Methylation Altered Dna Sequences As Markers Associated With Human Cancer

Abstract

There is disclosed 103 novel methylation-altered DNA sequences ("marker sequences") that have distinct methylation patterns in cancer, compared to normal tissue. In many instances, these marker sequences represent novel sequences not found in the GenBank data base, and none of these marker sequences have previously been characterized with respect to their methylation pattern in human cancers including, but not limited to those of bladder and prostate. These 103 sequences have utility as diagnostic, prognostic and therapeutic markers in the treatment of human cancer, and as reagents in kits for detecting methylated CpG-containing nucleic acids.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application is a continuation of U.S. patent application Ser. No. 09/699,243 filed Oct. 27, 2000 (issued as ______).

TECHNICAL FIELD OF THE INVENTION

[0002] The present invention relates to novel human DNA sequences that exhibit altered methylation patterns (hypermethylation or hypomethylation) in cancer patients. These novel methylation-altered DNA sequences are useful as diagnostic, prognostic and therapeutic markers for human cancer.

BACKGROUND

[0003] The identification of early genetic changes in tumorigenesis is a primary focus in molecular cancer research. Characterization of the nature and pattern of cancer-associated genetic alterations will allow for early detection, diagnosis and treatment of cancer. Such genetic alterations in vertebrates fall generally into one of three categories: gain or loss of genetic material; mutation of genetic material; or methylation at cytosine residues in CpG dinucleotides within "CpG islands." Among these, DNA methylation is uniquely reversible, and changes in methylation state are known to affect gene expression (e.g., transcriptional initiation of genes where CpG islands located at or near the promoter region) or genomic stability.

[0004] Methylation of CpG dinucleotides within CpG islands. DNA, in higher order eukaryotic organisms, is methylated only at cytosine residues located 5' to guanosine residues in CpG dinucleotides. This covalent modification of the C-5 position of the cytosine base by the enzyme DNA (cytosine-5)-methyltransferase results in the formation of 5-methylcytosine (5-mCyt), and gives this base unique properties (e.g., susceptibility to undergo spontaneous deamination). This enzymatic conversion is the only epigenetic modification of DNA known to exist in vertebrates, and is essential for normal embryonic development (Bird, A. P., Cell 70:5-8, 1992; Laird & Jaenisch, Human Molecular Genetics 3:1487-1495, 1994; Li et al., Cell 69:915-926, 1992).

[0005] The presence of 5-mCyt at CpG dinucleotides has resulted in the 5-fold depletion of this sequence in the genome during the course of vertebrate evolution (Schroderet & Gartler, Proc. Nat. Acad. Sci. USA 89:957-961, 1992), presumably due to spontaneous deamination of 5-mCyt to Thymidine. Certain areas of the genome, however, do not show such depletion, and are referred to as "CpG islands" (Bird, A. P., Nature 321:209-213, 1986; Gardiner-Garden & Frommer, J. Mol. Biol. 196:261-282, 1987). These CpG islands comprise only approximately 1% of the vertebrate genome, yet account for about 15% of the total number of genomic CpG dinucleotides (Antequera & Bird, Proc. Nat. Acad. Sci. USA 90:11995-11999, 1993). CpG islands contain the expected (i.e., the non-evolutionarily depleted) frequency of CpGs (with an Observed/Expected Ratio.sup.1>0.6), are GC-rich (with a GC Content.sup.2>0.5) and are typically between about 0.2 to about 1 kb in length. .sup.1 Calculated as: [number of CpG sites/(number of C bases.times.number of G bases)].times.band length for each fragment..sup.2 Calculated as: (number of C bases+number of G bases)/band length for each fragment.

[0006] Methylation within CpG islands affects gene expression. CpG islands are located upstream of many housekeeping and tissue-specific genes, but may also extend into gene coding regions (Cross & Bird, Current Opinions in Genetics and Development 5:309-314, 1995; Larsen et al., Genomics 13:1095-1107, 1992). The methylation of cytosines within CpG islands in somatic tissues is believed to affect gene expression. Methylation has been inversely correlated with gene activity and may lead to decreased gene expression by a variety of mechanisms including inhibition of transcription initiation (Bird, A. P., Nature 321:209-213, 1986; Delgado et al., EMBO Journal 17:2426-2435, 1998), disruption of local chromatin structure (Counts & Goodman, Molecular Carcinogenesis 11:185-188, 1994; Antequera et al., Cell 62:503-514, 1990), and recruitment of proteins that interact specifically with methylated sequences and thereby directly or indirectly prevent transcription factor binding (Bird, A. P., Cell 70:5-8, 1992; Counts & Goodman, Molecular Carcinogenesis 11:185-188, 1994; Cedar, H., Cell 53:3-4, 1988). Many studies have demonstrated the effect of methylation of CpG islands on gene expression (e.g., the CDKN2A/p16 gene; Gonzalez-Zulueta et al., Cancer Research 55:4531-4535, 1995), but most CpG islands on autosomal genes remain unmethylated in the germline, and methylation of these islands is usually independent of gene expression. Tissue-specific genes are typically unmethylated in the respective target organs but are methylated in the germline and in non-expressing adult tissues, while CpG islands of constitutively expressed housekeeping genes are normally unmethylated in the germline and in somatic tissues.

[0007] Methylation within CpG islands affects the expression of genes involved in cancer. Data from a group of studies show the presence of altered methylation in cancer cells relative to non-cancerous cells. These studies show not only alteration of the overall genomic levels of DNA methylation, but also changes in the distribution of methyl groups. For example, abnormal methylation of CpG islands that are associated with tumor suppressor genes or oncogenes within a cell may cause altered gene expression. Such altered gene expression may provide a population of cells with a selective growth advantage and thereby result in selection of these cells to the detriment of the organism (i.e., cancer).

[0008] Insufficient correlative data. Unfortunately, the mere knowledge of the basic existence of altered methylation of CpG dinucleotides within CpG islands of cancer cells relative to normal cells, or of the fact that in particular instances such methylation changes result in altered gene expression (or chromatin structure or stability), is inadequate to allow for effective diagnostic, prognostic and therapeutic application of this knowledge. This is because only a limited number of CpG islands have been characterized, and thus there is insufficient knowledge, as to which particular CpG islands, among many, are actually involved in, or show significant correlation with cancer or the etiology thereof. Moreover, complex methylation patterns, involving a plurality of methylation-altered DNA sequences, including those that may have the sequence composition to qualify as CpG islands, may exist in particular cancers.

[0009] Therefore there is a need in the art to identify and characterize specific methylation altered DNA sequences, and to correlate them with cancer to allow for their diagnostic, prognostic and therapeutic application.

SUMMARY OF THE INVENTION

[0010] The present invention provides for a diagnostic or prognostic assay for cancer, comprising: obtaining a tissue sample from a test tissue; performing a methylation assay on DNA derived from the tissue sample, wherein the methylation assay determines the methylation state of a CpG dinucleotide within a DNA sequence of the DNA, and wherein the DNA sequence is a sequence selected from the group consisting of sequences of SEQ ID NOS:1-103, sequences having a nucleotide sequence at least 90% identical to sequences of SEQ ID NOS:1-103, CpG island sequences associated with sequences of SEQ ID NOS:1-103, CpG island sequences associated with sequences having a nucleotide sequence at least 90% identical to sequences of SEQ ID NOS:1-103, and combinations thereof, wherein the CpG island sequence associated with the sequence of the particular SEQ ID NO is that contiguous sequence of genomic DNA that encompasses at least one nucleotide of the particular SEQ ID NO sequence, and satisfies the criteria of having both a frequency of CpG dinucleotides corresponding to an Observed/Expected Ratio >0.6, and a GC Content >0.5; and determining a diagnosis or prognosis based, at least in part, upon the methylation state of the CpG dinucleotide within the DNA sequence. Preferably, the DNA sequence is a sequence selected from the group consisting of CpG island sequences associated with sequences of SEQ ID NOS:1-103, CpG island sequences associated with sequences having a nucleotide sequence at least 90% identical to sequences of SEQ ID NOS:1-103, and combinations thereof. Preferably, the DNA sequence is a sequence selected from the group consisting of CpG island sequences associated with sequences of SEQ ID NOS: 2, 4, 6, 7, 9-16, 19, 20, 22-33, 35-43, 48, 51-55, 59, 60, 64, 71, 76, 78-81, 84 and 87-90, and combinations thereof. Preferably, the methylation assay procedure is selected from the group consisting of MethyLight, MS-SnuPE (methylation-sensitive single nucleotide primer extension), MSP (methylation-specific PCR), MCA (methylated CpG island amplification), COBRA (combined bisulfite restriction analysis), and combinations thereof. Preferably, the methylation state of the CpG dinucleotide within the DNA sequence is that of hypermethylation, hypomethylation or normal methylation. Preferably, the cancer is selected from the group consisting of bladder cancer, prostate cancer, colon cancer, lung cancer, renal cancer, leukemia, breast cancer, uterine cancer, astrocytoma, glioblastoma, and neuroblastoma. Preferably, the cancer is bladder cancer, or prostate cancer.

[0011] The present invention further provides a kit useful for the detection of a methylated CpG-containing nucleic acid comprising a carrier means containing one or more containers comprising: a container containing a probe or primer which hybridizes to any region of a sequence selected from the group consisting of SEQ ID NOS:1-103, and sequences having a nucleotide sequence at least 90% identical to sequences of SEQ ID NOS:1-103; and additional standard methylation assay reagents required to affect detection of methylated CpG-containing nucleic acid based on the probe or primer. Preferably, the additional standard methylation assay reagents are standard reagents for performing a methylation assay from the group consisting of MethyLight, MS-SNuPE, MSP, MCA, COBRA, and combinations thereof. Preferably, the probe or primer comprises at least about 12 to 15 nucleotides of a sequence selected from the group consisting of SEQ ID NOS:1-103, and sequences having a nucleotide sequence at least 90% identical to sequences of SEQ ID NOS:1-103.

[0012] The present invention further provides an isolated nucleic acid molecule comprising a methylated or unmethylated polynucleotide sequence selected from the group consisting of SEQ ID NO:1, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:18, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:32, SEQ ID NO:34, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:44, SEQ ID NO:51, SEQ ID NO:52, SEQ ID NO:62, SEQ ID NO:64, SEQ ID NO:65, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:74, SEQ ID NO:76, SEQ ID NO:82, SEQ ID NO:83, SEQ ID NO:84, SEQ ID NO:86, SEQ ID NO:90, SEQ ID NO:92, SEQ ID NO:97, and SEQ ID NO:100. Preferably the nucleic acid is methylated. Preferably, the nucleic acid is unmethylated.

DETAILED DESCRIPTION OF THE INVENTION

Definitions

[0013] "GC Content" refers, within a particular DNA sequence, to the [(number of C bases+number of G bases)/band length for each fragment].

[0014] "Observed/Expected Ratio" ("0/E Ratio") refers to the frequency of CpG dinucleotides within a particular DNA sequence, and corresponds to the [number of CpG sites/(number of C bases.times.number of G bases)].times.band length for each fragment.

[0015] "CpG Island" refers to a contiguous region of genomic DNA that satisfies the criteria of (1) having a frequency of CpG dinucleotides corresponding to an "Observed/Expected Ratio">0.6), and (2) having a "GC Content">0.5. CpG islands are typically, but not always, between about 0.2 to about 1 kb in length. A CpG island sequence associated with a particular SEQ ID NO sequence of the present invention is that contiguous sequence of genomic DNA that encompasses at least one nucleotide of the particular SEQ ID NO sequence, and satisfies the criteria of having both a frequency of CpG dinucleotides corresponding to an Observed/Expected Ratio >0.6), and a GC Content >0.5.

[0016] "Methylation state" refers to the presence or absence of 5-methylcytosine ("5-mCyt") at one or a plurality of CpG dinucleotides within a DNA sequence.

[0017] "Hypermethylation" refers to the methylation state corresponding to an increased presence of 5-mCyt at one or a plurality of CpG dinucleotides within a DNA sequence of a test DNA sample, relative to the amount of 5-mCyt found at corresponding CpG dinucleotides within a normal control DNA sample.

[0018] "Hypomethylation" refers to the methylation state corresponding to a decreased presence of 5-mCyt at one or a plurality of CpG dinucleotides within a DNA sequence of a test DNA sample, relative to the amount of 5-mCyt found at corresponding CpG dinucleotides within a normal control DNA sample.

[0019] "Methylation assay" refers to any assay for determining the methylation state of a CpG dinucleotide within a sequence of DNA.

[0020] "MS.AP-PCR" (Methylation-Sensitive Arbitrarily-Primed Polymerase Chain Reaction) refers to the art-recognized technology that allows for a global scan of the genome using CG-rich primers to focus on the regions most likely to contain CpG dinucleotides, and described by Gonzalgo et al., Cancer Research 57:594-599, 1997.

[0021] "MethyLight" refers to the art-recognized fluorescence-based real-time PCR technique described by Eads et al., Cancer Res. 59:2302-2306, 1999.

[0022] "Ms-SNuPE" (Methylation-sensitive Single Nucleotide Primer Extension) refers to the art-recognized assay described by Gonzalgo & Jones, Nucleic Acids Res. 25:2529-2531, 1997.

[0023] "MSP" (Methylation-specific PCR) refers to the art-recognized methylation assay described by Herman et al. Proc. Natl. Acad. Sci. USA 93:9821-9826, 1996, and by U.S. Pat. No. 5,786,146.

[0024] "COBRA" (Combined Bisulfite Restriction Analysis) refers to the art-recognized methylation assay described by Xiong & Laird, Nucleic Acids Res. 25:2532-2534, 1997.

[0025] "MCA" (Methylated CpG Island Amplification) refers to the methylation assay described by Toyota et al., Cancer Res. 59:2307-12, 1999, and in WO 00/26401A1.

Overview

[0026] The present invention provides for 103 DNA sequences (i.e., "marker sequences") having distinct methylation patterns in cancer, as compared to normal tissue. These methylation-altered DNA sequence embodiments correspond to 103 DNA fragments isolated from bladder and prostate cancer patients, and in many instances, represent novel sequences not found in the GenBank database. None of the instant sequence embodiments have previously been characterized with respect to their methylation pattern in human cancers including, but not limited to, those of the bladder and prostate. The significance of such methylation patterns lies in the value of altered fragments as potential prognostic, diagnostic and therapeutic markers in the treatment of human cancers.

