu uugagaugga gucuugcucu 120gucacccagg
cuggagugca guggugcgau cucagcucac ug 16216519RNAHomo sapiens
165aggccaaggc ugcgggguu 19166159RNAHomo sapiens 166gggcagccgu
ggggcgugga agccgcgcag aggccaaggc ugcgggguuc uucgucgucu 60acaggcuuuc
gcggcucagu guggaaaacc cgccguuucc ucgcgcccca cguccgaccc
120aggccuccug ggcacccuuc ggggaggccg cgaucucgg 15916721RNAHomo
sapiens 167aacgggaggc gcuagccaug g 21168161RNAHomo sapiens
168accgcaacuc ccuuuccccc acuuucccca aacgggaggc gcuagccaug
gaacauggca 60cauccagggc uaccuccucc caaguuaccc agaggucaug uguacaagca
gcaauucuaa 120caacaguccc ucaggcguga gcggcauuuu acaguuugca a
16116922RNAHomo sapiens 169accuuucagu gcaguuucuu uu 22170162RNAHomo
sapiens 170aaccagaacg ugguuugccu gaggcuguaa cugagagaaa gauucugggg
cuguguuaug 60aaaauauaga cauucucaca uaagcccagu ucaucaccau uuccuccuuu
accuuucagu 120gcaguuucuu uucacauuag gcuguugguu caaacuuuug gg
16217120RNAHomo sapiens 171auuuugggug gaagaggcau 20172160RNAHomo
sapiens 172guguguguau auauguauac auauauguau guguaugugu auauagagag
agagcugaga 60guuauucuau uuauuccuuu ucucuccuaa ucugaaaaug gguguucugu
auuuugggug 120gaagaggcau agaaggggau guguguuguc ucuuaagauu
16017318RNAHomo sapiens 173ggaaugagga gcuuugac 18174158RNAHomo
sapiens 174agacagaaau caggacuaag uccucugcuu caguuucauu guuaacgggc
cuuauucuga 60ucucaccugu cgcguagcuc uaauauucac auaaacugaa auaaagaagu
ggaaugagga 120gcuuugacau ucaaauuaug ugauguaauu uaucuucc
15817520RNAHomo sapiens 175aaaagcuggg uugagaggau 20176160RNAHomo
sapiens 176uauguguaag auagaaugaa uauugagcag gaugcuuuaa aagugaccaa
gcagauuuga 60aaaacauuaa aaauguuggc cuucucgucc caguucuucc caaaguugag
aaaagcuggg 120uugagaggau gaaaagaaaa aaaaagaaaa auuuagugga
16017722RNAHomo sapiens 177aguaagguca gcuaaauaag cu 22178162RNAHomo
sapiens 178cuccgugcua ccuaucacac cacguccuaa aguaagguca gcuaaauaag
cugucaggcc 60cauaccccaa aaauguuggu uacauccuuc cuguacuaau uaaccuauua
gcucagcuua 120ucaucuacuu uacuauuucu acagguaccc uuaucacaau gc
16217917RNAHomo sapiens 179gaauugacgg aagggac 17180157RNAHomo
sapiens 180cuucaggagu uggugguguu gacugggagu gaauugacgg aagggaccau
gggaauuuau 60auaucauuuu gaaacuuaug aaaccuuuug ucaaaguuuc acuuucugac
ucaggcucag 120uccaggacau uguucaauuc cccuggugua ggcauca
15718118RNAHomo sapiens 181uugugucuug ugucuuuu 18182158RNAHomo
sapiens 182ggaaguuugg gauaguaaag uuuguugccu uugugucuug ugucuuuuuu
ccuuuucuuc 60cuuucuuggg ggagauagau agauagacag acagacagac agacagacac
agagagagag 120agagagagag agagacagau aguguucaug gauccugu
15818316RNAHomo sapiens 183ucccuggugg ucuagu 16184156RNAHomo
sapiens 184guaaaaugaa aucacagugg uaugggccuc augggguuau cgaaagaaug
ggcugagguc 60augugggcca ugggcuuggu acagugccug agacauaaug aauacucagu
ucccuggugg 120ucuagugguu agaaaaauaa uaauaauaau aauaau
15618525RNAHomo sapiens 185ugaggcugua acugagagaa agauu
25186165RNAHomo sapiens 186cagaacccac caaccagaac gugguuugcc
ugaggcugua acugagagaa agauucuggg 60gcuguguuau gaaaauauag acauucucac
auaagcccag uucaucacca uuuccuccuu 120uaccuuucag ugcaguuucu
uuucacauua ggcuguuggu ucaaa 16518723RNAHomo sapiens 187uuagggcccu
ggcuccaucu ccu 23188163RNAHomo sapiens 188ugaaaucaac gaaucaccua
ccuaacuggg uuagggcccu ggcuccaucu ccuuuaggaa 60aaccuucugu ggggaguggg
gcuucgaccc uaacccaggu gggcuguaac acugcugugu 120uuucuaaggg
gcagaguuuu cuacuacuuu uccgcuggcc cag 16318924RNAHomo sapiens
189gagugugagu ggacgcguga gugu 24190164RNAHomo sapiens 190guggcgucgc
gugugaggcg cgugcagggu gagugugagu ggacgcguga gugugugagu 60gugcgcgcuu
ggagcguguu aggcgagugc gugcgcccac cccugcgccc cuccucccgc
120uuacacuuug aucuuauuug aucggaucgu gaccccagcc ccgc 16419122RNAHomo
sapiens 191auugagucug gcagucccug uu 22192162RNAHomo sapiens
192gggcuagugu guuuguguuu ccauucuaag auugagucug gcagucccug
uuuuuuugca 60uugggguaac ugcucuuuga uuuuuuuuaa uugcaguauu ugugugauug
caauaauaaa 120guuugguuug guuuuuacag ucaugcgcag ggacgauccu ug
16219321RNAHomo sapiens 193aaaauguuua gacgggcuca c 21194161RNAHomo
sapiens 194uguagcuuac cuccucaaag caauacacug aaaauguuua gacgggcuca
caucacccca 60uaaacaaaua gguuuggucc uagccuuucu auuagcucuu aguaagauua
cacaugcaag 120cauccccguu ccagugaguu cacccucuaa aucaccacga u
16119520RNAHomo sapiens 195gaauguuuau ggcaccugac 20196160RNAHomo
sapiens 196gugguauuug ugauuugguu aaucuguaua aaaauuguaa guagaaaggu
uuauauuuca 60ucuuaauucu uuugauguug uaaacguacu uuuuaaaaga uggauuauuu
gaauguuuau 120ggcaccugac uuguaaaaaa aaaaaacuac aaaaaaaucc
16019721RNAHomo sapiens 197ggaaauuuga gaccagcaag u 21198161RNAHomo
sapiens 198ugauauuacu caccauugau accucuguuu ggaaauuuga gaccagcaag
ugacuaucgc 60uguugccuua ggccacagag acuuuaucaa aaacgugauu acagggacau
aucagguggg 120cuguguuguc cugauuauug cugcuggugu uggcaacuuu g
16119922RNAHomo sapiens 199acaguagggc cuuuggagug au 22200162RNAHomo
sapiens 200gaaggauggu uauucccgcc uggagauccc acaguagggc cuuuggagug
auagacaucc 60ccaucucccu ccacacccug cccuaccgcc ccccaacccc caaagcuuca
acaaaggcuc 120cuuuuuaaag uuuuccggug cccuuugcuc uuuguuugcc uu
16220125RNAHomo sapiens 201cuugaagucu ggugaugcug ccauu
25202165RNAHomo sapiens 202ugguaagaaa cuggaagaug gcccuaaauu
cuugaagucu ggugaugcug ccauuguuga 60uacgguuccu ggcaagcccu uguguguuga
gagcuucuca gacuauccac cugugggucg 120cuuugcuguu caugaucuga
gacagacagu ugucgugggu gucau 16520315RNAHomo sapiens 203agaugagcug
aaggg 15204155RNAHomo sapiens 204ccuaaaauuc ucaauuaggc uauaaaugca
agaugagcug aagggaaaua ggugauuucc 60auucuguagu guguauauau gagguuuuau
ucucaugaca agaaacagac uaugcaaauc 120ucuuuaauuu cuggcauuuc
agcuuucuag aauua 15520518RNAHomo sapiens 205aaaaagggag ccaagaag
18206158RNAHomo sapiens 206ggcaagaaca aguaccuuac gaaagacagc
aaaaagggag ccaagaagug gcugauccau 60uuucuuuuuu ucuuuuuucu uuuuuuugag
acagucuugc ucuauccccc uggcuggaau 120acaauggugu gaucucagcu
cacugcaacc uccgccuc 15820722RNAHomo sapiens 207aguuuugugu
guuggcugcu cc 22208162RNAHomo sapiens 208cacuacggcc ucuggccaug
caccaguugu aguuuugugu guuggcugcu ccacuguugu 60cugccagccc acaggaggga
aagugaggcu ccuggaagga cgcuccuuca gauggaagca 120gcacuggaag
agccccaagu ugaggugcau gggacacaaa cu 16220918RNAHomo sapiens
209gguuuaguga gcagaguu 18210158RNAHomo sapiens 210gaaugaauga
uuuuauagau ugacuauagu gguuuaguga gcagaguuac aauuaugagc 60auuaauuccc
agaccaguuc cccuaacuca ccuugugcuc aaauaugaaa aaggauacca
120cagaaaaugu caguuacucu cagauuaagu aaaaaugg 15821125RNAHomo
sapiens 211aagacgagaa gacccuaugg agcuu 25212165RNAHomo sapiens
212acuuguuccc uaaauaggga cuuguacgaa uugcuacacg aggguucagc
ugucucuuac 60uuuuaaucag ugaaauugac cuaucuguga agagguggau auaaaaaaau
aagacgagaa 120gacccuaugg agcuuuaauu cauuaauaca aauaaaaacu caaac
16521322RNAHomo sapiens 213agcagggugc aggcuuggag uc 22214162RNAHomo
sapiens 214uggacacaga agaaacgagg gagcccgggu cuccuccgag ugugcaacaa
gcuggccugg 60ggccccccga aaggacgcug gagagaagcc caggaucacc cagucuuugc
agcagggugc 120aggcuuggag uccccccaag ggcggcuaga aucaggucca gg
16221520RNAHomo sapiens 215auguuggguu guuacagagu 20216160RNAHomo
sapiens 216ugugucuuuu aaacuggaaa aucuucuagc auguuggguu guuacagagu
auauuuuugu 60cugcagcugu uuguugcccc
auuccuaaga ggaguuuauc cauccugacu uguagcugug 120ugacuucuug
cagugccccc accccauccc cccgggagag 16021720RNAHomo sapiens
217acugaguuga cuguucccuu 20218160RNAHomo sapiens 218cagaagugac
uuuacuuucu caaguuugau acugaguuga cuguucccuu aucccucacc 60cuuccccuuc
ccuuuccuaa ggcaauagug cacaacuuag guuauuuuug cuuccgaauu
120ugaaugaaaa acuuaaugcc auggauuuuu uucuuuugca 16021922RNAHomo
sapiens 219acugggcagu gacaagcacg au 22220162RNAHomo sapiens
220ucacucgcca guaaccuguc ugcaugcaag acugggcagu gacaagcacg
augugcucac 60ugcccaagau uuugcuuuga uuuuguuuua cugcccaaga ucugaacauu
uuuugcaaac 120auagcagcuu cucuaccucu gcugcauuga cauauguuug aa
16222124RNAHomo sapiens 221agaaagcgug aguguccaga gccu
24222164RNAHomo sapiens 222agaagaaaac aaguuaauuu gaagagaguc
agaaagcgug aguguccaga gccuacugag 60cccuggaagu cacggauaaa aacaagaagu
gaagucaaca cucucgguga gaaagggagc 120gguacugaca aacuucuacc
aucccagugu gcccgguugc uccc 16422321RNAHomo sapiens 223gcuugagggc
aguuggugcg g 21224160RNAHomo sapiens 224cccucccggc gcgguugggu
ggcgccucag cgggugggca gcauggggcg gggagggugu 60ccccuccgcg ccguuaaaau
gaaacucuag uggcuggagu ccgggcagag cuugagggca 120guuggugcgg
ucggguuggu ucuuacaccc cggcgggagc 16022525RNAHomo sapiens
225ccgagccugg ugauagcugg uuguc 25226162RNAHomo sapiens
226gauucaugag uagcagugac aaaccuaccg agccugguga uagcugguug
uccaagauag 60aaucuuagac aacucccuau accagauccu cuaauuaauu uuaauaaagg
ccuucuauuu 120auacuagcca caucaagccu agccgucuac guacccacag aa
16222725RNAHomo sapiens 227uggagauggc uggcagaaug guucu
25228165RNAHomo sapiens 228gccuguugag aaagaccucu ggggcccugu
uggagauggc uggcagaaug guucucuuga 60ugagcuucau gauaaagcag acuugccaau
aauaccaaga gagaagacug gcucuacucu 120ccaaaggagu ccagggacag
agagucagac agaugacauc agaag 16522920RNAHomo sapiens 229aaguagaagc
cucagggaag 20230160RNAHomo sapiens 230ccaggagagg aaaaggaaau
gggaccucag aaguagaagc cucagggaag gaguaaagua 60gaaaucagaa gaaaagaagc
uucacuugau aguaauaagg uuuuuaacuu caaguaccuu 120cagaaaaugu
gauuuugaua agaggaaagg gcaaauuuag 16023120RNAHomo sapiens
231augguucugg acaguggauu 20232160RNAHomo sapiens 232agaaauaaua
augcaggguu ucuucaaaau augguucugg acaguggauu auaguuaccu 60ggagagcuug
uguuaaaaua ucugaggaug auuccaagua ccagggcuua uacacaggaa
120uacuugagaa ccacugcacu caagcauuua aaauuuuccu 16023318RNAHomo
sapiens 233gaugagucag gcuaggcu 18234158RNAHomo sapiens
234uccuagggcu uugacaccaa aaucuacuau gaugagucag gcuaggcuau
aaacuugcaa 60ggacuuagag cccagaaagu gacaagccca acuagccugc cucuuggagg
aaaaaagaag 120aauagcucaa aacacuuaag aagguaagga guccaagc
15823522RNAHomo sapiens 235augugagagc agcagaggcg gu 22236162RNAHomo
sapiens 236gggagggugg acaguccuua acugcucugc agguccagga uguuagaaag
gggcagggac 60aacaaauggg ugaccccaac cucaaccugc ugcuucucuc uccagucccc
augugagagc 120agcagaggcg gucuucaaca uccugccagc cccacacagc ua
16223721RNAHomo sapiens 237guuuuaagga cuuaagggua u 21238161RNAHomo
sapiens 238uaggucuguu auuuucacau acacuuggua acucagacug gucugaauau
aaaguagaaa 60uagcuaagaa ccauuuguaa ugaaugcaac ucuuauuugu uuuuaauggu
guuuuaagga 120cuuaagggua uuagaacuga caacaguuua uucaguuaag c
16123920RNAHomo sapiens 239ugauauguuu gauauugggu 20240160RNAHomo
sapiens 240acccccaaag cucuccugcc ugcuucugug ugauauguuu gauauugggu
uguuuaauua 60ggaaccaacu aaaugucaaa cauauucuua cagcagcagg ugauucagca
ccacccucuu 120ucauacuuca aucucugggg cuccugucuc uuuuacugaa
16024121RNAHomo sapiens 241agaaagccag gagcugugau u 21242161RNAHomo
sapiens 242auccguuuug gaaccugcgu cuggggcucc agucgcugcu cuugcuggcg
uccaucgccg 60ccucggacgg ccgugcauuu ucucgucuca cgcaguucga ggaggacccu
agaaagccag 120gagcugugau ugacaguagc uguagguuac cagacggcaa c
16124321RNAHomo sapiens 243auucuaaguc agucagucau c 21244161RNAHomo
sapiens 244uacauuuuag ggugguagag cuacuccuua cuuuaaaugc uaccuacuca
cugugacacu 60guuuaauaaa ugguuauuga cuagagaagu agggaucucu gucaccuagc
auucuaaguc 120agucagucau caguuuuugu agguuaucuc agaagcaaua g
16124523RNAHomo sapiens 245uucucuguuu uggccaugug ugu
23246163RNAHomo sapiens 246uucuucuuca gcaaacauua ggagaguauc
uucucuguuu uggccaugug uguacucaca 60gccccucaca cauggccgaa acagagaagu
uacuuuccua auauuugccu ccuuggagug 120ucucaagucc uggaagcaag
agauaauaag caauuaauau aca 16324723RNAHomo sapiens 247uucuaagcca
guuucugucu gau 23248163RNAHomo sapiens 248cccgacagau cgacuauguu
gaucuaacuu uucuaagcca guuucugucu gauaugccag 60cuugagcagc uccuuugucc
cagcuccccu gggcaucuag cugaugggag cucauuuuuc 120uguuuuuuca
uuucagguuu auuguuggcc aaaaccaggc uuu 16324920RNAHomo sapiens
249gguuuugaca ugucacuguu 20250160RNAHomo sapiens 250gggucaugga
ccagcgccuc agugcauuag ucauucgcuu uuccuuacag acaaaucaga 60uaacucuucc
ccagugauug ucaaauguau gaauguaucu cuguaaaugu gguuuugaca
120ugucacuguu acugaaggag aguauggaau ccccacagga 16025122RNAHomo