Identification of Methylation-Altered Marker Sequences in Genomic DNA

[0027] The MS.AP-PCR technique was used to scan the genomes of bladder or prostate cancer patients for DNA methylation changes relative to normal individuals, because the pattern is known to be highly conserved. A total of 103 DNA sequence embodiments (methylation-altered DNA sequences; "marker sequences") were isolated and characterized as having distinct methylation patterns in cancer, as compared to normal tissue.

[0028] Methods for the Identification of Marker Sequences in Genomic DNA. There are a variety of art-recognized genome scanning methods that have been used to identify altered methylation sites in cancer cells. For example, one method involves restriction landmark genomic scanning (Kawai et al., Mol. Cell. Biol. 14:7421-7427, 1994), another involves MCA (methylated CpG island amplification; Toyota et al., Cancer Res. 59:2307-12, 1999), and yet another involves MS.AP-PCR (Methylation-Sensitive Arbitrarily-Primed Polymerase Chain Reaction; Gonzalgo et al., Cancer Res. 57:594-599, 1997), which allows for a global scan of the genome using CG-rich primers to focus on the regions most likely to contain CpG dinucleotides. The MS.AP-PCR technique used in the present invention is a rapid and efficient method to screen ("scan") for altered methylation patterns in genomic DNA and to isolate specific sequences associated with these changes.

[0029] Briefly, genomic DNA from the tissue of bladder or prostate cancer patients was prepared using standard, art-recognized methods. Restriction enzymes (e.g., HpaII) with different sensitivities to cytosine methylation in their recognition sites were used to digest these genomic DNAs prior to arbitrarily primed PCR amplification with GC-rich primers. Fragments that showed differential methylation (e.g., hypermethylation or hypomethylation, based on the methylation sensitivity of the restriction enzyme, or upon DNA sequence analysis or Ms-SNuPE analysis; Gonzalgo & Jones, Nucleic Acids Res 25:2529-2531, 1997) were cloned and sequenced after resolving the PCR products on high-resolution polyacrylamide gels. The cloned fragments were used as probes for Southern blot analysis to confirm differential methylation of these regions in the tissue. Methods for DNA cloning, sequencing, PCR, high-resolution polyacrylamide gel resolution and Southern blot analysis are well known by those of ordinary skill in the relevant art.

[0030] Results. A total of 500 DNA fragments that underwent either hypermethylation (an increase in the level of methylation relative to normal) or hypomethylation (a decrease in the level of methylation relative to normal) were isolated from the scanned patients genomic DNA. A total of 178 of these fragments were sequenced, of which 103 were novel in that they corresponded to DNA loci whose methylation pattern had not previously been characterized. The corresponding sequences are disclosed as [SEQ ID NOS:1-103], wherein for certain sequences, the letter "n" refers to an undetermined nucleotide base.

[0031] Novel marker sequences identified by MS.AP-PCR. Table I shows an overall summary of methylation patterns and sequence data corresponding to the 103 DNA fragments identified by MS.AP-PCR. A total of 103 fragments were sequenced following identification as becoming either hypermethylated (gain of methylation; noted as having a hypermethylation pattern) or hypomethylated (loss of methylation; noted as having a hypomethylation pattern) relative to normal tissue. For the fragments of each category, the "Average GC Content" is shown, calculated as (number of C bases+number of G bases)/band length for each fragment, as well as the average Observed/Expected Ratio ("O/E Ratio"), calculated as [number of CpG sites/(number of C bases.times.number of G bases)].times.band length for each fragment. Additionally, the percent of fragments that qualify as CpG islands is listed, and corresponds to the percentage of all fragments within each category that have sequence compositions that satisfy the criteria of having a "GC Content">0.5 and an "0/E Ratio">0.6.

[0032] Thus, of these 103 fragments identified by MS.AP-PCR, 60 showed hypermethylation (Table I, upper row; Table II, [SEQ ID NOS:1-60]) while 43 showed hypomethylation (Table I, lower row; Table II, [SEQ ID NOS:61-103]). Moreover, 55 (43 hypermethylated, and 12 hypomethylated) of the 103 fragments correspond to CpG islands (i.e., fulfill the criteria of a GC content >0.5 and an Observed/Expected Ratio >0.6), whereas the other 48 (17 hypermethylated and 31 hypomethylated) fragments do not meet the criteria for CpG islands (see Table II).

TABLE-US-00001 TABLE I Summary of 103 DNA Fragments Identified by MS.AP-PCR Methylation Number of Percent that DNA Fragment Pattern (relative Fragments Average GC Average correspond to Type to normal) (103 total) Content O/E Ratio CpG Islands Hypermethylated Hyper- 60 0.54 0.72 72% Fragments methylation Hypomethylated Hypo-methylation 43 0.52 0.48 28% Fragments

[0033] Table II shows a summary of methylation pattern and sequence data for each individual sequence embodiment ([SEQ ID NOS:1-103]), corresponding to the 103 DNA fragments identified by MS.AP-PCR. Data for the 103 fragments was divided into either hypermethylated ([SEQ ID NOS:1-60]) or hypomethylated ([SEQ ID NOS:61-103]) categories. Table II also lists, for each sequence embodiment, the corresponding "Fragment Name," fragment "Size" (in base pairs; "bp"), "GC Content," Observed/Expected Ratio ("0/E Ratio"), "Description" (i.e., as a CpG island if criteria are met), "Inventor Initials" (IDCM=Isabel D. C. Markl, JC=Jonathan Cheng, GL=Gangning Liang, HF=Hualin Fu, YT=Yoshitaka Tomigahara), "Cancer Source," and "Chromosome Match" to the GenBank database. A dash ("-") indicates that no GenBank chromosome match existed, or that only a low-scoring partial match was found. Averages of the "GC Content" and "0/E Ratio," along with the percent of fragments that are CpG islands, are listed after the last member of both the hypermethylated and hypomethylated categories.

[0034] Therefore, the present invention provides for 103 DNA fragments and corresponding marker sequence embodiments (i.e., methylation-altered DNA sequences) that are useful in cancer prognostic, diagnostic and therapeutic applications.

[0035] Additionally, at least 55 of these 103 sequences correspond to CpG islands (based on GC Content and O/E ration); namely [SEQ ID NOS:2, 4, 6, 7, 9-16, 19, 20, 22-33, 35-43, 48, 51-55, 59, 60, 64, 71, 76, 78-81, 84 and 87-90]. Thus, based on the fact that the methylation state of a portion of a given CpG island is generally representative of the island as a whole, the present invention further encompassed the novel use of the 55 CpG islands associated with [SEQ ID NOS:2, 4, 6, 7, 9-16, 19, 20, 22-33, 35-43, 48, 51-55, 59, 60, 64, 71, 76, 78-81, 84 and 87-90] in cancer prognostic, diagnostic and therapeutic applications, where a CpG island sequence associated with the sequence of a particular SEQ ID NO is that contiguous sequence of genomic DNA that encompasses at least one nucleotide of the particular SEQ ID NO sequence, and satisfies the criteria of having both a frequency of CpG dinucleotides corresponding to an Observed/Expected Ratio >0.6, and a GC Content >0.5.

TABLE-US-00002 TABLE II Summary of MS.AP-PCR Fragments Sequenced Methylation Fragment Size GC O/E Inventor Cancer Chromosome [SEQ Pattern Name (bp) Content Ratio Description Initials Source Matches ID NO] Hyper- 11-1A 510 0.44 0.74 IDCM Bladder -- 1 methylation 14-3B 313 0.58 0.74 CpG Island IDCM Bladder 2 2 category 18-2B 165 0.57 0.45 IDCM Bladder 7 3 24-1B 601 0.51 0.72 CpG Island IDCM Bladder Xp11 4 26-1B 801 0.48 0.56 IDCM Bladder -- 5 26-2C 204 0.50 0.63 CpG Island IDCM Bladder -- 6 30-3D 205 0.55 1.25 CpG Island IDCM Bladder 14 7 32-3E 597 0.57 0.10 IDCM Bladder 20q12-13.1 8 34-2B 500 0.62 0.66 CpG Island IDCM Bladder 20 9 34-4B 343 0.70 0.81 CpG Island IDCM Bladder -- 10 34-5D 291 0.62 0.96 CpG Island IDCM Bladder 9 11 34-6A 266 0.64 0.93 CpG Island IDCM Bladder -- 12 35-1C 553 0.64 0.63 CpG Island IDCM Bladder -- 13 36-2D 156 0.60 0.58 CpG Island IDCM Bladder 10 14 38-1A 300 0.70 0.80 CpG Island IDCM Bladder 10 15 38-2B 196 0.56 0.89 CpG Island IDCM Bladder 15 16 7-8E 299 0.59 0.39 IDCM Bladder 17q21-22 17 83-4B 363 0.54 0.49 IDCM Bladder -- 18 84-1D 322 0.55 0.90 CpG Island IDCM Bladder 7 19 101-3E 255 0.57 0.83 CpG Island IDCM Bladder 17 20 M1-5A 406 0.45 0.96 IDCM Bladder 1 21 U2-8E 210 0.56 0.61 CpG Island IDCM Bladder 2 22 U12-1A 310 0.56 0.81 CpG Island IDCM Bladder 2 23 U7-4A 305 0.59 0.80 CpG Island IDCM Bladder -- 24 NU9-5A 379 0.67 0.83 CpG Island JC Bladder -- 25 3-17-8-B 625 0.48 0.72 CpG Island GL Bladder 18 26 4-10-4-A 499 0.55 0.30 CpG Island GL Bladder 7 27 1-1-1-A 561 0.58 0.98 CpG Island GL Bladder 20 28 3-17-8-A 717 0.50 0.68 CpG Island GL Bladder 17 29 G145-H 280 0.50 1.10 CpG Island GL Bladder 11 30 1-1-1-D 270 0.50 0.60 CpG Island GL Bladder 2 31 1-1-1-C 347 0.65 1.25 CpG Island GL Bladder -- 32 G178-A 342 0.55 0.85 CpG Island GL Bladder 2 33 34-A 370 0.62 0.44 HF Prostate -- 34 34-D 213 0.53 0.74 CpG Island HF Prostate 2 35 35-D 173 0.56 0.66 CpG Island HF Prostate 3 36 36-A 369 0.67 0.70 CpG Island HF Prostate -- 37 40-A 123 0.60 1.16 CpG Island HF Prostate -- 38 91-1 450 0.64 0.86 CpG Island YT Bladder 5 or 16q24.3 39 93-2 593 0.51 0.68 CpG Island YT Bladder Xp11 40 93-3 457 0.52 0.94 CpG Island YT Bladder Xp22.1-22.3 41 94-8 211 0.66 0.96 CpG Island YT Bladder -- 42 95-5 141 0.63 0.79 CpG Island YT Bladder 14 43 97-5 559 0.56 0.40 YT Bladder -- 44 98-1 433 0.46 0.96 YT Bladder 1 45 100-1 487 0.59 0.58 YT Bladder 14 46 100-2 403 0.60 0.47 YT Bladder 3 47 100-6 155 0.57 0.99 CpG Island YT Bladder 20 48 4-2 256 0.57 0.40 YT Bladder 7 49 5-8 224 0.47 0.96 YT Bladder 5 50 6-4 313 0.70 0.82 CpG Island YT Bladder -- 51 7-6 385 0.70 0.88 CpG Island YT Bladder -- 52 13-3 307 0.59 0.89 CpG Island YT Bladder 10 53 15-2 182 0.62 0.92 CpG Island YT Bladder 13 54 23-2 523 0.54 0.87 CpG Island YT Bladder Xp22.1-22.3 55 39-2 795 0.46 0.64 YT Bladder 13 56 40-2 438 0.62 0.51 YT Bladder 10 57 41-3 611 0.47 0.70 YT Bladder 18 58 105-4 291 0.58 0.71 CpG Island YT Bladder 5 59 107-8 226 0.53 0.96 CpG Island YT Bladder 11 60 AVERAGE 0.54 0.72 72% islands Hypo- 14-2B 580 0.55 0.51 IDCM Bladder 2 61 methylation 16-1B 633 0.56 0.39 IDCM Bladder -- 62 category 18-1B 703 0.45 0.35 IDCM Bladder 17 63 19-1B 420 0.66 0.87 CpG Island IDCM Bladder -- 64 20-1B 496 0.61 0.59 IDCM Bladder -- 65 21-2C 637 0.60 0.33 IDCM Bladder 9q34 66 29-1A 595 0.55 0.27 IDCM Bladder Xp11.23 67 29-2B 580 0.47 0.77 IDCM Bladder -- 68 32-1A 589 0.59 0.48 IDCM Bladder -- 69 34-1B 450 0.42 0.46 IDCM Bladder -- 70 34-3B 432 0.70 0.61 CpG Island IDCM Bladder -- 71 32-2B 748 0.47 0.24 IDCM Bladder 2 72 32-4B 599 0.57 0.15 IDCM Bladder 20q12-13.1 73 32-5B 614 0.58 0.20 IDCM Bladder -- 74 33-1A 552 0.54 0.32 IDCM Bladder 10 75 5-1E 501 0.61 1.04 CpG Island IDCM Bladder -- 76 6-1A 826 0.55 0.36 IDCM Bladder 22q13.32-13.33 77 7-5D 433 0.59 0.85 CpG Island IDCM Bladder 5 78 8-7C 424 0.58 0.83 CpG Island IDCM Bladder 5 79 30-6D 285 0.63 0.72 CpG Island IDCM Bladder 1 80 66-2E 401 0.54 0.82 CpG Island IDCM Bladder 16 81 78-1C 268 0.54 0.41 IDCM Bladder -- 82 97-2E 989 0.53 0.16 IDCM Bladder -- 83 M1-8C 250 0.64 0.99 CpG Island IDCM Bladder -- 84 M2-5A 402 0.50 0.45 IDCM Bladder 5 85 M1-4P 595 0.43 0.41 IDCM Bladder -- 86 M12-10A 304 0.53 0.76 CpG Island IDCM Bladder 7 87 M12-12C 296 0.51 0.64 CpG Island IDCM Bladder 17 88 M2-8M 220 0.67 0.62 CpG Island IDCM Bladder 6q27 89 NU4-3A 273 0.63 1.02 CpG Island JC Bladder -- 90 NU5-2A 361 0.44 0.73 JC Bladder 6q14.3-15 91 88-5 462 0.62 0.39 YT Bladder -- 92 90-1 591 0.66 0.45 YT Bladder 19 93 91-3 279 0.58 0.45 YT Bladder 5 or 16q24.3 94 91-4 351 0.55 0.30 YT Bladder 18q23 95 91-7 171 0.61 0.59 YT Bladder 11 96 89-3 743 0.55 0.43 YT Bladder -- 97 94-2 589 0.53 0.41 YT Bladder 22q13.31-13.32 98 94-3 538 0.53 0.49 YT Bladder 5 or 18 99 94-4 486 0.61 0.57 YT Bladder -- 100 94-5 450 0.60 0.45 YT Bladder 1p36.2-36.3 101 94-6 292 0.58 0.32 YT Bladder 8 or 9 102 96-4 395 0.63 0.54 YT Bladder 9 103 AVERAGE 0.52 0.48 28% islands

[0036] Diagnostic and Prognostic Assays for Cancer. The present invention provides for diagnostic and prognostic cancer assays based on determination of the methylation state of one or more of the disclosed 103 methylation-altered DNA sequence embodiments. Typically, such assays involve obtaining a tissue sample from a test tissue, performing a methylation assay on DNA derived from the tissue sample, and making a diagnosis or prognosis based thereon.