sapiens 251cucgcccgug gucucucguc uu 22252162RNAHomo sapiens
252ggugaggccc cgcgcgugug ucccggcugc ggucggccgc gcucgagggg
uccccguggc 60guccccuucc ccgccggccg ccuuucucgc gccuuccccg ucgccccggc
cucgcccgug 120gucucucguc uucucccggc ccgcucuucc gaaccggguc gg
16225325RNAHomo sapiens 253aacacgcaua cgguuaaggc auugc
25254165RNAHomo sapiens 254acaaggaugg aagaggcccu cgggccugac
aacacgcaua cgguuaaggc auugccaccu 60acuucguggc aucuaaccau cguuuuuuuu
uuuuuggugu uuuguuuuuu auuuuucuuc 120agacggaguc uuauucuguc
gcccagacug gagugcaaug gcgcg 16525525RNAHomo sapiens 255ugaaacagca
ucugaucuug aacuu 25256165RNAHomo sapiens 256acaugcaggg ugaaauccuc
acuauuuuua ugaaacagca ucugaucuug aacuuuuaug 60acucaccuca gucacuucac
cuguauuuug gcccugucag aucaugucuu uauuuaaacu 120uuugauauuu
uauucuuuau auguuuuugc auuaguuuuu auuuu 16525725RNAHomo sapiens
257gcguaaagag uguuuuagau caccc 25258165RNAHomo sapiens
258cacacgauua acccaaguca auagaagccg gcguaaagag uguuuuagau
cacccccucc 60ccaauaaagc uaaaacucac cugaguugua aaaaacucca guugacacaa
aauagacuac 120gaaaguggcu uuaacauauc ugaacacaca auagcuaaga cccaa
16525924RNAHomo sapiens 259guccguuucc ugucagagug aucc
24260164RNAHomo sapiens 260cagagcaacu ggcuccuggc agcugugcuu
guccguuucc ugucagagug aucccagguu 60uccuccuggc ccgucccaug gucccuccac
aggaguguga gaggaugggg gaagcacugu 120gggaagacca ccaaagaugg
cuggacagug ggagagagca cguu 16426124RNAHomo sapiens 261ccaucgguga
ucccagugac aagu 24262164RNAHomo sapiens 262aucaaacacu uauccuauua
aacacagcau ccaucgguga ucccagugac aaguaauuga 60auguuaguuc uggagucuuu
ccugggguga uggccuggag aagccucucu uuuaaggauu 120agauucagag
guagagguaa augaguguug agcaccagga agag 16426323RNAHomo sapiens
263gaaauguuga guguuuaccc ugu 23264163RNAHomo sapiens 264guuggcuuuu
ccagggccag cgugaguggu gaggccagcu cucucaguga ccaucagaga 60caaggccuug
gccaguccag gggucuuggg gcuccacuuu ucugaauuau gaaauguuga
120guguuuaccc ugucaauaua uauaucauuu auauauuuuu ugu 16326521RNAHomo
sapiens 265gaagaaacag cucaugaggc u 21266161RNAHomo sapiens
266gaaagagaaa gccaagaucc acuaccggaa gaagaaacag cucaugaggc
uacggaaaca 60ggccgagaag aacguggaga agaaaauuga caaauacaca gagguccuca
agacccacgg 120acuccugguc ugagcccaau aaagacuguu aauuccucaa a
16126723RNAHomo sapiens 267augaaccacc aguccaagaa ucu
23268163RNAHomo sapiens 268gaagcccaag uuugaauugg gaaggcucau
ggagcuucau ggugaaggca guaguucugg 60aaaagccacu ggggaugaga caggugcuaa
aguuggacga gcugauggau augaaccacc 120aguccaagaa ucuguuuaaa
guucagacuu caaauagugg caa 16326917RNAHomo sapiens 269ggaagguugg
ggggugu 17270157RNAHomo sapiens 270cucucagcuc ugcagcuguc ugcggugggg
ggaagguugg ggggugucug gaggcauguu 60ccccucacca ccccccgugg gucucaggga
ggccgggugu gaccucaucu uucucauggu 120gcuauccugg ugcuauuggg
guggggagcu cccuccc 15727118RNAHomo sapiens 271uucuagguug uggcauuu
18272158RNAHomo sapiens 272auccuuuagc acguuuggau aaaguuggcc
uucuagguug uggcauuuca acugguuaug 60guccugcugu gaacacugcc aagcuggagc
cuggcucugu uugugccauc uuuggccugg 120gaggauuugg aucggggguu
accaugggcu guaaagug 15827324RNAHomo sapiens 273ugauauguuu
gauauugggu uguu 24274164RNAHomo sapiens 274acccccaaag cucuccugcc
ugcuucugug ugauauguuu gauauugggu uguuuaauua 60ggaaccaacu aaaugucaaa
cauauucuua cagcagcagg ugauucagca ccacccucuu 120ucauacuuca
aucucugggg cuccugucuc uuuuacugaa ccuc 16427522RNAHomo sapiens
275agaauacagc agaauuggcc uc 22276162RNAHomo sapiens 276cauuaauuag
guaauauuuu ccucauuucu uuacugcugc cauuuuccuc accaguauuc 60cagagauggu
cauagcucau uacucuacca ccaagaaccu aaaaggaauu agaauacagc
120agaauuggcc ucagugaaga gcuuaaaauu guucuccucg ua 16227721RNAHomo
sapiens 277cugauggaga gaagaaggca u 21278161RNAHomo sapiens
278ugguguggcc aaggccaaca cugagucgac cugauggaga gaagaaggca
uguguccacu 60ggcuccugau gaccaugcuu uggauguugc caacaaaauu gggaucaucu
aaucugaguc 120cagcuugcua auucuaaagg uauauaugua ucuuuucacc a
16127922RNAHomo sapiens 279acucggcgug gcgucggucg ug 22280162RNAHomo
sapiens 280acucccugac agauaucucc cucuuccauu ucaucaagac ccagcugagu
cacugucacu 60gccuaccaau cucgaccgga ccucgaccgg cucgucugug uugccaaucg
acucggcgug 120gcgucggucg ugguagauag gcggucaugc auacgaauuu uc
16228122RNAHomo sapiens 281augagguggc aagaaauggg cu 22282162RNAHomo
sapiens 282auauaaaaac auuaggucaa gguacagccu augagguggc aagaaauggg
cuacauuuuc 60uauauccggc aaaucucaca acaaccuuua ugaaaucuaa gggcucaagg
aggauuuagu 120aguaaaccaa gcgcagagug cuugguugaa uaaggccaug aa
16228325RNAHomo sapiens 283aauuuugaca gaugcucaag gcugu
25284165RNAHomo sapiens 284uuucauuuuc ugugauuauu uuuaaauuag
cuucugugua aacucacuaa cuuguuccca 60caugacaauu uauagcaguc caaagauuuu
uuuauagcca ugguuguuau aauuuugaca 120gaugcucaag gcuguuguuu
gcauuguucu ucagaauuuc aucuu 16528518RNAHomo sapiens 285aaaagcuggg
uugagaag 18286158RNAHomo sapiens 286cauuacacuc cagccugggc
aacaagagca aaacucuguc ucaaaaaaau gaaaagaaaa 60gaaaauaccu ccauggggcc
uucucuuccc aguucuuccu ggagucgggg aaaagcuggg 120uugagaaggu
gaaaagaaaa aacaaaccuu gacugggc 15828723RNAHomo sapiens
287auugaugguu aagcucagcu uuu 23288163RNAHomo sapiens 288ggaguagcug
uaacauuaug uggaaagcaa gugggagaau caugaaaaaa aauaauccca 60uagauggaga
agaauagaaa gaaggaaagg agcauugccu aguguugguu auugaugguu
120aagcucagcu uuuauuuauu caauaggccu gcagauguag acu 16328924RNAHomo
sapiens 289auggauugag augugaucaa aggc 24290164RNAHomo sapiens
290uguaggauuu uuuguuuuug uagcuaacuu auggauugag augugaucaa
aggcuuuauu 60aaauuuguac uucagcauau gauggcugcg uucugcauuu cauuccgcca
uaugccugga 120ccguucacac uuggguaucu gggcuuaggg agcauguagg cuuc
16429123RNAHomo sapiens 291auuuggcauu uggaagauag guu
23292163RNAHomo sapiens 292ucuagcucug uuauaaagaa aacauuuagg
aaauucucuc uuucucucuu ucaccuaucc 60uacuuuuugu guguccuuug uaguuuugca
ccaucauucc uaacgaauuu auuuggcauu 120uggaagauag guuagcaaaa
auuuuacuau auuugaaagg cua 16329323RNAHomo sapiens 293acugcugagg
aacugucacu ugu 23294163RNAHomo sapiens 294ggucaccagg cugagaaagc
aggagaugcu acugcugagg aacugucacu ugucauuuca 60agguccacuc cuccacccuc
uggcagcaug agucgcucug aaagauuuug aagcugggac 120aggagagggu
gagugaggug aggccuccgc augccagguu uuc 16329524RNAHomo sapiens
295aggugagggg caggaccuga aggu 24296164RNAHomo sapiens 296auucaucucu
gguuuucuug ccacccucug ggagucccca ucccauuuuc auccugagcc 60caaccaggcc
cugccauugg ccucuugucc cuuggcacac uuguacccac aggugagggg
120caggaccuga agguauuggc cuguucaaca aucagucauc augg 16429720RNAHomo
sapiens 297agugucugug ugugcuugcu 20298160RNAHomo sapiens
298acuaauaagc uacaaaacau uuaaaugacu agugucugug ugugcuugcu
aguauuauua 60uaccaucaga aaguaaaaau ggacauacau guuaugcauu aaacccacaa
gagagaaaac 120uugaggacug auuaauuuaa guaguaaaug aauccaagaa
16029920RNAHomo sapiens 299acgaagaugg cgaccguaac 20300160RNAHomo
sapiens 300acuguuuaua agucgguguu guaaaucuga ugugaauuuu uguuucuuuu
uucuuagauu 60uuugccuuua ugacgacagc uuguuauggu ugcaguuugg gucuggcuuu
acgaagaugg 120cgaccguaac acuccuuaga aacuggcagu cguauguuag
16030123RNAHomo sapiens 301aacuggaaug gcggcaaggu ccu
23302163RNAHomo sapiens 302aguaggcaac ugaggacuga uuucucaggg
ugauuagaaa ggaaagggug gcggccuccu 60uucauacuuc ggaaagucuu guucccauca
gccuuuccuc auggugccau aacuggaaug 120gcggcaaggu ccucuuuccu
gugccugugu cuuaaguuuc ugg 16330321RNAHomo sapiens 303aguugaguca
gggccugugu g 21304161RNAHomo sapiens 304gcgcccccaa aguguccccu
ccugcuguga cuucuagcca agaagacauu ucucccaugg 60ccaagugauc ucugauagau
ccuguaggac cacugaaguc agacaggaca aguugaguca 120gggccugugu
guccagugcg cagcaugcuu ggggagugac a 16130519RNAHomo sapiens
305gccgcccggg gccauggcg 19306159RNAHomo sapiens 306uaacgcaugc
gcggggaggg cggagcuggg cguugccgug gcuacuggga acgcauuuca 60cgggggcggg
gcgugguucc ggggcggggc gcggccgccg gaagugcgug gccgcccggg
120gccauggcga cacucagcuu cgucuuccug cugcugggg 15930724RNAHomo
sapiens 307aagucuaagu cuaacauucg gugu 24308164RNAHomo sapiens
308ucgaugggug uucuuuuaaa auacgguucu aagucuaagu cuaacauucg
guguaucuaa 60ccgaauguua auugauggag acaaggugau acggguucag aaaauagaau
ucagaaaaga 120aaaggaagaa uuggcaaaau ucagaaauca auuuuaagaa aaau
16430916RNAHomo sapiens 309cggcgggagc ccgggg 16310156RNAHomo
sapiens 310cccacggauu cgccccgccg cgccucuccg cgcguagauu ggccggagcg
aggcgaacgg 60gcccggccuu gguagccgcc gaccgagcgc uggcuguccu ggaaccuagg
cggcgggagc 120ccggggcgcc ucgcggcacg gaagagcggc gagaug
15631121RNAHomo sapiens 311acguggaugg cguggaggug c 21312161RNAHomo
sapiens 312ccuguucccc ccaaaaccca aggacacucu caugaucucc cggaccccug
aggucacgug 60cgugguggug gacgugagcc aggaagaccc cgagguccag uucaacuggu
acguggaugg 120cguggaggug cauaaugcca agacaaagcc gcgggaggag c
16131318RNAHomo sapiens 313guuuagacgg gcucacau 18314158RNAHomo
sapiens 314uuuuacauaa guagacacag gugggaaacu guuuagacgg gcucacauca
ccccauaaac 60aaauagguuu gguccuagcc uuucuauuag cucuuaguaa gauuacacau
gcaagcaucc 120ccauuccagu gaguucaccc ucuaaaucac cacgauga
15831521RNAHomo sapiens 315auuucugcag ucaggugaga c 21316161RNAHomo
sapiens 316uggaaaucac agcaacccau ugaaaacugc ccuccccacc agaacgugcu
acguucuuuc 60uucaugccua ugugugcucc auuccucauu ucuacuuggc ucaagaaaac
auuucugcag 120ucaggugaga cuuuuacaaa agaggagaaa aucaaugccu c
16131722RNAHomo sapiens 317aagaaugacc gcugaagaac gu 22318162RNAHomo
sapiens 318cugaucauag uauucuguca gauaaugccu aagaaugacc gcugaagaac
guugacccau 60uugaguaccc ggucucaguc gucauuuuua aguccaguga gcauuguggu
aguuguucuu 120agauugcagu uucuuauguu uugaguuuga aguugauuuu ca
16231920RNAHomo sapiens 319aaauaugagc cacugggugu 20320160RNAHomo
sapiens 320aaagucuuag aauggaaaaa guaaagaaau aucaacuucc aaguuggcaa
guaacuccca 60augauuuagu uuuuuucccc ccaguuugaa uugggaagcu gggggaaguu
aaauaugagc 120cacugggugu accagugcau uaauuugggc aaggaaagug
16032123RNAHomo sapiens 321aagguccucu gaggcagcag gcu
23322163RNAHomo sapiens 322gaagaucacu acaacaauuu gucugccucc
aagguccucu gaggcagcag gcucuggggc 60uucugcuguc cuuuggaggg ugucuucugg
guagagggau gggaaggaag ggacccuuac 120ccccggcucu ucuccugacc
ugccaauaaa aauuuauggu cca 16332321RNAHomo sapiens 323gucgaggucu
uugguggguu g 21324161RNAHomo sapiens 324caccccgugc cuuuugaucu
agcacagacc cuucaccccu caccucgaug cagccaguag 60cuuggauccu ugugggcaug
auccauaauc gguuucaagg uaacgauggu gucgaggucu 120uugguggguu
gaacuauguu agaaaaggcc auuaauuugc c 16132521RNAHomo sapiens
325gguuucgggu uugaaggcag c 21326161RNAHomo sapiens 326cccucuggca
ugguucauua gggccaauua auguggcugg guuauuugca acuuaaacug 60ggggauaaug
ucgcuugagg gagcguuuuc guuuuaggaa auauuguuuu gguuucgggu
120uugaaggcag cugucaaaaa agcggcaugg aaauucauug g 16132722RNAHomo
sapiens 327gcuggugagu gcaggcugcu uc 22328162RNAHomo sapiens
328gagcuaguac cuucuccccu uagcaacuuc cucauucuaa aaugggggug
gcagaaccau 60uguuuggcuc caguuguccu cagaaaggug gcuuccagau gccagugacu
gcuggugagu 120gcaggcugcu ucaguauuuc cuggccagcu gacaaggugu ua
16232920RNAHomo sapiens 329gacagaggug gcaucaagcu 20330160RNAHomo
sapiens 330aaucccuguu ugcuucaggg cgagaugugu gacagaggug gcaucaagcu
cuuacagucc 60caacccucca acggaaaugg gcgaagaucu caggaauggc aucggucaca
ggaaaucgau 120aguggcuggc ugcuagcaug gccacuuggg gcuuaggcag
16033123RNAHomo sapiens 331cugcccuggc ccgagggacc gac
23332163RNAHomo sapiens 332ggaaccugcu uggacaaguc uucuggcucg
accucgacau gcuccaucgg augaauuguu 60gguguuagcc cugcggcccc acgcaccagg
guaagagaga cucucgcuuc cugcccuggc 120ccgagggacc gacuggcugg
gccugccuuc ugcccagcuc acc 16333320RNAHomo sapiens 333ucgugucgcg
uggggggcgg 20334160RNAHomo sapiens 334ucgcgggucu guggcgcggg
gccccggugg ucgugucgcg uggggggcgg gugguugggg 60cguccgguuc gccgcgcccc
gccccggccc caccgguccc ggccgccgcc cccgcgcccg 120cucgcucccu
cccguccgcc cguccgcggc ccguccgucc 16033525RNAHomo sapiens
335gggaugccag gcaagugagc agguc 25336165RNAHomo sapiens
336ggucaacaag gugagucugg augaggggca gggaugccag gcaagugagc
aggucuggga 60gucaggccuu gcucaggccc uguucuucuc ccuugcagcu ucugucuggc
cccaaagaga 120ccccugcugc ccagagcccc accagaggcc ccucugacac caaga
16533725RNAHomo sapiens 337agggccguca ggacacggga ggguu