[0037] The methylation assay is used to determine the methylation state of one or a plurality of CpG dinucleotide within a DNA sequence of the DNA sample. According to the present invention, possible methylation states include hypermethylation and hypomethylation, relative to a normal state (i.e., non-cancerous control state). Hypermethylation and hypomethylation refer to the methylation states corresponding to an increased or decreased, respectively, presence 5-methylcytosine ("5-mCyt") at one or a plurality of CpG dinucleotides within a DNA sequence of the test sample, relative to the amount of 5-mCyt found at corresponding CpG dinucleotides within a normal control DNA sample.

[0038] A diagnosis or prognosis is based, at least in part, upon the determined methylation state of the sample DNA sequence compared to control data obtained from normal, non-cancerous tissue.

[0039] Methylation Assay Procedures. Various methylation assay procedures are known in the art, and can be used in conjunction with the present invention. These assays allow for determination of the methylation state of one or a plurality of CpG dinucleotides (e.g., CpG islands) within a DNA sequence. Such assays involve, among other techniques, DNA sequencing of bisulfite-treated DNA, PCR (for sequence-specific amplification), Southern blot analysis, use of methylation-sensitive restriction enzymes, etc.

[0040] For example, genomic sequencing has been simplified for analysis of DNA methylation patterns and 5-methylcytosine distribution by using bisulfite treatment (Frommer et al., Proc. Natl. Acad. Sci. USA 89:1827-1831, 1992). Additionally, restriction enzyme digestion of PCR products amplified from bisulfite-converted DNA is used, e.g., the method described by Sadri & Hornsby (Nucl. Acids Res. 24:5058-5059, 1996), or COBRA (Combined Bisulfite Restriction Analysis) (Xiong & Laird, Nucleic Acids Res. 25:2532-2534, 1997).

[0041] COBRA. COBRA analysis is a quantitative methylation assay useful for determining DNA methylation levels at specific gene loci in small amounts of genomic DNA (Xiong & Laird, Nucleic Acids Res. 25:2532-2534, 1997). Briefly, restriction enzyme digestion is used to reveal methylation-dependent sequence differences in PCR products of sodium bisulfite-treated DNA. Methylation-dependent sequence differences are first introduced into the genomic DNA by standard bisulfite treatment according to the procedure described by Frommer et al. (Proc. Natl. Acad. Sci. USA 89:1827-1831, 1992). PCR amplification of the bisulfite converted DNA is then performed using primers specific for the interested CpG islands, followed by restriction endonuclease digestion, gel electrophoresis, and detection using specific, labeled hybridization probes. Methylation levels in the original DNA sample are represented by the relative amounts of digested and undigested PCR product in a linearly quantitative fashion across a wide spectrum of DNA methylation levels. In addition, this technique can be reliably applied to DNA obtained from microdissected paraffin-embedded tissue samples. Typical reagents (e.g., as might be found in a typical COBRA-based kit) for COBRA analysis may include, but are not limited to: PCR primers for specific gene (or methylation-altered DNA sequence or CpG island); restriction enzyme and appropriate buffer; gene-hybridization oligo; control hybridization oligo; kinase labeling kit for oligo probe; and radioactive nucleotides. Additionally, bisulfite conversion reagents may include: DNA denaturation buffer; sulfonation buffer; DNA recovery regents or kit (e.g., precipitation, ultrafiltration, affinity column); desulfonation buffer; and DNA recovery components.

[0042] Preferably, assays such as "MethyLight" (a fluorescence-based real-time PCR technique) (Eads et al., Cancer Res. 59:2302-2306, 1999), Ms-SNuPE (Methylation-sensitive Single Nucleotide Primer Extension) reactions (Gonzalgo & Jones, Nucleic Acids Res. 25:2529-2531, 1997), methylation-specific PCR ("MSP"; Herman et al., Proc. Natl. Acad. Sci. USA 93:9821-9826, 1996; U.S. Pat. No. 5,786,146), and methylated CpG island amplification ("MCA"; Toyota et al., Cancer Res. 59:2307-12, 1999) are used alone or in combination with other of these methods.

[0043] MethyLight. The MethyLight assay is a high-throughput quantitative methylation assay that utilizes fluorescence-based real-time PCR (TaqMan.RTM.) technology that requires no further manipulations after the PCR step (Eads et al., Cancer Res. 59:2302-2306, 1999). Briefly, the MethyLight process begins with a mixed sample of genomic DNA that is converted, in a sodium bisulfite reaction, to a mixed pool of methylation-dependent sequence differences according to standard procedures (the bisulfite process converts unmethylated cytosine residues to uracil). Fluorescence-based PCR is then performed either in an "unbiased" (with primers that do not overlap known CpG methylation sites) PCR reaction, or in a "biased" (with PCR primers that overlap known CpG dinucleotides) reaction. Sequence discrimination can occur either at the level of the amplification process or at the level of the fluorescence detection process, or both.

[0044] The MethyLight may assay be used as a quantitative test for methylation patterns in the genomic DNA sample, wherein sequence discrimination occurs at the level of probe hybridization. In this quantitative version, the PCR reaction provides for unbiased amplification in the presence of a fluorescent probe that overlaps a particular putative methylation site. An unbiased control for the amount of input DNA is provided by a reaction in which neither the primers, nor the probe overlie any CpG dinucleotides. Alternatively, a qualitative test for genomic methylation is achieved by probing of the biased PCR pool with either control oligonucleotides that do not "cover" known methylation sites (a fluorescence-based version of the "MSP" technique), or with oligonucleotides covering potential methylation sites.

[0045] The MethyLight process can by used with a "TaqMan.RTM." probe in the amplification process. For example, double-stranded genomic DNA is treated with sodium bisulfite and subjected to one of two sets of PCR reactions using TaqMan.RTM. probes; e.g., with either biased primers and TaqMan.RTM. probe, or unbiased primers and TaqMan.RTM. probe. The TaqMan.RTM. probe is dual-labeled with fluorescent "reporter" and "quencher" molecules, and is designed to be specific for a relatively high GC content region so that it melts out at about 10.degree. C. higher temperature in the PCR cycle than the forward or reverse primers. This allows the TaqMan.RTM. probe to remain fully hybridized during the PCR annealing/extension step. As the Taq polymerase enzymatically synthesizes a new strand during PCR, it will eventually reach the annealed TaqMan.RTM. probe. The Taq polymerase 5' to 3' endonuclease activity will then displace the TaqMan.RTM. probe by digesting it to release the fluorescent reporter molecule for quantitative detection of its now unquenched signal using a real-time fluorescent detection system.

[0046] Typical reagents (e.g., as might be found in a typical MethyLight-based kit) for MethyLight analysis may include, but are not limited to: PCR primers for specific gene (or methylation-altered DNA sequence or CpG island); TaqMan.RTM. probes; optimized PCR buffers and deoxynucleotides; and Taq polymerase.

[0047] Ms-SNuPE. The Ms-SNuPE technique is a quantitative method for assessing methylation differences at specific CpG sites based on bisulfite treatment of DNA, followed by single-nucleotide primer extension (Gonzalgo & Jones, Nucleic Acids Res. 25:2529-2531, 1997). Briefly, genomic DNA is reacted with sodium bisulfite to convert unmethylated cytosine to uracil while leaving 5-methylcytosine unchanged. Amplification of the desired target sequence is then performed using PCR primers specific for bisulfite-converted DNA, and the resulting product is isolated and used as a template for methylation analysis at the CpG site(s) of interest. Small amounts of DNA can be analyzed (e.g., microdissected pathology sections), and it avoids utilization of restriction enzymes for determining the methylation status at CpG sites. Typical reagents (e.g., as might be found in a typical Ms-SNuPE-based kit) for Ms-SNuPE analysis may include, but are not limited to: PCR primers for specific gene (or methylation-altered DNA sequence or CpG island); optimized PCR buffers and deoxynucleotides; gel extraction kit; positive control primers; Ms-SNuPE primers for specific gene; reaction buffer (for the Ms-SNuPE reaction); and radioactive nucleotides. Additionally, bisulfite conversion reagents may include: DNA denaturation buffer; sulfonation buffer; DNA recovery regents or kit (e.g., precipitation, ultrafiltration, affinity column); desulfonation buffer; and DNA recovery components.

[0048] MSP. MSP (methylation-specific PCR) allows for assessing the methylation status of virtually any group of CpG sites within a CpG island, independent of the use of methylation-sensitive restriction enzymes (Herman et al. Proc. Natl. Acad. Sci. USA 93:9821-9826, 1996; U.S. Pat. No. 5,786,146). Briefly, DNA is modified by sodium bisulfite converting all unmethylated, but not methylated cytosines to uracil, and subsequently amplified with primers specific for methylated versus unmethylated DNA. MSP requires only small quantities of DNA, is sensitive to 0.1% methylated alleles of a given CpG island locus, and can be performed on DNA extracted from paraffin-embedded samples. Typical reagents (e.g., as might be found in a typical MSP-based kit) for MSP analysis may include, but are not limited to: methylated and unmethylated PCR primers for specific gene (or methylation-altered DNA sequence or CpG island), optimized PCR buffers and deoxynucleotides, and specific probes.

[0049] MCA. The MCA technique is a method that can be used to screen for altered methylation patterns in genomic DNA, and to isolate specific sequences associated with these changes (Toyota et al., Cancer Res. 59:2307-12, 1999). Briefly, restriction enzymes with different sensitivities to cytosine methylation in their recognition sites are used to digest genomic DNAs from primary tumors, cell lines, and normal tissues prior to arbitrarily primed PCR amplification. Fragments that show differential methylation are cloned and sequenced after resolving the PCR products on high-resolution polyacrylamide gels. The cloned fragments are then used as probes for Southern analysis to confirm differential methylation of these regions. Typical reagents (e.g., as might be found in a typical MCA-based kit) for MCA analysis may include, but are not limited to: PCR primers for arbitrary priming Genomic DNA; PCR buffers and nucleotides, restriction enzymes and appropriate buffers; gene-hybridization oligos or probes; control hybridization oligos or probes.

[0050] Kits for Detection of Methylated CpG-containing Nucleic Acid. The reagents required to perform one or more art-recognized methylation assays (including those identified above) are combined with primers or probes comprising the sequences of SEQ ID NOS:1-103, or portions thereof, to determine the methylation state of CpG-containing nucleic acids.

For example, the MethyLight, Ms-SNuPE, MCA, COBRA, and MSP methylation assays could be used alone or in combination, along with primers or probes comprising the sequences of SEQ ID NOS:1-103, or portions thereof, to determine the methylation state of a CpG dinucleotide within a genomic sequence corresponding to SEQ ID NOS:1-103, or to CpG island sequences associated with sequences of SEQ ID NOS:1-103, where the CpG island sequence associated with the sequence of the particular SEQ ID NO is that contiguous sequence of genomic DNA that encompasses at least one nucleotide of the particular SEQ ID NO sequence, and satisfies the criteria of having both a frequency of CpG dinucleotides corresponding to an Observed/Expected Ratio >0.6, and a GC Content >0.5.

Sequence CWU 1

1

1031510DNAHomo sapiensmisc_feature(15)..(15)"n" refers to an undetermined base 1ttgcaagccc ccttngctct tcctttgncc tcgcctacat attcagggga tcgcaatctc 60actcgcgaaa taatttnttt ctgtaagagg aagccgcctt tcccctctcc caccgccaag 120gtaaaggctg ctaaagtagc tcttcttgga aggaaaaata ttttaaaaag cagctgggtt 180gctctccaca agaagatggc agttttggga aaacccatta tgtgtccaaa tgccggtttc 240cttttcttgt ttaacgcttt ttagagggca aaaatgacgc tcatgtgaag cccacaggct 300cgagccaatg tcgctgggct aattatgagt ctgcttatcc cactcccaaa tatccgagac 360gactcactca naagacattt ttactcttcc aagaattgng aattcagaan cagcttcccc 420acattctaag agaaaaaaaa acttgtttaa cgggcacgtt tttgattttt ttgccgctgg 480cgaccttaat taaaagccgg gagctncnna 5102313DNAHomo sapiens 2gcactcttaa aacgcctctc tgcagtccca ggtccgcgct ccccaagaac tggccagatc 60gcgccgggct tggcccctga caactctgcc tcctccacct gttgcgttta ctccgtttag 120ttggctgtgc agtctctggc cccaggtgtg cttttaaaac tcgaggaacg cgggtgttgg 180actcattcgc agcctcttgc ctctggttcc cgtgatccca cggtggcgag cttccaggct 240cagcgaggag atctgggttt gaacattcat ctcccatgtt actcttttct tgctcctcgc 300gtccccaagc cga 3133165DNAHomo sapiens 3gcttagcaaa tttccctttt ttattgttgg ttttgtctgt tggctcttac cccctttcct 60tttcctgctt cccctgagtc agcaatgctg agcccagcga agcacagggg gccaaaggga 120gagacacacg gagcgccccg gggtccccca gcctcggcgg ccaaa 1654601DNAHomo sapiens 4ggggggagtc gtgcgtgtca gatttaggcc aggaagcgga agtcgccagc agcgagagtt 60taacctctgt gggcgcagag ggttgcgggg attcagcgcc cgggaccgtg gatctgtgca 120gggagtcata ggtgtgtgtg acatcagtgg tggaacattt tggctcgttt tcacaattca 180gtcattatcc tttctgcttt cctcctggaa gcattaaggt tgaagttttc ttctaaagat 240caaagttttg atttgttata ttagttcgga tttgtttgat ttttgtttgt gttcggtttc 300aagtgctgat ttgtaacttt tctccccccc cacacacacg ccttttgacc cctgaattat 360ttaaaagtcc attgttggag tggcaaacat cctccgagac tcaaagggca aggccatggg 420cgctttattc cggctgctgc tccaggaacg tgggaaagca gcggagtttt attctagggg 480aaggaaacaa aggcggccga gtgccagctg cacgtttggt gggatttggt catcaggggt 540ggacatgctg cccaatggag ctgtcggcag tttgacccag cttggtccgt cgcgtcccga 600a 6015801DNAHomo sapiensmisc_feature(467)..(467)"n" refers to an undetermined base 5gagtacgcgg ggcagaacca gcgcaataca gcattctggt aggggaacta attttgacta 60aaatatttgc caattctaat ccccaattcc tggacctccg ggtagctggc aaggtatttt 120atgttagatg tgtctggagt aaggtgcacc ggagtatttc gacaagagac tcaattcaat 180gcgtattaaa acttgattga gagagggaga gagagaggtc attttataaa gaaagacctg 240tgaacactgt agattggaaa tttatgtttg caaaataaaa ggatgggttt atcaagtgga 300tgcatttaca aaatgtggca tccaggtttc gtaaaattag ctgaattcta cgggtaagat 360tatgaatgtg gctcataaat aattaatagg tagtgaaaaa gaatgtattt tgcattaggc 420agtgcattca atagtatttc ggaaatgagc acttcgattt cctcggnttc catgcgnggc 480cacctctcca gagcagggca ggcacccagg gnggtgccca cacaaacaag cgcgtgtggg 540cattttcttg gctcgtgcgc tgaagtgcac gctgggcctt ggtgcccgca ccctcagcct 600gggagatagg gaggtggtgc tacctgcagg ccgattgtgt ccccgccata ggacactagt 660gggcggcaaa cctcacaaga ctcttgcagc cagccttcag cagagccagc aaacccagcc 720gccaccgagg gaggactgct ccatgcagat ggtcaggggc tttcttctga agacgcctcc 780cccacgatct ctcaagttca c 8016204DNAHomo sapiens 6ccggttgctg ctggaggatg ggactacgaa ggatggggac tccgctcggc caccgctcct 60gaatggcctc taatctcggt gttaaatact ttatgagagt atcaatacca cctaatcctt 120tgctgagaat tactgctaga aatgtagatt ctgaggttcc gaaagtttgt ttttggttac 180cccctccagc tcctcccgcg gcaa 2047205DNAHomo sapiens 7agacctttat cgggcgtgag tgaaatcggt cgttcgtgtt tttcgtgggt cttccaacca 60caggccgcct gagatggttc tagtcccttt gagtatacag acccttcctg tgcattgacc 120gacacagctc ggcccggatc ccgaaatgaa cgtttctacc ttcggaacgc tgcgtctcgg 180atccttctga acccgcacgt cgcaa 2058597DNAHomo sapiensmisc_feature(361)..(361)"n" refers to an undetermined base 8gaccatgaaa tcgtgtggct ctagcccctt ctgggcctct tgttggtaat gaagccactc 60taaagcgccc cctgttattc agagggctcc ccagctgcca tgatatgtgt atggggaggg 120catagcaggt ccttttgccc cggcagccat tcttctgctc acaaggggct ggctctgggg 180acagggatgt ctttgtcatc agtgaccact aatccccctc ctcattggcc tccagggctg 240ctccccttca ctctcttggt tgaagttgta ggggctgagg ttaccctgag aaacacctgt 300tcttggagcc catagaccca accttggaga tgcaggggga gccactggct gggctctgca 360ngtggggcca gctgatcccc anctgctggc acctccaggc atccacagag cttggagtcc 420cagccacatt tcctccttgg ccttagaggg agaggaagtc ctttgattgc ctagtccaag 480atccctttat ttcctgccct gggattatgg ggnagcaagc catgcccttc atgggaagct 540gttctccctt cctcggggtt gggtctggcc tcagctcggg caacagtcat gatgggc 5979500DNAHomo sapiensmisc_feature(10)..(10)"n" refers to an undetermined base 9gccaaacgcn ataccctctg cggggtgaga atgcgggccc gcccggctcc tcccgtgagg 60ccagggcctc ctgttctcct agacacccca aggagccaac tcctccgcag aagttccccg 120cttctgctct tatttccaag cttcgcgctt tctacaaact ccctgttgcc ttgactttga 180tttccagccg tggtgagggt cagagtgaac cccggcgcgc tccccgacgg catccccgca 240caccaggata ggagaaattg gagggcctgg ggcctcgggc tccgcagtcg tcggaggaag 300aacccaccgc ggggtccgca agggaaagtg aagaggcccg ggatttttcc aaagcgctgg 360ccaggacccc gaaggaaggg gaggagtcac ctgaagccgg ggaaggcccc ttgggtgctc 420tgccttggat ccttatgttc actgactttc gcgacccctg gaggggggca aatccgcgct 480gtttccccca acttggcttc 50010343DNAHomo sapiens 10gccaacccac accagtacct gggaccgggg ggagcccggt ccggccgcta aaccgggctg 60gctggcgcca gggctccggg aggtgcggtc cggcggggaa gccgtgatgg gaagcgactc 120tgtccaggga gtgtccttca ccaccacact cctcacgtcc aggcagtgat cgacggcctg 180gcggcaccct cacagcgggc ccatagcacg gggccacaca cgtcccctga gcttagcctg 240ggcacattcg tctgccgccg agggcttaag ccagtctgca gcccgcgccc cgtcactcgg 300acgcaagtcc gtcgtccgct ctgccacgcg gccgctaagc cga 34311291DNAHomo sapiens 11gtcctacaca ctccgcacac aacgcggccg gtgttaagtc tccaaacgcc ccgagagctc 60caaggaccgc gcgcgcgaag gcgccgtagc aagtgggcac acaccagaca ccaccccggc 120gtgttccgcg ggagaagcca gtgcacacat cctcccgcaa ggcggggttg ccagtgcaac 180acaggaatcc tgcccttttt ctagaaaagc cccctccccc actttccctc caatacactc 240acctgcgtct caacagtttc cttcttgcgc tacacgcggc cgctaagccg a 29112266DNAHomo sapiens 12gtccggatca gtttccccgg ccaggtcgct tcccggtctc aaccatttcg cgctctgctc 60tgtccgctgg tttgtccctg cccggttcct ctccccgggc ctgtcagcct ccgcttctct 120ggaggttcct gggactcatc tctgatccac cgtcttgcgt tctctgggcg catcgacttc 180tctccatctt cgggctcact cctgactccc tcgctgccgc ccccgggggt ttccacgcgt 240gtctctaacc gcggccgcta agccga 26613553DNAHomo sapiensmisc_feature(497)..(497)"n" refers to an undetermined base 13gatcctggtc catcgaaacc ttgtgtgcat cggttagtgc ttcctgggcg tttgcttcta 60gccgacgctg acagtggagt gccagaaaga gggagaggac cgtcatggct actctgcccc 120tggtgtcacc atgcgctctc ccccggcacc ggcgaggcga aacgtttcgc tagtccccgg 180gaggcccctc ggtcagggca gcagcatccc tgcaccctct ccgcaggtgg tctccccgac 240gccacaggtg gccagcaggg cgcgggtggg ggcaggagcg cctctcccct gcccaggcct 300cccgctcctt ctcggagcgc tgtggcgggg tggagagaca gccttctaca gctagtctag 360ctcggcgcgg ttcccgtctg tggcctccta atcccacagc cacagcgcct tcctctaacc 420tccctcggtg ggcttaaagc ctcccgttcc ttctgtctca ttccttctgc tccctccccc 480cgaaaccccc agatganagc tgggaacctg gcnccantna ctgagcnaac agtgttgacg 540ggccgnggcc caa 55314156DNAHomo sapiens 14gcgcacacag tgggtacaag gatgagctcg gtgtaaggaa tggaaagccc ccagtctaaa 60ccaccgcccc ctagacacgg gtgaaaacct gcctaaaagc taactcaggc agtgactcta 120tcacccgaag gggccctggg ccgcggccca agccga 15615300DNAHomo sapiensmisc_feature(117)..(117)"n" refers to an undetermined base 15gttcacagcc cataaggtgg gggtggcccg aacctgaaac ggagcctgag ccaggatcct 60gcaaccaaag tctgaagcgc cccccggtgg gggccgagag cgctgcaggc aggtggnggc 120gcggggcagg cgggcgggcg aagggagctc cggntacgca ganaacgcgg agcgccccct 180tcccacctgc gcgagggcat cctgcccggg ggaggaaagg cgggagtccg aggcgggtcg 240gattcccagc cagctccctc ctcacaggag gcggcccatt atccggcgtc gcaaagccga 30016196DNAHomo sapiens 16ggcgcccagc aggggagcga gggaggaggg tgcagaaaga ggctccgaaa ttgggggaaa 60ctgacccgtg cttctctacc ttcggaggtg ggacagttgc acgaagtgct agttagaccg 120gatcagttgg aactgacgga ggactgcaaa gaagaaacta aaatagacgt cgaaagcctg 180tcctcggcgt cgcaaa 19617299DNAHomo sapiensmisc_feature(21)..(21)"n" refers to an undetermined base 17acaccaggag aggggaagaa nccagcacct accgacaggg gtggagctgg gtcaagaatg 60gtgtggtccc tgctttgggg gaatgctggg gaggtagaaa gccccttcta acggggcgtc 120actgcaatta ctgcttcctc tttcccataa aactccccct agtgtatcag aacccccaag 180gagtttcagt aagcggttct tctgttgtct ccggctgaga ctccagggga acctcaagct 240cacatggccc tggccgggcc cctgggcagg agcaggcgag aggtctgcgc ggccgctaa 29918363DNAHomo sapiens 18gggtatgtgt tacacatccg agataactac acaggcatcg accctgtcca cccggggatg 60ctagaggggc tgcgctggtt ttactccagg ccatggtgag agccaccgtg aacacagggc 120tctctcctct gagctgcaga agctctgtgc cctgtcccct gccacaagtc acagactttc 180ttcatgtgtt ttacctcatg ttaatgaagg agatcttctc caggggcttg atctagtggg 240aaacagagga gggggggatt ttaaatttca gtccgtccaa ccctgtagat ctgctgtcct 300acagtaacgt aaaggatcac caggtaaaac gctgcttctc ccggacgccg ccccgcaagc 360cga 36319322DNAHomo sapiens 19ccggcccgtc cctcttaata tggcctcagt tccgaaaacc acagaataga accgcggtcc 60tattccatta ttcctagctg aggtatccag gcggctcgga cctgctttga acactctaat 120tttttcaaag taaacgcttc gggctgcagg acactcagct aagagcatca ggggggcgcc 180aagaggcaag gggcggggat gggtggtggc tcgcctcgtg gcagaccgcc cgcccgctcc 240caagatccaa ctacgagctt tttaactgca gcaactttaa tatacgctat tggagctgga 300attaccgcgg ccgctaagcc ga 32220255DNAHomo sapiens 20taataagata ccaaatcggg cgagaaacga aaagctcctg gcctccgtat ttggggccag 60agacaccgca gggagtcagg tccccgccga caaatcggaa gaggcctgcg ggagttagcc 120agataatgct ctccctgtcc tacccgtccc caccaatttg ccttttacct gccgcagagc 180ttgcttgaac caaaggggtt tgcggtcttc tcctcctcaa cttgcgatcc ccaggccttc 240gcgtcccgaa gccga 25521406DNAHomo sapiensmisc_feature(6)..(6)"n" refers to an undetermined base 21atgtgnnaag gctcgctntc catttctctt ttcctccttc tccctctctc atgtgcggtc 60tccctcaaca tccaaaccaa ccgagtgcgt ctgaggtgaa atcgtgccag acttagagac 120ggctgccagg tttctctcaa gtcttggctt aacaaaagaa agcaaattac aaaaatggaa 180attttcaaac tagcgttcag tggtattcaa atcgacgttt gggtagcgca caggcacaga 240ccgcattcgt gctattttgt gattaaaatg ataccaaaaa tacctccttg ctttggtttt 300cgtcttcgaa aacgacttct ttccttcttc taatttcccc cttacttttg ggagcggcaa 360acccctgacc actctagaat tgctaacatt tggaccggcg tcgcaa 40622210DNAHomo sapiensmisc_feature"n" refers to an undetermined base 22gcacgttcgn gcnncgtgta ccatnagctg ccaactggan gcaccnnggn aagggtgggg 60gcctcctgga gacttngggg agagggatag ccggntaaag ctcctgtcct ttctataggc 120ataagcgggt ggtcaccacg gattggggat cccgaatccc tggctccaga tagacttaat 180gaagaagcac ctggatccgg gccgcgncaa 21023310DNAHomo sapiensmisc_feature(9)..(9)"n" refers to an undetermined base 23tcacgcttnc naaggctctg aatcctgagg gncagatctc caagaaggag ggaggctggt 60cctagttccc gaggtcctnn actaggtcta gatcactggg taaaagaagg ggagcggcan 120cacgtatggg gtaggcgctc tcactactca catctcgaga cctttgccgg cgtagggctg 180tccgggggga acgacccgcc ttttccggta tcggttgtca tggcggcgcc cagcccagcc 240tggttttttc cggtagccaa ttgaactaac aaccccgttc cctttaggac taatctgtca 300cgtcggcgca 31024305DNAHomo sapiensmisc_feature(13)..(13)"n" refers to an undetermined base 24ctctggtctg tgntggatac gcgtgttctt ctgcggagtt aaagggtcgg ggacgggggt 60tctggactta ccanagcaat tccagccggt gggcgtttgg cagtcactta aggaggtagg 120gaaagcagcg agcttcaccg ggcgggctac gatgagtagc atgacgggca gcagcagcag 180ccagcaaaag ccctcgcaaa gtgtccagct gctgcactgc cgcggggact cccacagcac 240catgactagt tcgtgcgact ctgcancanc aaacggcttc cgaggaacac angatcgcgg 300gggca 30525379DNAHomo sapiensmisc_feature(6)..(6)"n" refers to an undetermined base 25aaaacncatn tgnagagcnc ntcggcagag ncgcagctgg ctgacccagg agaaggcgcg 60ctgggtgtgg ctgggacggc caaggccgcg gcttcccgcg tggggatgcg ctntggcgca 120aagctggtcc cggcggggcc aggcgtttgt gggcgggtga cggggatcta gggcttccgc 180tcgngattcc tcttgggctg tctttncggg tttggactcg cctgccaggc tgtgtgcagg 240gttcccgctg cctctggccg gcaggcgtcc gggctgcagg tgggccggca ggcaggtgtt 300agcgggaagg gagcacaggt agcgaggtgg gatcggcgac ctggctaggg tgtcggcaga 360atggaatgcg cggccgcta 37926625DNAHomo sapiensmisc_feature(8)..(8)"n" refers to an undetermined base 26gggacgcnag ccagggantt tgatccgttt tgaatgaaaa gaaagagaan ccaaaccaaa 60cctntcagtc atccaaaacc ttcaggcttc cagggaggtt ttgctataat tttctctaag 120catgactgtt tctgggggag gggaaagggg tggttgtatt tactgaaaat tcaaatcgaa 180ataataaatg gccaaatttg gacacttacg gacccaaaca gttttgctca cgccagagaa 240accgagagca cagggcttgc gtgaagccta tctcggcaga aggcaacatt ctaataaagc 300ccgtgggaaa acagattaca ttttcgccat gaataagtca tgcagtgaaa aatattgcct 360acagcctgtc gacttatatt attatcacgt ttttcaactc ggcgtgagga gggagaggag 420tgttcatatt tgactaggaa ttgcaggatc gatgcaaact ccagggcagc agccagactg 480gcatatgtgg ggctctccgg ttactttctc tgtatgtcgc gggtgagagg aacagcgagg 540acaatttagc gcaaacacac gaagggtcgg atctcaaggg ggcagcgctg ggagaaaggt 600tagggctgna gagcgnanag ncaaa 62527499DNAHomo sapiensmisc_feature(2)..(2)"n" refers to an undetermined base 27gnctccncgt tcccctcggg cggaacggag gcaactttcc ggagtctatt tttgttaaga 60caatcaactc caataactga gctgaagttt ttgtttaaaa agaaaaaaat ctgataagtg 120atgattttac ctacttgtgg acactagatt tcaattagga aggttttttt aaacggcttt 180ttgtaacttc gctgcaggaa gcaggtttgt ttctttttct tttcttttta agagaaggtg 240tatttcactg gtgcaatggc ttggcacctc cggggcctgg gaggacctca gacctcccca 300gccctgggtt tctccgtctt caagaccaac taggaagggt caagcgggga gagggagtgg 360agggtcaggt gagatctcag agctgccccg gccggccccc gtctctttct acctcctctt 420ccagagaacc agcggctcac acccttctca acgcaggaca tgctcggcgg ccaaagccga 480attctgcaga tatccatca 49928561DNAHomo sapiensmisc_feature(20)..(20)"n" refers to an undetermined base 28gggcgattgt tattcaaacn ngntanctct ctgcggggnn gagnaatgng ggcctcgcac 60ggctncatcc ccgtcgagcn cagggcctcc ctgttctnct agacatncca aggagccaac 120tcctccgcag aagttccccg cttctgctct tatttccaag cttcgcgctt tctacaaact 180ccctgttgcc ttgactttga tttccagccg tggtgagggt cagagtgaac cccggcgcgc 240tccccgacgg catccccgca caccaggata ggagaaattg gagggcctgg gcctcggctc 300ccgcagtcgt cggaggaaga acccaccgcg gggtccccaa gggaaagtga agaggcccgg 360gatttttcca aagcgctgcc aggaccccga aggaagggga ggagtcacct gaagccgggg 420aagctccttg ggtgctctcc ttggatcctt atgttcactg actttcgcga ngccccctgg 480aggnggaaaa tccgcgctgt ttcccccaac ttaacttcac gcggccgcta agccgaattc 540tgcngaaatc attacactng c 56129717DNAHomo sapiensmisc_feature(13)..(13)"n" refers to an undetermined base 29actctccgcg gtntcntggt gcctcacagg aggtggggct ccctccaccc ggtccccagg 60cctctccctc tgcccgagct tcccggtcct gcctccttcg cctcgcctgc ctgcccgact 120ctgaaccctg ctcctcttct aactaaaagt cagtgtttta tttcctccgc agtccaatgc 180ccgcgtttta ccttattcaa taagaagggc ttcatttatg gcaagacagg acagccaggt 240aataagggcc tctgcacacg cgggcccatt ggaggggcgg aactgcgaag tcttcccgga 300agagcttcct ggagagaagg ggaacgagcc agcgtttatt gagcatctat tatactaagc 360atctgcttgg cagttcacga cggtcgcatt ttttcatcct tacagcgatc cctattgtgt 420cgcttgcttt aaagcctcac agctcacaaa gggctgggat ttattccaga tctctctctc 480agatgccatc tcacttccag gtgtctctgc tgctttgaac gcgggaaacc cacgcaaagg 540agtgatttcc aaggccttct gtttggaata tctttaatcc tccccttatt aactggaaaa 600actcccacgc atccttcagg gctcagctca aatgtccttt atntctgcag ngaaactttc 660ccaaggaaaa ttagttacac agctaatttt agataaattg agccagttga tagaatt 71730280DNAHomo sapiensmisc_feature(30)..(30)"n" refers to an undetermined base 30tgatggatat ctgcagaatt cgggctttgn gacgccgggc acgcagtagg gaaaacagta 60ttaaaacgcc ctacagaaaa tctcggcgaa gtccccggag aactctggtt tctaagatca 120gctgggcgca ctttctccgg gacgtccctt cttctcggtc tcagcgcctt cctgccctca 180gccgcgccng tnttgttttg gtggcaaact gaaataagaa atggaaatat attggccttt 240gctgctgcca gggatgagag gttgttgacg tcggcgcaaa 28031270DNAHomo sapiensmisc_feature"n" refers to an undetermined base 31gnggnngnna nncggcgatg gatntnngna ganttnggtg atggatatct gcagaattcg 60gcttagcggc cgcgaacaaa gagcgaacca aaggatgctt cgaattttta aaacggaatc 120tctgcaccca aatgcaggac tggtgactta aggagctgcg aagtctgatt taccgggcct 180actctcgacc tgccccccac ccccagctca gggggacctt tttatcntga acgccagagc 240tacnnaccaa gtcgggtggc cacnnccaaa 27032347DNAHomo sapiensmisc_feature(7)..(7)"n" refers to an undetermined base 32tttggannta ngggggcgtg gcgtggatcc agtttccccc ggccaggtcn gcttcccggt 60ctcaaccatt tcgcgctctg ctctgtccgc tggtttgtcc ctgcccggtt cctctccccg 120ggcctgtcag cctccgcttc tctggaggtt cctgggactc atctctgatc caccgtcttg 180cgttctctgg gcgcatcgac ttctctccat cttcgggctc actcctgact ccctcgctgc 240cgccccgggg gtttccacgc gtgtctctaa ccgcggccgc taagccgaat tctgcagata 300tccatcacng aantctgcag anatncatcg ncgaannnca ccgcact 34733342DNAHomo sapiensmisc_feature"n" refers to an undetermined base 33gtagggcgcc gccgtgacag attagtccta aagggaacgg ggttgttagt tcaattggct 60accggaaaaa accaggctgg gctgggcgcc cgccatgaca accgataccg gaaaaggcgg 120gtcgttcccc ccggacagcc ctacgccggc aaaggtctcg agatgtgagt agtgagagcg 180cctaccccat acngtcggcc ggctcccctt cttttaccca gtgatctaga cctagtctag