25338165RNAHomo sapiens 338cuguguugug uccugacacc uccaaguucu
agggccguca ggacacggga ggguuugggg 60acagaguguc cuuccucugu ccucucaucc
caguccugau ggccgcuugg ugagugucug 120gugcccuggu ggccugcccc
agcucucuuc uggcuuucug agcag 16533921RNAHomo sapiens 339gaaaagcugg
guugagaggg u 21340160RNAHomo sapiens 340uuacaauggu ucuaugagga
cguggcccca caguaaguug aggagcacug gguauguaug 60aauaaaaugg caugacaggc
cuucucuuuc caguucuucc cagaauuggg aaaagcuggg 120uugagagggu
aagaaaagaa aaacaaauaa auuuuuuaaa 16034117RNAHomo sapiens
341aacuagcuag ggguucg 17342157RNAHomo sapiens 342gcgagacucu
uuuuuucucc aggaccugcg gagcagccag gcuucaugag uuaaaugcag 60aucugaacca
uacccaguug ggauuggggu acacacucua cuccucugaa aacuagcuag
120ggguucgaac uuggugagag ggagaguggg acagagc 15734323RNAHomo sapiens
343ggguuugggg gaugucagag ggc 23344163RNAHomo sapiens 344aaguucugag
aguccaggag gcagaggcug ggguuugggg gaugucagag ggcaaaucug 60gggcuugggg
ggcccaggaa gcagagauga agguuuuaga gucuccagag aacaaaucug
120guacuuuuaa ggcccaggaa gcggaggcug gggucuuggg aaa 16334522RNAHomo
sapiens 345ugaagggaga ugugaagaag cc 22346162RNAHomo sapiens
346aauuugaaaa uccauauaaa gguacgucca cauuuauguu auuaugagug
agucauauug 60gugaagucag gacaauggcc ugucauuaga ucuuugauuc uuguuugcag
ugaagggaga 120ugugaagaag ccauguucuc ugaacgugcu gcuuggagga cu
16234720RNAHomo sapiens 347agaaagucca aguguucagg 20348160RNAHomo
sapiens 348ucucuaauua gcuuucccag uauacuucuu agaaagucca aguguucagg
acuuuuauac 60cuguuauacu uuggcuuggu uuccaugauu cuuacuuuau uagccuaguu
uaucaccaau 120aauacuugac ggaaggcuca guaauuaguu augaauaugg
16034918RNAHomo sapiens 349ugaacgggua uuuuacug 18350158RNAHomo
sapiens 350uuuuuuuuuu aacaccuaua uaucacccau ugaacgggua uuuuacugaa
cacaguacag 60uagacuguuu aaaacucaca uccugguaac uuucacuacu ugaaauuaca
aagugcuuuu 120guuaauugca uauuuuugcu cagccaucuu agaauugu
15835121RNAHomo sapiens 351cugagacaug cacuucuggu u 21352161RNAHomo
sapiens 352aagcuggccc uggcuggaga uggcuagccc cugagacaug cacuucuggu
uuugaaauga 60cucugucugu ggggcagcag aaacuagaga aggcaagugg cugccccacc
ccaaggcgug 120accaggagga acagccugca gcucacucca ugccacacgg g
16135321RNAHomo sapiens 353aauugaggau gugugagguu u 21354161RNAHomo
sapiens 354uugguggugu guauaagaau guuucuugcu aauugaggau gugugagguu
uaaggcugug 60agcugaucuu ugaaaaauag uuuccuguuu cuaaagugac auuacccagu
auuugcuuac 120ugcuuugugc cuuaucuccc gcuuucuuuu uaguauuucu g
16135524RNAHomo sapiens 355aguuucuaug aguguauacc auuu
24356164RNAHomo sapiens 356ucauucugag gucacauaac acauaaaauu
aguuucuaug aguguauacc auuuaaagaa 60uuuuuuuuuc aguaaaaggg aauauuacaa
uguuggagga gagauaaguu auagggagcu 120ggauuucaaa acguggucca
agauucaaaa auccuauuga uagu 16435725RNAHomo sapiens 357agguggagau
caagcccgag augau 25358165RNAHomo sapiens 358ugggcgucua caacggcaag
accuucaacc agguggagau caagcccgag augaucgacc 60acuaccuggg cgaguucucc
aucaccuaca agcccauaaa gcacggcggg cccggcaucg 120gggccagcca
cuccucccgc uucaucccuc ucaagcagug gcuca 16535921RNAHomo sapiens
359gaagacagca auaaccacag u 21360161RNAHomo sapiens 360guagcccaca
uggauagcac aguugucaga caagauuccu ucagauuccg aguugccuac 60cgguuguuuu
cguuguuguu guuguuguuu uucuuuuucu uuuuuuuuuu gaagacagca
120auaaccacag uacauauuac uguaguucuc uauaguuuua c 16136124RNAHomo
sapiens 361guuucuguug aguguggguu uagu 24362164RNAHomo sapiens
362aguccgugcg agaauaauga uguaugcuuu guuucuguug aguguggguu
uaguaauggg 60guuugugggg uuuucuucua agccuucucc uauuuauggg gguuuaguac
ugauuguuag 120cggugugguc ggguguguua uuauucugaa uuuuggggga gguu
16436321RNAHomo sapiens 363aucgccgugg agugggagag c 21364161RNAHomo
sapiens 364ugccugguca aaggcuucua ccccagcgac aucgccgugg agugggagag
cagcgggcag 60ccggagaaca acuacaacac cacgccuccc augcuggacu ccgacggcuc
cuucuuccuc 120uacagcaagc ucaccgugga caagagcagg uggcagcagg g
16136523RNAHomo sapiens 365ggaugaagug cacugaggcu cuu
23366163RNAHomo sapiens 366guaagauggu uaucgauagu aguuaaugau
ggaugaagug cacugaggcu cuuaaaagau 60acuuaggauu uuugacuuua cucuguaggu
ucuaaaguaa acauauauga gguuuuuaau 120uucucagaua cuauaccugc
agcucuuuuu gcugacucaa gau 16336727RNAHomo sapiens 367uccuguuggc
cgaguggaga cuggugu 27368165RNAHomo sapiens 368ucuacaaaau uggugguauu
gguacuguuc cuguuggccg aguggagacu gguguucuca 60aaccuggugu gguggucacc
uuugcuccag ucaacauuac aacagaagua aaaucugucg 120aaaugcacca
ugaagcuuug agugaagcuc uuccugggga caaug 16536924RNAHomo sapiens
369guggagucug ucaucgaggu gcgu 24370164RNAHomo sapiens 370gucacucagg
acagacuucu ugaaguaggu ggcccuucaa cugagcugaa gggaagagaa 60gggcauugca
ggcugaggga ugaucccagg gugcccccca gaucuaaagu guggagucug
120ucaucgaggu gcguagccuc uccagaggug ucacuguguu ccau 16437124RNAHomo
sapiens 371uugcagcugc cugggaguga cuuc 24372164RNAHomo sapiens
372cugcaaacag ccuuuccacu gacgcagugc cuugggggcu cugccaagcg
accccuagaa 60uggggauugu ggggggucgc ucuaggcacc gcagcacugu gcuggggaug
uugcagcugc 120cugggaguga cuucacacag uccucucugc cuccaggguc accc
16437322RNAHomo sapiens 373ugcagccagc gucccaugcu cg 22374162RNAHomo
sapiens 374guggauauga gugaagacgg ggcaggcagg ccacaucucu uagaagagga
aggugauugc 60cacgucuccu uccuccaugc ugauggcaag gcgugcgggc uguguucucu
ugcagccagc 120gucccaugcu cgguggcccc agaaaaguca guguguaggc cu
16237517RNAHomo sapiens 375aguuuugugu guuggcu 17376157RNAHomo
sapiens 376aggaaacaca agaagaaacg ccuggugcag agccccaauu ccuacuucau
ggaugugaaa 60ugcccaagau gcuauaaaau caccacgguc uuuagccaug cacaaacggu
aguuuugugu 120guuggcugcu ccacuguccu cugccagccu auaggag
15737722RNAHomo sapiens 377acggccaagc agaaaauguu uu 22378162RNAHomo
sapiens 378cauagauaug uauucagcuu gucuucaaau acggccaagc agaaaauguu
uuauauuuua 60uaaaucaucu uuugacucug uauuuaaauu cuaugauacu gaaaauaaag
gcauucugga 120aaaauacuga cugauuuugg ugcagaaguu uugaguauca ag
16237924RNAHomo sapiens 379gauuagggug cuuagcuguu aacu
24380164RNAHomo sapiens 380ggcggcggga gaaguagauu gaagccaguu
gauuagggug cuuagcuguu aacuaagugu 60uuguggguuu aagucccauu ggucuaguaa