240gacctcggga actaggacca gcctccctcc ttcttggaga tctgaccctc aggattcann 300nnctttgctc acgagctcca acccnacnca tccaaannnc aa 34234370DNAHomo sapiensmisc_feature(267)..(267)"n" refers to an undetermined base 34cattgtttac tttcgtctaa acgcggtgga agcccatgga agaaagcggt tagcagcaag 60gcagagccct gctccctctg cagccccagc tcccagcgcc ctgggctttc caggcacctg 120tccgggtagg ggattgaggg ccgtggccag gcccgcactt tcctgctagc cgcagctggc 180cacatgccca tctgaccctc cgagttctcc tctaaaaatg gggctgacag ccgctacctc 240acaaagtcca caccgggctc aacccgntgc cttcctcccc aacaggactc tgccaccctc 300cctcaggatg cctgagggcc ccganctgca cctggccagc cantttgtga atgaggcctg 360nggggcgntt 37035213DNAHomo sapiensmisc_feature(8)..(8)"n" refers to an undetermined base 35aaaatacnan taaagcgatg cttcgaattt ttaaaacgga atctctgcac ccaaatgcag 60gactggtgac ttaaggagct gcgaagtctg atttaccggc ctactctcga cctgcccccc 120acccccagct caggggacct tttgtctgaa cgccagagct actgaccagg tcggggggcc 180gcggcccaag ccgaattctg cagatatcca tca 21336173DNAHomo sapiensmisc_feature(4)..(4)"n" refers to an undetermined base 36gacnncgggt ttgtgtgtaa cagggtcagt ccccgtatct actttgcgaa agcttcgagg 60cgagcgtgaa gtcaagggct gcggtggatg ggggtaaaan gcctcctcnt cccactgcct 120gcnccgtctt ggggtaaccc ctanccccca cccggngttn cnctttaatg ctc 17337369DNAHomo sapiensmisc_feature(22)..(22)"n" refers to an undetermined base 37tcactgtgcc gggtctctcc tncccggtcc aactccctta cttgtcctca tctctgtccc 60caaggtccgt gacccgcgga ggtgatgggg gggataggag agccccaggg accgcagagg 120tgacacaatc gcccgcccgt cctccctcgc tgggagccga ttcagcctgt gccgagcctc 180tcccttcgcg tgcctctgcg cacagcggtg gcaccgcagg actccgggtc ccccccggct 240ctccatcggg aagccggcaa atgcgcttcc tcagccagac cgcggcgggg tgggggcggg 300gggggcggaa gttgaaatac tgggacagaa acacctgccc gtcccaaggg acggaaaact 360ggatgccaa 36938123DNAHomo sapiensmisc_feature(20)..(20)"n" refers to an undetermined base 38gtcccttcgc cccgcttttn ctttccccna ggtcccagcg nccgaaccgg cggatgtcca 60cgaaacatag ggcgagccgg gggccangcg gggccgtgta aaatctcntg tggtcatttt 120gtg 12339450DNAHomo sapiensmisc_feature(32)..(32)"n" refers to an undetermined base 39ctagccctgg aagagaatcc gaggctcagc cntgctgcag cacccaggac actgcatccc 60agcacctgcc cgaagatcag cccaggaccc aaaggaaagc aggctccaag ctccccggaa 120gccaaggaaa ataggaaaac atatcctgcc ccggggacac cttctggaac tatgaccaca 180tgcacttgac cttccggaac aatcaccgca tgcacctgac ctcccggaac tgtcaccacc 240gcgcgcacct gacctcccgg cactgtcacg accgcgcgca cctgacctcc cggcactgtc 300atcaccgcgc gcacctcacc tcccggaact gtcaccaccg cgcgcacctg acctcccggc 360actgtcacga ccgcgcgcac ctgacctccc ggaactgtca tcaccaggcg cacctgaccc 420cccggcactg tcacgaccgc gcgcacctca 45040593DNAHomo sapiens 40ggaccaagct gggtaaactg ccgacagctc cattgggcag catgtccacc cctgatgacc 60aaatcccacc aaacgtgcag ctggcactcg gccgcctttg tttccttccc ctagaataaa 120actccgctgc tttcccacgt tcctggagca gcagccggaa taaagcgccc atggccttgc 180cctttgagtc tcggaggatg tttgccactc caacaatgga cttttaaata attcaggggt 240caaaaggcgt gtgtgtgggg ggggagaaaa gttacaaatc agcacttgaa accgaacaca 300aacaaaaatc aaacaaatcc gaactaatat aacaaatcaa aactttgatc tttagaagaa 360aacttcaacc ttaatgcttc caggaggaaa gcagaaagga taatgactga attgtgaaaa 420cgagccaaaa tgttccacca ctgatgtcac acacacctat gactccctgc acagatccac 480ggtcccgggc gctgaatccc cgcaaccctc tgcgcccaca gaggttaaac tctcgctgct 540ggcgacttcc gcttcctggc ctaaatctga cacgcacgac tccccccgcg gca 59341457DNAHomo sapiens 41accaccaacc aaatagggcc tttcctgtta acgaccacgc ggcaaggggg ccgggccctc 60gcacgcctcg acggcctccc ccactccaaa gggactccga tttcgcagga tctcccgcct 120cccgcctctg ctcccaacac cctacgtttt tctcttcctc ctcatttacg tatttacaat 180aaaacagcga agctgcacag tctgtctcta aatcaaacgc ggttaccatc aaagcctcag 240actctatgtc tcaaccgcaa aaggtctgac aggaaatcaa ctcgggagtt tgtcaattct 300ttaaactcaa agctctgtta acgaaatctg gatctttcct cgctccccac ctgcctcccc 360tgacaggaga atgactgtaa aaggatcctg tcgtccccga aagtcagcac caagcacttc 420acaaattgtc aaatctcaaa agcttacacg cgcggca 45742211DNAHomo sapiens 42gcctgacctg aatgacgcgc atgttgaggc cggtctcctg cgccagctgc tcgcggatgt 60ggcgggtggg cttgggtgta gcagcgaagg cggccttcag cgtctccagc tgcttggctt 120tgatggtggt gcgcggtccc cgccgcttgg cgcccaggtt ctggtcgtca ttctcgttgc 180tacccgcttc cttgtccgac acgtcggcgc a 21143141DNAHomo sapiens 43aaatcatctc cgggggccca gcacggacac gctccagacc cgtgagttcc ccagcgccgt 60gccgggaggt caggggcgct gaaagaagga agaattcagc cacctctcag catccctgtt 120acctcgagga cgcgcctctc a 14144559DNAHomo sapiens 44acccactttc cattaacact aaataaaacg catccatgga tttcctctcc attccgaggc 60aacaggagtg catggcacat tgccctactc ccctgaagct cttcgctaac ctaagactcc 120agggtgagga agttagctgg agctttttaa agtgcatctc caaagagaat tttgctcaca 180ccatgagagc ccccaagaaa caccagggcc cccttagatg ccggagacca cgccctccag 240gaataagccg caccctctgc ccagcagatc cttgcgcgag tagccctctt tccctggggc 300taatcaagtg catgccacat gtcaccactc tcagctggca attcttcctc agaggcgcag 360actttcacgg aatccccagc agggggggtt aagagattca ggggaggccc cgcccgtgcc 420ttccacaaaa gtcgctttac cgtggctcgt gtcctgcggc cccaaggggg tagcctggga 480cgtgtattgg gagggcatag aggctccttc caggacaagc tgccagcctc cagtgggcaa 540ccatgtgaga ggcaaaatt 55945433DNAHomo sapiens 45gcgaacagca caaaggcttc attcctacga gagattaagt tttagagcaa atggacacga 60tcgttaaaga atttgatatt tccatgtaaa ctgcattagc aggttatgcg atccaaactc 120acaggaacaa ctccaactct cggccatgcc ctatttcatg tctagatttg tttaaccgac 180ttacatcata atccaagaat acgaactaca gtatattctt acagcaaagt tattccttaa 240aagcaaaacc gagccacctt tgaaaacacg cacacacatt atccacggca ctaaaacccc 300agtcttgacc gagaaagacc aacaacttgg gggggaagaa aacaacttca gagccagagc 360tcccaaagca gaaagcgctg gcggctgaag ggcacacgag gttccgctcc cgggcgaacg 420ggcggcgtcg caa 43346487DNAHomo sapiens 46cccttagtat tccatgagcc accattttcc ccacgatccc tccagcctga acgatcacat 60cctactgtgg accacgactc tcccagcagc gggcgtttaa tatccagtta gcaggttctc 120accaccccct cgctggctcg aatacagcat ctgcaccgag ttcccgagaa tcgtcaaccc 180agcaaatccc ttaattggtg gacatgaaaa tccagggctt tgtgctgtaa taacagagtc 240ctgggggcct ggggagtttg tgccgcttgg agctcaggtt tctgggacag aggctgagcg 300cagggcaggg aggcaggtct cacctggcac ctcccagagt cctcgccgag cagatggaag 360cagaggctct cgcgcccggc ccccgccggg agacctctct ctctttccct cggcctgctc 420tgccctctcc cgccttctcc ctgtctgatc cttctctgct gtcatgttct ttgtcctcgc 480gccccga 48747403DNAHomo sapiens 47gtcatataag cacaaccatt cccagggcca ccctggatgc atcagatcag tccccccact 60ggtgaccaca atggctggct cagagtgcct ttgaacagac aggagaaaca gacttcttgg 120agggagggac cttcccacag ggaatggcca aggagctagg tcttcagggc ttgcatggcg 180tggagtgtgt gctcaggtgc acagtgaagc aaacctgagg ggacttgggc cctgcgtcct 240ccagcacaca cgcacccttt cgccgtcaca tccggggcac ccacccgtgg aatatgtgag 300ccgcacttgg ccagccacga gttccagggc caggaagtcg tgcttctcgt tcaggcgccc 360gttgtagaag agcagcccgc tctgctgcac tgtcgcgtcc cga 40348155DNAHomo sapiens 48ggcgtggaga ggagggggca gaaactcagc cgcccctacg tttgctaaac tgcgtccgcc 60agggggcgta tttttctaaa acgcacaaga cgtttcgtgg gttatcgatg gtctcttgag 120cctccttgac tgatggggat tgaccgggcg ggata 15549256DNAHomo sapiens 49tctactgagc ttttctttaa gtggaaccag aagtgctggg atgagaggga aaggatggga 60gtgcgtccaa aggtggacag caggtcccca tccctggtgg gagtgagact ggacggcatc 120ccccggaaag gtggtttggg ccttggacaa ggctagaggc aggagtccat gatgcagaga 180tgacacagtg cccctccgcg tgtgagtcca cgaaggtcac tactgaggct ttgtgcttgt 240aaaaggccgc cccgca 25650224DNAHomo sapiens 50tgcggggtcg tgggggaacc ggcgggagct gttcgctggc cggcctcact ggagtaggaa 60ttttagatga aactgagtcc gtttctcctt gaaggcaggc agtattctta gatctactat 120tcatttaaaa agaaggaaaa gaaaaaaaaa tgactgctac ttactgagaa gaaaatttct 180gttctcctcc gattccgctg atcccgcttt atccgcgcac ctca 22451313DNAHomo sapiens 51gtggctggga cggcccaggc cgcggcttcc cgcgtgggga tgcgctgtgg cgcagagctg 60gtcccggcgg ggccaggcgt ttgtgggcgg gtgacgggga tctagggctt ccgctcgtga 120ttcctcttgg gctgtctttc cgggtttgga ctcgcctgcc cggctgtgtg cagggttccc 180gctgcctctg gccggcaggc gtccgggctg caggtgggcc ggcaggcagg tgttagcggg 240aagggagcac aggtagcgag gtgggatcgg cgacctggct agggtgtcgg cagaatggaa 300tgcgcggccg cta 31352385DNAHomo sapiens 52tacgttgcgc attcattctg ccgacaccct agccggtcgc cgatgccacc tcgctacctg 60tgctcccttc ccgctaacac ctgcctgccg gcccacctgc agcccggacg cctgccggcc 120agaggcagcg ggaaccctgc acacagccgg gcaggcgagt ccaaacccgg aaagacagcc 180caagaggaat cacgagcgga agccctagat ccccgtcacc cgcccacaaa cgcctggccc 240cgccgggacc agctctgcgc cacagcgcat ccccacgcgg gaagccgcgg cctgggccgt 300cccagccaca cccagcgcgc cttctccagg gtcagccagc tgcggctctg ccgaagcgct 360cctccgctcc tttctcgcgc cccga 38553307DNAHomo sapiens 53aacccggctc ggttcggcaa ggttcaggga gacaaggtag agaaggctgg ggtgagcaag 60aagtcgggcg gccgatcgtc agggccacga gcctcgcctt gccttcttgg aatcccaccc 120aactttaaag gcccaaagat cctgaaaatt ccgaaagcga aactgcgggc tggtctccag 180aagtttgaga acggtctccc aggctttcca gcgtcgtccc gggattctcg gacaccacaa 240acgccatcaa ccacgagcac cggtgtccgt ggctattgcc ccgaatggtc cccatccgcg 300tccccta 30754182DNAHomo sapiens 54cgatgtcgaa gccgtttgga gggaacagcg gtttccaagt tcctgctgac ttgagaagcc 60tctgcgggtt tccgaatctc cggcgcactc ctgggcgcgc tgcgggagct gtagctcagc 120cagccaggga gtagcggctt tcatccgccg ggaggagtct ttcgagttca atcgcggggg 180ca 18255523DNAHomo sapiens 55tcgggtttga tccgccccaa ccaaataggg cctttcctgt taacgaccac gcggcaaggg 60ggccgggccc tcgcacgcct cgacggcctc ccccactcca aagggactcc gatttcgcag 120gatctcccgc ctcccgcctc tgctcccaac accctacgtt tttctcttcc tcctcattta 180cgtatttaca ataaaacagc gaagctgcac agtctgtctc taaatcaaac gcggttacca 240tcaaagcctc agactctatg tctcaaccgc aaaaggtctg acaggaaatc aactcgggag 300tttgtcaatt ctttaaactc aaagctctgt taacgaaatc tggatccttc ctcgctcccc 360acctgcctcc cctgacagga gaatgactgt aaaaggatcc tgtcgtcccc gaaagtcagc 420accaagcact tcacaaattg tcaaatctca aaagcttaca cgcgcgggca ctccggaaag 480gctgtgggga ccacccaaag cacccccctc cacaccgcgg gca 52356795DNAHomo sapiensmisc_feature(526)..(526)"n" refers to an undetermined base 56tttactttct tccggctgac gtccatctcc tcaaatttct caggaatgtg gggaagctcc 60tagccctgcc tgcctttcta gagggcttct tggatttgca gcttctaaca agttgctctc 120gccacggaga agctgttatt atgacaaaat atttggggca ttatcaaaat cacacaggct 180gctgggctgc tgtcggtttc tcgccagggc cagtaagcag ttacatttgg agttgctacg 240tgttgtttgg gggccgggct gtggagagtg actgagccag tatttttcat ccaaaattct 300gcaaattgaa ttaaccacaa ttctagtctc acctcccgtc tttaaaaaaa taagttgaag 360aaaaggtaaa tattagagat aaggcagcat ctagtgactg cggagaggca caagctggtg 420ggcgagggtt gggggagtca gcaaagccct tcaaaacctc cccgtttaat tttctggctg 480tctctgcatc ctgttgccag aattccaaat gcttggagtc atttanaggt gcgagaactc 540aaacgtcgtt ccacttggaa aggggaccgt ttaacgttaa attccattag cacctaaatt 600gtttcttaaa gacatccgct cagacacagg actcgaaagc gagcatttca tgcaaataaa 660tttctcaaat tttaaacctt gttaaaagct tgtctcgcac ctcggctccc tccccttccc 720cggaaganaa caataggccg ntggcgcatc cccacttcgg antaaatatt gacgggggaa 780gttgctaaaa acatc 79557438DNAHomo sapiens 57gcctgtgtgt aggggactgg aggtggggga acctgttctt ttcttgtgtc tgatcctggg 60gctcgcttcc tgggtcctag aacagcagcc aggacggaag aaactgttca cgttgcaccc 120ctttctctaa gattcccagg ccaagagtag ctgcagaagg tggccctgaa tctatggcct 180ccttctctct gcctgacccg gctagtggat ccggagaggg gaccagggag agctcctccg 240agcaggggtc cttcgggaga cagagagggg tccaggctga gagaactctt caagcatggc 300gagtctgcgt tatagaatcg ggcgggcggc tcaacttggg ggaagcacca agaagagctg 360ggcgacctgg agcgcagaac cggctttggg gagccacccg gcggggcagg ggtagcacgg 420agcccgggcc gcggccca 43858611DNAHomo sapiens 58gcttccccct tcctttctcc cgcgctgccc ccttgagatc cgacccttcg tgtgtttgcg 60ctaaattgtc ctcgctgttc ctctcacccg cgacatacag agaaagtaac cggagagccc 120tacatatgcc agtctggctg ctgccctgga gtttgcatcg atcctgcaat tcctagtcaa 180atatgaacac tcctctccct cctcacgccg agttgaaaaa cgtgataata atataagtcg 240acaggctgta ggcaatattt ttcactgcat gacttattca tggcgaaaat gtaaactgtt 300ttcccacggg ctttattaga atgttgcctt ctgccgagat aggcttcacg caagccctgt 360gctctcagtt tctctggcgt gagcaaaact gtttgggtcc ataagtgtcc acatttggcc 420atttattatt tcgatttgaa ttttcagtaa atacaaccac ccctttcccc tcccccagaa 480acagtcatgc ttagagaaaa ttatagcaaa acctccctgg aagcctgaag gttttggatg 540actgagaggt ttggtttggt ttctctttct tttcattcaa aacggatcaa actccctggc 600tcgcgtcccc a 61159291DNAHomo sapiens 59gagtttggca ggccccggat tccacaaagg agtaggcgcg gccagccgcc tccagccctg 60agctcagtaa attcggtgtc ctgaatgctc ccttcctgtc cttaccactg cgagctctct 120tgggacagct ttctaggttc cactgcgacc tactttccgc tccctgagtg cttctttgct 180gaaactgcag gcgaaaagat ctctttccca gaccgcagcg cactttgaga aggggctcaa 240agtcgcccgc tctgaatccg gcaccggcaa ataggagtag ccgcatgcgc a 29160226DNAHomo sapiens 60gaaaacagat aaaacgccct acagaaaatc tcggcgaagt cccggaggac tctggtttct 60aagatcagct gggcgcactt tctccgggac gtcccttctt ctcggtctca gcgccttcct 120gccctcagcc gcgcgcagct ttgttttggt gacaaactga aataagaaat ggaaatatat 180tggcctttgc tgctgccagg gatgagaggt tgttgacgtc ggcgca 22661580DNAHomo sapiens 61ctgtgatgca ctcggcggat ctcggtggca gctgcctcct tcatctccag tgacgcctgc 60atgctgtcct aggcagtgtg aggagtgaag atgagatttg gcgcatcttt caacggagtc 120tgagcaaagc taaagggctc cgattcgtgc aagccaaggg ctgcccctcc tatcctgtcc 180tccttgagga cctgtgctaa ggctttctca tccaccaggc caccatgggc tgcgttcaca 240aggaatgctc cctgtctcat ctgctttata gtaaagtcat tgacgaggtg gtggttatgt 300tcattgagat tgctgtgcaa cgagacacag tcactctgat acagcaaacc ctgcagggtg 360tatcagggtc ccctctgcat gccctgggac ctctctatct tgtcctacaa gtaggggtca 420taaaatacga cgctgaatcc aaaggccttg gctcaaactg caaccgcctg cctcatgcaa 480ccgaagccca tgaggcctag cgtcttccac gaatgagggc cactcccatg gccacctcga 540gaatctgctc cacgctctga acccgcgcac ctcaagccga 58062633DNAHomo sapiens 62gcccaggaga agccctccac ggtgggcgtc ctcctagaca accagcaccc cctgcaggca 60ccctcgtctg gcagaatcag ccctttccca cctgcaggcc cttctcagcg cctctgactt 120cccacacaca gcacaggtta caaactggtc cctggcagtg cactctagcg ggcctctctc 180acaagttctg cgggcctcgt ttcatggaaa gcgggttgtg gattcctgct gcccttggat 240ggcccctgcg cacgcacacc tctgagcggg cactgagcga gcgtggggag ctgctccctg 300ggaactaggc aggagctttt aaacaccctt acacacagcc attctgcggg aatacatgct 360ttcccggtaa ggcttttact gttcattcca ggtaaattgg aagtcgcaca ccccaagctc 420caaatacaac tcgttagctg gcaggtctct gaagccaatt ccttctgagg aaaatggaga 480taatagcagc taccctccca ggtgactggg ggagaataaa gtggctgtgc atagtggtgt 540ttgcagctgg tggctgctat tatccttcat tacagcttgt aaaaagggtg tctaggccat 600ttacacacag ataggccggg tggggtaagc cga 63363703DNAHomo sapiens 63gcctatgaat ggatttataa ttgctttatt tttgtcccat ttagacagaa gtcagagaca 60gaggagagaa ccaaaaaact tggatgtttc cgtaaactag attcgtcaat cctcgataat 120tgaaagtagt tccagtatgt cagccaccgg ggttccctgg ggagctaacc agtcctgaag 180gaagtatgaa gaggaagagg aggtcttcag ttaaggggat gaatttgtgc agtcctaagc 240cctgcaaagg tgctggaggg aggaagaagg gcaggaaata aaagatggaa gaaaatttgt 300tttttatcca cttagagttt tatctttaat gatgggaaac agtgctgctc tcaggaaact 360cagtgtggag atctaggagt tcacggttca tagtccatta ggagcaggaa aaggatagag 420gacatttata aagtaacatc caagtccaaa gtaaaatggt ataaattgtt tcccatgata 480aaggctggct gagtaggtca ggaaaggtct tgtcagacca tatgtgctgt ttcaggctgc 540ttcaaattct tttaggacag tggtggatat gagtgaagac ggggcaggca ggccacatct 600cttagaagag gaaggtgatt gccacgtctc cttcctccat gctgatggca aggcgtgcgg 660gctgtgttct cttgcagcca gcgtcccatg ctcggcggcc aaa 70364420DNAHomo sapiens 64gtgacgtgcg gaatacacgt gatgtcgggg acaggagcgg gctgaagagg gcacgatccc 60acgcggaggc cacccctcac ccggggtagg agcccgctgc acttgctgtc gctcagcccg 120ggcgctgcac cacggcagcc gccatgctgc ttaaagccgg tcatgtgacg cgggagccag 180ggtggaaggg gtccccgcgg gcaagccttc gacacgtgac ctgccacccg actacggaag 240cctcttgggc gttccgcccg ggctcacatg tcatgtgacg gccggccggt cgccggagta 300accaggaact ttcccagacc ctgcggtccc tggagcgtca aaaagagcgt ccccgtgact 360aggtggagtc gcctgccctt ccgaatctca gctgtcttat ctggaacccc cacgcggcaa 42065496DNAHomo sapiens 65gcgctgcacc aatttagagg gtagaaaaag gagttagaag caaagaggaa aaaataaata 60aacaggcaac aaaaacccaa cccagccagc ctgagccatt tgcattagtg ttcatttagg 120aaattagcag acgggaaacg ctggggagtg gagtgggccc cggccttggg gactgcagag 180cccgctcagc cctgggtggc tgggcccaca tggctgtcgc caggagcaca ggaggaccca 240gaggtggccg agggagcctc gccgggctcc ggtatgggtc ctggcccctc acaggtgcga 300gcctggccca gtgactgtgg acgctgtggg agagcaggcc tccgatacgc agggctggga 360ctgctgacct ggaaggtggt gccgggcgtg tctggtgaag gcgccgttgg cagctagaga 420gagacggcgg atggggtgac gccataaccc acggtcccag ttttgaggct tgacggtgac 480ggaaaaggac gtcggc 49666637DNAHomo sapiensmisc_feature(612)..(612)"n" refers to an undetermined base 66cgccgagccg ggatgagcaa ggcttcctgg aggagagggc cggcctgagc ttggaaggat 60ggggaggagc cactggctac aagggtgtag aggtgagaac cagtgtgacc tgcccatcgc 120tggtcgtctc tgggtcattc agctgaaatg gcatctctga gctgagagga gtgttgcctg 180taaggagcta ggcatcagcc cccagtagag gggcggccca