gggcuuagcu uaauuaaagu ggcugauuug 120cguucaguug augcagagug
ggguuuugca guccuuagcu guug 16438121RNAHomo sapiens 381ccuucgaggc
ggcugagacc c 21382161RNAHomo sapiens 382cguucuccgc acuccugcuc
cgcgagggcc ccuucgaggc ggcugagacc cgagugccgg 60acucccgccg cuggagcggg
gcucggguuc ggcagccgga aggaggugug cccccggggc 120gcuugggggc
gccugagguc ccgaggggag gcaagauggg a 16138323RNAHomo sapiens
383aggugguggc agcuggaggg acc 23384163RNAHomo sapiens 384agccugugac
cccucgggac ugccuggugc aggugguggc agcuggaggg acccaugcag 60cacccagguc
agagcagacc cuccccugcc ggccugcgcc agcuggaccu gauggccccc
120uguggcgccu ugaccugcug ggccaggcug cccugggacu cuc 16338523RNAHomo
sapiens 385agaagaagga cggcaagaag cgc 23386163RNAHomo sapiens
386ggccccgcau ccgcguucgu cuaggcgcuc uugucaccuc gccaugccgg
agccaucgcg 60ggcggcuccg gcuucuaaaa agggcuccaa gaaggccauu accaaggcgc
agaagaagga 120cggcaagaag cgcaagcgcg gccgcaagga gagcuauucu auc
16338723RNAHomo sapiens 387aaaagcaaau guugggugaa cgg
23388163RNAHomo sapiens 388gaaacaucuu aaugcacagc cacaaguuac
aaugcaacag ccugcugcuc auguacaagg 60ucaggaaccu uugacugcuu ccauguuggu
aucugcccau gcucaagagc aaaagcaaau 120guugggugaa cggcuguuuc
cucuuauuca agccaugcac cuu 16338919RNAHomo sapiens 389ggcggagggg
ccgcgggcc 19390159RNAHomo sapiens 390cagaguggga ccggcagcuc
ccagacuuga ggcggagggg ccgcgggccg gagcucccug 60cagccgcuag ccugggaaga
cuggagugcg cugcccaccg agggucugcg ccgcgccggc 120cgccccgggc
cgcuuugugc gcgcccgcgc ggucuguac 15939119RNAHomo sapiens
391uugcagcugc cugggagug 19392159RNAHomo sapiens 392cugcaaacag
ccuuuccacu gacgcagugc cuugggggcu cugccaagcg accccuagaa 60uggggauugu
ggggggucgc ucuaggcacc gcagcacugu gcuggggaug uugcagcugc
120cugggaguga cuucacacag uccucucugc cuccagggu 15939324RNAHomo
sapiens 393ucucuccagg ugacagaaag ggcu 24394164RNAHomo sapiens
394acauaaaauc uuaucuaugu gcagcaugac ucucuccagg ugacagaaag
ggcucuagac 60agcugagagg accugaucau guagggaggg acggggaggg gagccaggac
ccaggagcug 120cauggcugua agaggaaggu ccuuggaggg uaucagcagu cuca
16439521RNAHomo sapiens 395cguucuugcu cugccucggu c 21396161RNAHomo
sapiens 396gggcagaagu cugagccagu guuucaucau cguucuugcu cugccucggu
cuguacaucu 60gugaaauggg acucccucuc uguuguggag gcccugggga cagcugggag
gacuggaggg 120guggugggga gguugugguc cuuauuagac auucagauac c
16139722RNAHomo sapiens 397augccaagag ggccaguguc uu 22398162RNAHomo
sapiens 398aaacaauaaa aaacuggcug cuaucgaagc ccuaaaugau ggugaacucc
agaaagccau 60ugacuuauuc acagaugcca ucaagcugaa uccucacuug gcccuuuugu
augccaagag 120ggccaguguc uucgucaaau uacagaagcc aaauacugcc au
16239922RNAHomo sapiens 399uuuugugugu guguuuguuu uu 22400162RNAHomo
sapiens 400cugguuauau caggauaaau ucauaaaggg uuuugugugu guguuuguuu
uuguuguugu 60uguuuagggu uuuuuuuuuu uaaacagggu cuugcuuugu ugcccaggau
gaaaugcaau 120cacacacaau cauggcucau ugcaucacua ucuauguauu ca
16240125RNAHomo sapiens 401augaggaaca cugacuuuau uaagc
25402165RNAHomo sapiens 402ucaguuaagc caauacauuu aaaguuuugc
augaggaaca cugacuuuau uaagcauuuu 60cagauguggu gguuguauuu uugccccaag
aaguguuugg auaaccacac aaaagcauga 120ugaaaaggcu ucuuguaguc
ccauaauuuc uugugaacua auguu 16540324RNAHomo sapiens 403gggucggagu
uagcucaagc gguu 24404164RNAHomo sapiens 404uuuagaaguu ucagucgcac
acuccuaccc gggucggagu uagcucaagc gguuaccucc 60ucaugccgga cuuucuaucu
guccaucucu gugcuggggu ucgagacccg cgggugcuua 120cugacccuuu
uaugcaauaa auucgguaua aucugucacu cuga 16440519RNAHomo sapiens
405ggagguucag aguuggaag 19406159RNAHomo sapiens 406aguugcgaca
agacagaguu ggagaauaga ggagguucag aguuggaaga aaugggagua 60ggugauggca
acaccgaguu gucagaguga gcugaggcaa cauccucuac uucuagcuca
120cugaugaaaa uauccaggau agcgggucug ggguccagu 15940723RNAHomo
sapiens 407gagagucuca ggaaagaaag guc 23408163RNAHomo sapiens
408auaaugagua aaaauguuca ucuuuaauaa agcaaaaaua gagcaaccca
ccaaauaguu 60aacacuugcc uggagagauu uaggaacacc aguauuccac ugagauugcu
gagagucuca 120ggaaagaaag gucuaacuua aauuguauuu uaccauuucu gag
16340943DNAartificialPCR primer 409gcctccctcg cgccatcagc tnnnngacct
tggctgtcac tca 4341061DNAartificialPCR primer 410gccttgccag
cccgctcaga cgagacatcg ccccgctttt tttttttttt tttttttttt 60t
6141121RNAHomo sapiens 411acccgucccg uucguccccg g 2141296RNAHomo
sapiens 412ccggccgcug ggcgcacccg ucccguucgu ccccggacgu ugcucucuac
cccgggaacg 60ucgagacugg agcgcccgaa cugagccacc uucgcg 9641319RNAHomo
sapiens 413accgggugcu guaggcuuu 1941494RNAHomo sapiens
414ucucggaagc uaagcagggu cgggccuggu uaguacuugg acgggagacc
gccugggaau 60accgggugcu guaggcuuuu ucuuuggcuu uuug 9441594RNAHomo
sapiens 415ucuuggaagc uaagcagggu cgggccuggu uaguacuugg augggagacc
accugggaau 60accgggugcu guaggcuuug gccgggcgug gugg 9441629RNAHomo
sapiens 416acgcggguga ugcgaacugg agucugagc 29417104RNAHomo sapiens
417ggcguagggg ggccggccug cugugaugac auuccaauua aagcacgugu
uagacugcug 60acgcggguga ugcgaacugg agucugagcc ugcccgagcg gagc
10441820RNAHomo sapiens 418acucggcgug gcgucggucg 2041995RNAHomo
sapiens 419cacugucacu gccuaccaau cucgaccgga ccucgaccgg cucgucugug
uugccaaucg 60acucggcgug gcgucggucg ugguagauag gcggu 9542021RNAHomo
sapiens 420acucggcgug gcgucggucg u 2142124RNAHomo sapiens
421acucggcgug gcgucggucg uggu 2442221RNAHomo sapiens 422agagcagugu
guguugccug g 2142396RNAHomo sapiens 423ugcugcaggu guuggagagc
aguguguguu gccuggggac uguguggacu gguaucaccc 60agacagcuug cacugacucc
agacccugcc gucaug 9642422RNAHomo sapiens 424agaguugagu cuggacgucc
cg 2242597RNAHomo sapiens 425cgccccgggc cgcggcuccu gauuguccaa
acgcaauucu cgagucuaug gcuccggccg 60agaguugagu cuggacgucc cgagccgccg
cccccaa 9742619RNAHomo sapiens 426aggacggugg ccauggaag
1942794RNAHomo sapiens 427aguaccaaga aguuaucauu uccauaugac
ugucauugcu uaaaacuagc uaguaugagc 60aggacggugg ccauggaagu cgaaauucgc
uaag 9442820RNAHomo sapiens 428aggacggugg ccauggaagu 2042921RNAHomo
sapiens 429aguuggugga gugauuuguc u 2143022RNAHomo sapiens
430aguuggugga gugauuuguc ug 2243123RNAHomo sapiens 431aguuggugga