ggcacagccc atagccgcag 240acttagtgag tctagctagg gagacagtag aggggccaaa atgaggacac aggtcaccaa 300aaatcctggc caggtcctgc cactacctgg ctcagcgacc tgcccccccg agcctcagtt 360tcccccattg gtggaatgga gtgaggaaga cgcgcctccc ggggctgcga tggagaattg 420agtcagagtc tgggggtgct gggagggctg gggagcagcc tccctgagcc tcagtttccc 480tggctgggga atgaggacct tgctcgtccc ccctcataag gggaagctgt caggaaagtg 540ctttcaacgc tgagccattt cccagtggtg cacaattagc tttccagagg attttggtgg 600attctagagc tngagggctg ggggatnggc ggccaaa 63767595DNAHomo sapiens 67gccctgagct cttgagggcc tctgcagttc ttgggacaat tctgggacta tatctttggg 60ccttggtgag atctagaggc tctaaagtct ttgggagggg tcctgagctc cgtggacggc 120agggtcttgg gcactcactt gcattcttga ggggtgtgtt tggcctcgtc cgtgcaggtg 180tagaatttcc cctgtagaga ggatgtctgt caagtaggtt cacccttcat cacactcccg 240cccagacccc tgcctggcat tccctccagt gtttgcccca ccttgaagag ctgcaccccg 300atgcaggcga acataaattg cagaagtgtg gtgacaatca tgatgtttcc gatggtccgg 360atggccacaa atacacactg caccacatgc tgcgggcacc caagcatatg gctactgaac 420actacaggcc acagtggtca tggggcaggg actctggtca tagatgcagc tgagggactt 480gggctgggga catgtggtga tgggtcaggg atgtatggtt agcaacatgt gttcaagagg 540cagtgttatg ggctagagac gtgtgggcat ccaccaggaa taagtgtttg ccggg 59568580DNAHomo sapiens 68gagtcaggac ggaggacgcg gcaggtcaca gagcccacca agtccgaagc tggaagttca 60gattctttga tattcaaagg tggatcatct gtgctttttt ttttttatca gtctctcact 120ttttatccat catctaattg tgacagctta tttgccttta taccataaga tggggagtag 180ggttgagatg aaatccaagc atcgtttccc ttccccgatg gtcgcctccc tggggtgaga 240cgttcgacgt gtcagacttc accaagagca tctcccgcct cggtgcagta atgaacttgg 300aaacgattta ctccggcact tggttcctgt ctccataaat gcggctgctt taaagggaat 360gtaaaaaggg ctgtaaattg gtattgattg ccggtggtct tgaagaaccc caactgagga 420ttgaccgttc cttggagtga aggctccgca ttcagacgcc tttcgcctta cgtcatcata 480attgagaagg gaaaggagac gtgttagttt cagtctgatt atttaccatc aaggcataaa 540cacttctcag aggcagcgga acccattaaa ccggcccgta 58069589DNAHomo sapiensmisc_feature(559)..(559)"n" refers to an undetermined base 69acacgggggg caacctcttg cacctggctc cctgccctcg gtgccacgtt tccagggttc 60ctccacgtcg caggctgtgt cagcctcgct ccttccactg cagaattgcg gtccacagcc 120tggatgggcc actctccatg tatccacctg tccctccgtg gctgctgggc tgagtcgctt 180ctgatgctaa caagaggcgt ccggctggac taaggccccg gaagctgaga actggagggc 240aggtgcgggc atcgggcaga gcagctccag caggcaggac ctggggcctc caccctgcac 300ccctgtgccc cgcgtgtggc ggaaccgccc cgaggggagg ctgtcaccac ggtgacaggc 360agccccacgc gagcctgaga accctcagcc cacctttttc tgtaatcaca gcaggcatct 420ctccggcaag tcaatccagt tccagctggt gctgcctccc ttgcctcatg ggctttattt 480tagaactctg agcaataata aaaaagacgc tacccgctac aatagatgtg gcagagaatc 540tggctcttca cttcatcana gatcaccctg aaatgatggt tgttgttaa 58970748DNAHomo sapiensmisc_feature(10)..(10)"n" refers to an undetermined base 70gctacatctn ctctacattc taactaacac ttgttatttt ctgtttttgt ttgtttgttt 60ttaatagcca ttctagtagg catgaagtgg tgtttgcctg ctttttttga tggaggtgga 120ggaatagggt ggaattggtc cttaaccatc aattaagctg ggggccttag acctctgtga 180attggctgtg acaatagcta aaggaggctg ctacctcata ctgaagagat gtttcctaag 240tttgtcaccg gagagggcac cgaaccaact tattgtcttg gagggaagaa gcagcaaggc 300agaagacttg aacttctcag agaaaaaaac agtctacaga cttcatttta tgctgtcctc 360acacactact gaaagctcta ccctggggac ctggcttgac ttctaaccta cncctgtgtt 420atttaggaag agctcccagc tgctctgagt ctcagtctcc caatcagtga aatggaggca 480atagcacctg cctggctgca tcgccccaca gtgctgcaat gagcatccaa cgagagaaag 540cttgtcacct gtgttgcaaa ctaagttaca caaatgcagg cagtagcagc tagaagaaaa 600tggttgggaa tctgaaaaga attaaagccc cccatgaatt tcttctcacg cctcctccaa 660aagccaggga ctgcttcacc ccgcctccag gactgctcgc tccagcattt ccggcagctg 720ctgacagaat gtatgttgcg gctgtccc 74871599DNAHomo sapiensmisc_feature(491)..(491)"n" refers to an undetermined base 71gatgactgtt gcccgagctg aggccacgac ccaaccccga ggaagggaga acagcttccc 60atgaagggca tggctgctgc cccataatcc cagggcagga aataaaggga tcttggacta 120ggcaatcaaa ggacttcctc tccctctaag gccaaggagg aaatgtggct gggactccaa 180gctctgtgga tgcctggagg tgccagcagc tggggatcag ctggccccac ctgcagagcc 240agccagtggt ccccctgcat ctccaaggtt gggtctatgg gctccaagaa caggtgtttc 300tcagggtaac ctcagcccct acaacttcaa ccaagagagt gaaggggagc agccctggag 360gccaatgagg agggggatta gtggtcactg atgacaaaga catccctgtc cccagagcca 420gccccttgtg agcagaagaa tggctgccgg gcaaaaggac ctgctatgcc ctccccatac 480acatatcatg ncacctgggg accctctgaa taacaggggg cngctttaga gtggcttnat 540taccaacaag aggcccagaa gggctagagc acacgatttc atgntcggcc gcatgncaa 59972614DNAHomo sapiens 72gtgcgctatc acgactgttg cccgagctga ggccagaccc aaccccgagg aagggagaac 60agcttcccat gaagggcatg gctgctgcca ccataatccc agggcaggaa ataaagggat 120cttggactag gcaatcaaag gacttcctct ccctctaagg ccaaggagga aatgtggctg 180ggactccaag ctctgtggat gcctggaggt gccagcagct ggggatcagc tggccccacc 240tgcagagccc agccagtggc tccccctgca tctccaaggt tgggtctatg ggctccaaga 300acaggtgttt ctcagggtaa cctcagcccc tacaacttca accaagagag tgaaggggag 360cagccctgga ggccaatgag gagggggatt agtggtcact gatgacaaag acatccctgt 420ccccagagcc agccccttgt gagcagaaga atggctgccg gggcaaaagg acctgctatg 480ccctccccat acacatatca tggcagctgg ggagccctct gaataacagg gggcgcttta 540gagtggcttc attaccaaca agaggcccag aaggggctag agccacacga tttcatggtc 600ggccgcatgc gcaa 61473552DNAHomo sapiens 73aagcgcccac agatggccaa gcatgtggag gagagcacaa tattttattt aaatatccaa 60atacgaacac attcccgcat ggcaccaaca gccgcctgaa cacgcccgat gccggcttgt 120gctttttccg ttttgtctag aaatttgggt tgcactaaat tctcagctga atgaagatga 180gaaggggctg gcagaggggg tggctccagc tctctgagaa cctggctcct tcccgggtgg 240cagggagaga tggcccctgg ggagacgggg agggtgcact gcctcatgcc caaaccacca 300gcttctagtt gagaaatcag aattttctct gcagaataag gaaaaagcat tgtcaccatg 360attcacgtgg agctggccac actcaggaaa ttcaatgggg tcccacaggg gctccgaggg 420ggaaggagag ggcctgggac atgcccctcc agccatcatg gaacaggatg ggcagggccg 480gccctcactg ctctctaaca gtgaaaagcc acatctccac tttggaaaac acaggcatgt 540gagagcctgg gg 55274450DNAHomo sapiensmisc_feature(378)..(378)"n" refers to an undetermined base 74tggaggcttc gagggaagtg aggttccctc ggacacccta gtgggaaggc tccacgcggt 60aatggaacca cgctgtgaaa cctttgcctt tgggtgtcat ggtggaagca aatcttagaa 120gacatttaat ttaaaaaatt cagttttaaa aaatgttgac ttaaaaagca gttttgaaaa 180acaacctgga attagcctga gatcgatgcc aactcttagc agtctgtata ctaaacacag 240ttaaacaact gtagctgctg gcaagctgga acctttttgt aaagaagcac ataaaaagga 300cagaactggt ggaaggtgca ctggtctttc cacatcgcca ccaggcgttt tgaagcgtgc 360tgctgacacg ctactcanat gcttctggaa gccaaacaat aanaaaaanc cccattgttt 420cccttgctgg gttttacccn ccatggtgga 45075432DNAHomo sapiensmisc_feature(417)..(417)"n" refers to an undetermined base 75ggacaatgag gagggggtgc acgtggaatc cccacggata ggccggacgc cgggcaggag 60cctttgcagg ggtgcacagc ctcctctgga agccctggtc gctgcctggt gcctgctgca 120ccctgcgggc tccgcagcgg tggagccagg cctgaactgc ctgctcttgg ccccgcctgc 180ggccctctgc cctttgtctt gcccgtgggg cccggggcct caagctggcc cggggttcct 240gaagttagct gacgatgggc tggcctctgg ggctgggtcg tgggccttgt gcactggccg 300ccacgtcacc agcgccaggc ctacccgcgg tgctgctgga gacgcgggat gcccgggctc 360gggctgtgct ggatcccctg gcgctgcgaa ccccgtaccc ctttccaatc gcgggcncgg 420nttaaagccc ga 43276501DNAHomo sapiensmisc_feature(18)..(18)"n" refers to an undetermined base 76gacgagacct agccggcncc atgcgcgcct tgagcctggc gaacagttcg gctggcgcga 60cgcgcctgat gctcttcgtc cagatcatcc tgatcgacta gaccggcttc catccgagta 120cgtgctcgct cgatgcgatg tttcgcttgg tggcgaatgg gcaggtagcc ggatcaagcg 180tatcgagccg cccgattgca tcagccatga tggatacttt ctcggcagga gcaaggtggg 240atgacaggag atcctgcccc ggcacttcgc ccaatagcag ccagtccctt cccgcttcag 300tgacaacgtc gagcacagct gcccaaggaa cgcccgtcgt ggccagccac gatagccgcg 360ctgcctcgtc ctgcagttca ttcagggcac cggacaggtc ggtcttgaca aaaagaaccg 420ggcgcccctg ccgttgacag ccggaacacg gcggcatcag agcagccgat tgtctcgttg 480tgcccagtca tagccgaatt c 50177826DNAHomo sapiens 77gcgccctgtg gggatgacgc accatcctgt ttgtttgcac caagtcattt atctcgtgca 60ccccaggggg ccgtggtccc tgccgggcca tcatgtctgc ttcccttatt tgggttttct 120gccccctcac ttcatttctc acttcgcttt tcctccttat ccctttgcag tcttgctttt 180gggggcattg ctcagccagt aatttgaggg acacctcgtg gagccctagt gtggagccgt 240cagagcctgg gtaggattct ccgtggtgag gtgctcaggg agacacagga gcattccggc 300gcctgttcct tgtgcacatc cgcaagtgtc tgcagtgaga ggcatgggtc ccatcttgaa 360tgccaacaat gtggcaccca caccccactt gatggggccg agccacagct ggccaggttg 420accaccatgg acgtgccaga ggcatccgaa acccagctct tgcccagctg ttccactgcc 480aactccagcg ttagcaaagc agctctccct tgctttgtct tctacagcag agaacagatt 540aaaagagaag ctgcaggcag agaaatgcct cttggagcca gatgccccaa aggatctctt 600tgaacaaagg gttgctcagg tcagcgttag ttcctggcat caagcaacaa aatcagagat 660gctaacagtt ctcagattca ctccaagtga agactcaaag ctggatttat aaatccccac 720agagccgctg tgcagaggta gagggccggt ttcaggatga ggaagccctc ttggaagcac 780cgtcctccgg ctaacaagcc tccaacctac tgtcggcagg gagaac 82678433DNAHomo sapiensmisc_feature(16)..