gugauuuguc ugg 2343224RNAHomo sapiens 432aguuggugga gugauuuguc uggu
2443396RNAHomo sapiens 433gggauugaca gauugacagc ucuuucucga
uucugugggu gguggugcau ggccauucuu 60aguuggugga gugauuuguc ugguuaauuc
ugauaa 9643419RNAHomo sapiens 434agaacgcggu cugaguggu
1943594RNAHomo sapiens 435ugcaacugcc gucagccauu gaugaucguu
cuucucuccg uauuggggag ugagagggag 60agaacgcggu cugagugguu uuuccuucuu
gaug 9443619RNAHomo sapiens 436augaggaugg auagcaagg 1943794RNAHomo
sapiens 437auacguguaa uugugaugag gauggauagc aaggaagccg cucccaccug
acccucacgg 60ccuccguguu accuguccuc uaggugggac gcuc 9443820RNAHomo
sapiens 438cggagcuggg gauugugggu 2043995RNAHomo
sapiens 439gggugugaga agccggaucc uguggugacc caguggccua auggauaagg
caucagccuc 60cggagcuggg gauugugggu ucgaguccca ucugg 9544022RNAHomo
sapiens 440cggcggcucc agggaccugg cg 2244197RNAHomo sapiens
441ccgggagcgg ggaggcggcg gcuccaggga ccuggcggcc gccgaucggg
gcugcgaggc 60cccauggcgc cgcccccagc cccgcuccug gcgccga
9744220RNAHomo sapiens 442cggcggggac ggcgauuggu 2044395RNAHomo
sapiens 443gggccgggaa ugccgcggcg gggacggcga uugguccgua uguguggugc
caccggccgc 60cggcuccgcc ccggcccccg ccccacacgc cgcau 9544429RNAHomo
sapiens 444cggggaucgc cgagggccgg ucggccgcc 29445104RNAHomo sapiens
445uggugggggg agccgcgggg aucgccgagg gccggucggc cgccccgggu
gccgcgcggu 60gccgccggcg gcggugaggc cccgcgcgug ugucccggcu gcgg
104446104RNAHomo sapiens 446ggggaggaga cgguuccggg ggaccggccg
cgacugcggc ggcgguggug gggggagccg 60cggggaucgc cgagggccgg ucggccgccc
cgggugccgc gcgg 10444723RNAHomo sapiens 447cucggcgugg cgucggucgu
ggu 2344898RNAHomo sapiens 448acugucacug ccuaccaauc ucgaccggac
cucgaccggc ucgucugugu ugccaaucga 60cucggcgugg cgucggucgu gguagauagg
cggucaug 9844919RNAHomo sapiens 449cugcccagug cucugaaug
1945094RNAHomo sapiens 450aaugcagugu gauuucugcc cagugcucug
aaugucaaag ugaagaaauu cagagaagcc 60uggguagccg ggcguggugg cucacaccug
uaau 9445127RNAHomo sapiens 451cugcccagug cucugaaugu caaagug
2745224RNAHomo sapiens 452cugcccuggc ccgagggacc gacu 2445399RNAHomo
sapiens 453augaauuguu gguguuagcc cugcggcccc acgcaccagg guaagagaga
cucucgcuuc 60cugcccuggc ccgagggacc gacuggcugg gccugccuu
9945418RNAHomo sapiens 454cuggaggagc uggccugu 1845593RNAHomo
sapiens 455cuucucccca gccagaggug gagccaagug guccagcguc acuccagugc
ucagcugugg 60cuggaggagc uggccugugg cacagcccug agu 9345621RNAHomo
sapiens 456cuuggcaccu agcaagcacu c 2145796RNAHomo sapiens
457gcacccagau cagugcuugg caccuagcaa gcacucagua aauauuuguu
gagugccugc 60uaugugccag gcauugugcu gagggcuuug ugggga 9645821RNAHomo
sapiens 458gacuauagaa cuuucccccu c 2145996RNAHomo sapiens
459aguugguaga gggcagaggg augaggggga aaguucuaua guccugagau
cuaauuacag 60gacuauagaa cuuucccccu caucccucua cccuua 9646022RNAHomo
sapiens 460gagagcagug uguguugccu gg 2246197RNAHomo sapiens
461gugcugcagg uguuggagag cagugugugu ugccugggga cuguguggac
ugguaucacc 60cagacagcuu gcacugacuc cagacccugc cgucaug
9746217RNAHomo sapiens 462gaggaaggug gggaugc 1746392RNAHomo sapiens
463gagggcggga gacaaaaucu cgcaauucug accugccuuu ggacauaauu
gaggcuuuau 60gaggaaggug gggaugcggg aguggcgauc cc 9246430RNAHomo
sapiens 464gagugagagg gagagaacgc ggucugagug 30465105RNAHomo sapiens
465ggcguugcuu ggcugcaacu gccgucagcc auugaugauc guucuucucu
ccguauuggg 60gagugagagg gagagaacgc ggucugagug guuuuuccuu cuuga
10546621RNAHomo sapiens 466gaugccuggg aguugcgauc u 2146796RNAHomo
sapiens 467ugugagacga auuuuugagc ggguaaaggu cgcccucaag gugacccgcc
uacuuugcgg 60gaugccuggg aguugcgauc ugcccgaccu uauuca 9646823RNAHomo
sapiens 468gaugccuggg aguugcgauc ugc 2346918RNAHomo sapiens
469gcaugggugg uucagugg 1847093RNAHomo sapiens 470gugaacggcg
gcugguggcg aguuccgcug ugccagcuuc cguuggcguu ugccaucggu 60gcaugggugg
uucaguggua gaauucucgc cug 9347119RNAHomo sapiens 471gcaugggugg
uucaguggu 1947231RNAHomo sapiens 472gcaugggugg uucaguggua
gaauucucgc c 3147319RNAHomo sapiens 473gcauuggugg uucaguggu
1947494RNAHomo sapiens 474ccaucaccac ugugaaucag agcaacaaaa
cagcuggagg cagaacagca cucagcugga 60gcauuggugg uucaguggua gaauucucgc
cugc 9447519RNAHomo sapiens 475gcguuggugg uauaguggu 1947694RNAHomo
sapiens 476cgaggaagua gugaccugcc acuggccacc ugcggaacca gaguucccca
cuggagggcc 60gcguuggugg uauaguggug agcauagcug ccuu 9447723RNAHomo
sapiens 477gcguuggugg uauaguggug agc 2347819RNAHomo sapiens
478gcuguggcug ugacuggcg 1947994RNAHomo sapiens 479ccgacccggg
ccugggcugu ggcugugacu ggcgcugccg ugggcgccgc agcccucgcg 60ggagccggac
gcgguaaugc cccagcggcg cagc 9448019RNAHomo sapiens 480ggacgguggc
cauggaagu 1948194RNAHomo sapiens 481guaccaagaa guuaucauuu
ccauaugacu gucauugcuu aaaacuagcu aguaugagca 60ggacgguggc cauggaaguc
gaaauucgcu aagg 9448219RNAHomo sapiens 482ggggauguag cucaguggu
1948394RNAHomo sapiens 483cugcugagag uaggugggga uguagcucag
ugguagagcg caugcuuugc auguaugagg 60ccccggguuc gauccccggc aucuccagug
uagu 9448418RNAHomo sapiens 484gguucccuca gaccuggu 1848593RNAHomo
sapiens 485gccgacagcc agcuggaucu ccugucccga gcccugggua cugggggugc
cccugaguug 60gguucccuca gaccugguga ucgggcccug gag 9348626RNAHomo
sapiens 486ggaaaguuug gcugcgcggg uucccc 26487101RNAHomo sapiens
487cucccucucc ucccccgggg uucggugcgc ggccggggcc ggaguucgcu
gcaagucggc 60ggaaaguuug gcugcgcggg uucccccgaa guucaggugc g
10148825RNAHomo sapiens 488ggaauaccgg gugcuguagg cuuuu
25489100RNAHomo sapiens 489gucugaucuc agaagcuaag caaggucggg
ucuaguuagu acuuggaugg gagacugccu 60ggaauaccgg gugcuguagg cuuuuggccu
aucguucccu 10049023RNAHomo sapiens 490gugaacgggc gccaucccga ggc
2349198RNAHomo sapiens 491agagagcccc ggggugcaga uccuugggag
cccuguuaga cucuggauuu uacacuugga 60gugaacgggc gccaucccga ggcuuugcac
aggggcaa 9849224RNAHomo sapiens 492gugaacgggc gccaucccga ggcu
2449325RNAHomo sapiens 493gugaacgggc gccaucccga ggcuu
2549426RNAHomo sapiens 494gugaacgggc gccaucccga ggcuuu
2649521RNAHomo sapiens 495guugguggag cgauuugucu g 2149696RNAHomo
sapiens 496ggauugacag auugauagcu cuuucucgau uccgugggug guggugcaug
gccguucuua 60guugguggag cgauuugucu gguuaauucc gauaac 9649722RNAHomo
sapiens 497guugguggag cgauuugucu gg 2249823RNAHomo sapiens
498guugguggag cgauuugucu ggu 2349920RNAHomo sapiens 499guugguggag
ugauuugucu 2050095RNAHomo sapiens 500ggauugacag auugacagcu
cuuucucgau ucugugggug guggugcaug gccauucuua 60guugguggag ugauuugucu
gguuaauucu gauaa 9550121RNAHomo sapiens 501guugguggag ugauuugucu g
2150218RNAHomo sapiens 502gaacgcgguc ugaguggu 1850393RNAHomo
sapiens 503gcaacugccg ucagccauug augaucguuc uucucuccgu auuggggagu
gagagggaga 60gaacgcgguc ugagugguuu uuccuucuug aug 9350420RNAHomo
sapiens 504uaccgggugc uguaggcuuu 2050595RNAHomo sapiens
505aucuuggaag cuaagcaggg ucgggccugg uuaguacuug gaugggagac
caccugggaa 60uaccgggugc uguaggcuuu ggccgggcgu ggugg 9550695RNAHomo
sapiens 506aucucggaag cuaagcaggg ucgggccugg uuaguacuug gacgggagac
cgccugggaa 60uaccgggugc uguaggcuuu uucuuuggcu uuuug 9550720RNAHomo
sapiens 507uccuguacug agcugccccg 2050895RNAHomo sapiens
508auccucccug gggcauccug uacugagcug ccccgaggcc cuucaugcug
cccagcucgg 60ggcagcucag uacaggauac ucgggguggg aguca 9550921RNAHomo
sapiens 509ucgaggagcu cacagucuag u 2151096RNAHomo sapiens
510acaucaagug acugugcuug gcuguggggc uaccaagaug aagaaggaau
gcuccugccc 60ucgaggagcu cacagucuag ugggagggaa caaugc 9651120RNAHomo
sapiens 511ucggacaccg gcgcgucucu 2051295RNAHomo sapiens
512ggggcucgcg gacccggccc agagggcggc gguggcggca gcuacuuuuc
uggucagggc 60ucggacaccg gcgcgucucu caagcucgcc ucuuc 9551319RNAHomo
sapiens 513ucugcccagu gcucugaau 1951494RNAHomo sapiens
514uaaugcagug ugauuucugc ccagugcucu gaaugucaaa gugaagaaau
ucagagaagc 60cuggguagcc gggcguggug gcucacaccu guaa 9451521RNAHomo
sapiens 515ucugcaagug ucagaggcga g 2151696RNAHomo sapiens
516guuccuugug uucuuccagu ccgcccucug ucaccuugca gacggcuuuc
ucuccgaaug 60ucugcaagug ucagaggcga ggaguggcag cugcau 9651722RNAHomo
sapiens 517ucugcaagug ucagaggcga gg 2251822RNAHomo sapiens
518ucuucucugu uuuggccaug ug 2251997RNAHomo sapiens 519aacauuagga
gaguaucuuc ucuguuuugg ccaugugugu acucacagcc ccucacacau 60ggccgaaaca
gagaaguuac uuuccuaaua uuugccu 9752022RNAHomo sapiens 520ugauauguuu
gauauugggu ug 2252197RNAHomo sapiens 521cugccugcuu cugugugaua
uguuugauau uggguuguuu aauuaggaac caacuaaaug 60ucaaacauau ucuuacagca
gcaggugauu cagcacc 9752223RNAHomo sapiens 522ugauugucca aacgcaauuc
ucg 2352398RNAHomo sapiens 523ccgggccgcg gcuccugauu guccaaacgc
aauucucgag ucuauggcuc cggccgagag 60uugagucugg acgucccgag ccgccgcccc
caaaccuc 9852422RNAHomo sapiens 524ugcuggauca gugguucgag uc
2252597RNAHomo sapiens 525guugugcugu ccaggugcug gaucaguggu
ucgagucuga gccuuuaaaa gccacucuag 60ccacagaugc agugauugga gccaugacaa
gucccca 9752620RNAHomo sapiens 526uuagggcccu ggcuccaucu
2052795RNAHomo sapiens 527accuaccuaa cuggguuagg gcccuggcuc
caucuccuuu aggaaaaccu ucugugggga 60guggggcuuc gacccuaacc caggugggcu
guaac 9552822RNAHomo sapiens 528uucucuguuu uggccaugug ug
2252997RNAHomo sapiens 529cauuaggaga guaucuucuc uguuuuggcc
auguguguac ucacagcccc ucacacaugg 60ccgaaacaga gaaguuacuu uccuaauauu
ugccucc 9753017RNAHomo sapiens 530uucugcccag ugcucug 1753192RNAHomo
sapiens 531uaccucccag cuguguucug cccagugcuc ugcugacugg acgccucugu
guccuuagac 60caugugcagg gccugaugca ggaaagagcu ga 9253220RNAHomo
sapiens 532uucugcccag ugcucugaau 2053395RNAHomo sapiens
533uuaaugcagu gugauuucug cccagugcuc ugaaugucaa agugaagaaa
uucagagaag 60ccuggguagc cgggcguggu ggcucacacc uguaa 9553424RNAHomo
sapiens 534uugcagcugc cugggaguga cuuc 2453599RNAHomo sapiens
535accccuagaa uggggauugu ggggggucgc ucuaggcacc gcagcacugu
gcuggggaug 60uugcagcugc cugggaguga cuucacacag uccucucug
9953621RNAHomo sapiens 536uuggccaugg ggcugcgcgg g 2153796RNAHomo
sapiens 537ucguccagcg agggcgcgcu ggcccugggc agcguguggc ugaaggucac
cauguucucc 60uuggccaugg ggcugcgcgg ggccagcagg uccacg 9653820RNAHomo
sapiens 538uuggggaaac ggccgcugag 2053995RNAHomo sapiens
539cagcuccgca gggguuuggg gaaacggccg cugagugagg cgucggcugu
guuucucacc 60gcggucuuuu ccucccacuc uuggcugguu ggacc 9554021RNAHomo
sapiens 540uuggggaaac ggccgcugag u 2154119RNAHomo sapiens
541uugguggagc gauuugucu 1954294RNAHomo sapiens 542gauugacaga
uugauagcuc uuucucgauu ccgugggugg uggugcaugg ccguucuuag 60uugguggagc
gauuugucug guuaauuccg auaa 9454320RNAHomo sapiens 543uugguggagc
gauuugucug 2054420RNAHomo sapiens 544uuuccggcuc gcgugggugu
2054595RNAHomo sapiens 545ccggcggggc gaagcccgcg gcugcuggac
ccacccggcc gggaauagug cuccugguug 60uuuccggcuc gcgugggugu gucggcggcg
gggcc 9554617RNAHomo sapiens 546aaagcugggu gagaggg 1754792RNAHomo
sapiens 547ccugagcaag gucacaaagc ugggugagag gggcugggau ucacauucaa
gcacuguguu 60ccuaggcccc uugaccccug gcaucugugg gg 9254817RNAHomo
sapiens 548aaagcugggu ugagagg 1754918RNAHomo sapiens 549aaagcugggu
ugagaggg 1855019RNAHomo sapiens 550aaagcugggu ugagagggc
1955192RNAHomo sapiens 551cugaaauguc uucaaaugua ucaauaagcc
uucucuuccc aguucuucuu ggagucagga 60aaagcugggu ugagaggagc agaaaagaaa
aa 9255292RNAHomo sapiens 552guauguauga auaaaauggc augacaggcc
uucucuuucc aguucuuccc agaauuggga 60aaagcugggu ugagagggua agaaaagaaa
aa 9255392RNAHomo sapiens 553ucuucuaccu aagaauucug ucucuuaggc
uuucucuucc cagauuuccc aaaguuggga 60aaagcugggu ugagagggca aaaggaaaaa
aa 9255492RNAHomo sapiens 554gaaacuagug uuucaaaaau aaauuaaucc
cucucuuucu aguucuuccu agagugagga 60aaagcugggu ugagagggca aacaaauuaa
cu 9255518RNAHomo sapiens 555aaccuuggag agcugagc 1855693RNAHomo
sapiens 556cgcggugggg aagccaaccu uggagagcug agcgugcgac cggcccggcg
cgggggucuc 60cgggagcugg cgagucgcua gcaccgaguc aca 9355718RNAHomo
sapiens 557aagcuggguu gagagggc 1855893RNAHomo sapiens 558cuucuaccua
agaauucugu cucuuaggcu uucucuuccc agauuuccca aaguugggaa 60aagcuggguu
gagagggcaa aaggaaaaaa aaa 93
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