(16)"n" refers to an undetermined base 78tgcgcagctc cgcgangtgc ccggcgggcc cgaccctcag actcgcttgt ccctggagac 60caaccctagc gaccaggctc tgccggatcc cgtcgggttt caactcctat tccgaaggtc 120ctttctcccc taatcacaac acccactcgc ctctttttcc tcctcttcct cagcttccac 180cgccgaccgg gcagccccag ttacccgata acggctccca aggccccgtg tttacattct 240ttcccactgg aagcagaaat tatcacgccc aaattcctac ctgccttccc tggattcctg 300gtttcctaag aaacgggttt ggcccacccc tgggcgttcg aacagtccac agaagcgggc 360aaaggaaaga cgactcagtc tttcccctcc gccaatctct tctccgggac cacagatccc 420agaagtcacc gcg 43379424DNAHomo sapiens 79ggcgggcccg accctcagac tcgcttgtcc ctggagacca accctagcga ccaggctctg 60ccggatcccg tcgggtttca actcctattc cgaaggtcct ttctccccta atcacaacac 120ccactcgcct ctttttcctc ctcttcctca gcttccaccg ccgaccgggc agccccagtt 180acccgataac ggctcccaag gccccgtgtt tacattcttt cccactggaa gcagaaatta 240tcacgcccaa attcctacct gccttccctg gattcctggt ttcctaagaa acgggtttgg 300cccacccctg ggcgttcgaa cagtccacag aagcgggcaa aggaaagacg actcagtctt 360tcccctccgc caatctcttc tccgggacca caaatcccag aagtcaccgc ggccgctaag 420ccga 42480285DNAHomo sapiensmisc_feature(14)..(14)"n" refers to an undetermined base 80caaccggggg gcanaggcga tcaaaantgg ggtgcgctgt ggtgggcgac acgtgtggcg 60cgggtctcat tatccgccct tttcacttcc tggactggaa atggcagacc atatgatggc 120aatgaaccac gggcgcttcc ccgacggcac caatgggctg caccatcacc ctgcccaccg 180catgggcatg gggcagttcc cgagccccca tcaccaccag cagcagcagc cccagcacgc 240cttcaacgcc ctaatgggcg agcacataca ctacggcgcg ggcaa 28581401DNAHomo sapiens 81cagatatgta tcctcctctt tccaaccctg cgtccctttg aggcctggtc ggcgttccca 60acctgcccct accccaccaa cccctgtccc tttggccatt agtcccggat tatctagcga 120tgccccgtgt accgtctggc tttgctgttt actccgcgct cggccagttg aggccttttg 180tatttattcc tgattttctc ataggggtaa agtgccttcg ggaggatagg acaagtccca 240tcctgttcat acgaattaca gctcggactt cgggcccttt tacactgcct tttgtatctg 300ttaacttgcg ctaaaaacga ttcggttctt ttttttgagg aagggggttg gggggcggag 360actctgtcgc ccagtcctga gggccgcggc gcgcaagccg a 40182268DNAHomo sapiens 82atagcgcgca caactgtgtc tcttacccag gcacatgcac tatccctgat cccggtgcat 60gatgggaatg tagtcctgca gccctgtgac caaagggctg ggagtgttta tgagacagca 120tctctcagca agcaaagcaa ggcctgcaca gccccgcctt ttcctccagt gaggcgcact 180gttcattaag gagtgttcat gagattacat tttccatcaa gcccagccag tcacgcacag 240ctctacctct tcctctgccg ccccgcaa 26883989DNAHomo sapiensmisc_feature(878)..(878)"n" refers to an undetermined base 83gggtaatggg ggtgaacaga gagggatgcc gaggccagct tgtagtgtgg ctgttggtct 60tgtccatcct atggcacaac cctgtcacca cccagatttt gttaggagtc ctcccccaac 120ttgagagtgg aagctccttt ggcacaaaaa ggggttctgc atcatccccc agcccccagc 180cctgagcctg ggtctggctc tgaactagac ctccatgaat gaatgcacag catcagtggg 240gatccaccat catggggaaa tagtagatac aggaatgatt ttccaaccag attacagact 300atttcaagcc cagccagagc ctaccaggcc aacattcccc aggcttgtgc ctctccgagc 360ctcagattgc tcatccttca aacgagggac agctctgctg gcattacctg aactctaggg 420tcctttataa gctcagactc cagcttagag cacacattga gaggctgctg caccccagag 480ccacatacgt gcaacagagg gtggtccaga ccccttattg gtccccatgg ggtttgagag 540agaagcctcc agaccagctc aacttctccc tcatctcact taggcctttg cacccagctc 600ttaggaggtt gtcaggtcac agtgccccat ttcttttctc ttccccagaa atcatgcggg 660ggatacctgc tcagacagga ccttcatgaa agccaggctg tgaggtgtgt tggggaatgc 720ataattgata ggccatcgtt cggaggccct cctggaggac caaaatgtaa tcagcagtgg 780cgagcttgtt cacgacagga attcctttta catcctggtg aggccaaaga cctggcaagc 840aagtccctct ggtcattaaa gaagcatcct gacttgangc aggncacctt aggtcactgc 900agccacaaaa atctttgntg ctggattcna aagtaggcat tggggctggg atctgggctc 960tggcatcctt gancgtgtcg ggggccaaa 98984250DNAHomo sapiensmisc_feature(37)..(37)"n" refers to an undetermined base 84cgggctcgaa acttcgaaga ccgcggaacc cgaagcngcn cttggctcna atcgcttcgg 60ctcgaggcgc ccgtncgggt cacgtgaggt gggggcgggc cgaagagggg ggctcccctc 120ctcctgccgc agggttggcc gcaagtgcgc ttcaagaggc gcttgatgac ggttaatgtt 180gcagcccgga agatgacttt tttctcctcc ttgggttgcg gcaggccgtt agtgggaggt 240cgcgtcccga 25085402DNAHomo sapiensmisc_feature(224)..(224)"n" refers to an undetermined base 85ttctcccttg tcatcccctt accagagcca cagaaattat ccctgtgggc tcccttgtcc 60tcactcggcc ttttctggag ttaagagatc caagccaact actgggtctg ttccctgcta 120aaatcttagg ccggcgtccc atccacccat ccccatgcct aggactttta agctggcaac 180ggtacctggg tttagttttc ccttcgtata tcactatctt cgtngcttac cttcttgtgc 240ctaaagttcc accgatgtgc aaggngatta accactaaag tgcacctgac actactcttg 300acaaattgca gttgggaggt gagttgatga ctggccggta aatcaaaagt gcttatttag 360ggagtgaggg ggcccgcggc anaagccgan ttccagcaca ct 40286595DNAHomo sapiensmisc_feature(157)..(157)"n" refers to an undetermined base 86gatcccagaa ggttctggag ccgagtatca gagtttgagc agcgagtcca gcctagcaga 60agcgggtgtt gaccggagac ttttcaatgg tgcaaaatga cacactgctt ttgacttggg 120gatctgtccc ttgtggcacc agaagctaca acaggtncac ctggattcca gctctagctg 180gactcggtaa ttgctaagtg ccagctctga agtctgtgat tccgtggaaa tccctttcaa 240gcccgaattc tgttttttat gggcctcttg tccaaacagt ttgacttgtg aactctgttt 300ctgtcaagtt gacacttggg cttggcaccc attcatgagc cagatgaaag cggctaaatg 360cccgaaaaaa taaaggnttt tacctttttt ttgaaccatt ggtgagcatn taaaaaaatt 420agggaaggta aaacccaacc nggncaaacc caactnaaca nttttttttt ccnaaacaag 480ggggggctan tttttcactt ggaaaaacaa acaattttaa ttgantcttg ananggtgga 540naaccaaaat tttttgttgg gttgggttcc gnagnccgaa ttntgcaaat ttctt 59587304DNAHomo sapiensmisc_feature(279)..(279)"n" refers to an undetermined base 87cgtggcccga tgcattcagg gagccctctg tgttggccgc atagcaggtg tagttgccgg 60catcctggat gaagacgggc gcgatctgta gaccccccga ttcaagaagc atgaacctag 120gaatccggac agagccactg gccagaatgt ggttttctaa agaacagtgg agaaaagagg 180catgttacag tcgtaacgct tgaaggaaat gaagatagtg gttagagcca taagcaagta 240atatggttcg gctccgtgtc cccacccaag tctcgtctng aattgcaatc cccacgtcgg 300cgca 30488296DNAHomo sapiensmisc_feature(9)..(9)"n" refers to an undetermined base 88ggctttcgnt aggagttaat ggggcattgg ngggtgggat ggcagggctg ccagcatctg 60acccaggagg ctgggaggag gctgctgtgt gaatacacgc tcggcctctc acagtggctg 120ccgccgcatt agccccttgt gcttcaggga acagagcatc cgtgatggat gagactttaa 180ttaaagtaat gagacattta taatcgcggt tatctccaaa attaggcctt ttagcaatta 240ttcctgggga atattcctcc ggtagatagc tcccttttta gaacaacgtc ggcgca 29689220DNAHomo sapiensmisc_feature(10)..(10)"n" refers to an undetermined base 89attggcccgn caggcgggaa acangctgnn nttctctnac cgttntccag cactgcccag 60accaggaggc gcagggagag gaggggncag cggttccgng accgctcctc ccgctgtccc 120tgctctccag cctntgcctc tgcaggagcc cgcgggantt gccccaggcc cctgtcccca 180cctgtggctc ccgtcctggt cgctcccggg gccgcggcaa 22090273DNAHomo sapiensmisc_feature(2)..(2)"n" refers to an undetermined base 90gnagggnggn ggtcgcggac gccggtgggc agttcttgtt cggtgatgtg ggttaaaaag 60gactgcagcg aggagccggg gcggcgctcg gagtaatcac cggcggcatc aaaaagcgcc 120atcatggcat cgaggtcgcg gtctgcttgg gagccggtgg cgccgccgcg caaggcagat 180gcctgcaggc gcatatccag ctcggtagcg ctccatacct cccacaggat ttcttccaca 240gaggcttggg cttgtatagc ctgccgcccc gca 27391361DNAHomo sapiensmisc_feature(10)..(10)"n" refers to an undetermined base 91acggcttctn tnctaagtga cacggtgtgt gaaattcggt tggggaggta gttctgtaaa 60ctgcgtctcc ccgccagcta aggaagttga gtgaagggag cgttgccgtc tgggaatcgt 120agtcctcaca aaggcgtgag taggcggcaa ataaggattt gggtttagcc ttggggattc 180actcctgtca aagctgttag agaagctccc anaactcnta aagtaacaga aactacttgc 240ggcaacattt gtaacttcca cctggctcat tatcttccac tgttaccttg tgttctagat 300aagttataat ttattctaca tatcgttcag aagtcttgtg cctgttccat attgtnagca 360t 36192462DNAHomo sapiens 92gctgcccaca ctggatggga aggaccggcg cctgcagcat ctgccctcca agccttcgta 60gctccctcct tcctgcagga taaactctaa actccttagc acaacgtggg agccttctca 120gagactgggt ccaacccatc tccagccgca gcctcccctc ctggccccac tgccacaccc 180ccgggcctcc ggccacactg agcctctccc ggtttcccag gatacaacac tcgcccattc 240atagtgtggt gccttttgca cgtgctgttc ctctgcttgg ggatgctgtt ggtctttctc 300agccaggtga agaggacgct gaatgtcacc tgcttgagta tcaggaccgg ggactgggcg 360ctggacctag actcttggcc ctggagagaa gccctgcatg

gggccgcagc ctgcccccgt 420ccctgctcac agaaaagctc agccttgcag ccgcgtggga ga 46293591DNAHomo sapiens 93caaagtcacc tccacggtgc ggctcagcag ctcggcacac ttggtcatgg tgtcggggaa 60ggcgccctcc agctgtaggt gggtagtggc agaacaggag ggtgagggga gagtccgaac 120tgtccccact tggccgttcc ctccccactg gggggccctg agccagtggc ctcctctctc 180ggggcctccc cggaaggagc caaggtctgt ctgcgaggca ccggtccccg gccacggcca 240tcagccccca gaggtggatc agggcatcac ccccactcca cagctgaggc cagggggtca 300gggaggcaac cagggcagac ctggaacctg gctctgagac aggacggccg agggcccctc 360cactctccct ccctcggggt gggcactgac ctggacgcca aagatgtcct cacactggtg 420gcgtttgagt agggcccact cggacatctg gccctgcagc aggttggtgc agacggccat 480ctctccacat gtcacatccg ccccgaagcg cttgcagatc cgtcggaagg gcaggttccc 540acactgcggg gggagcagga cagacacaca tgctcttgca cgcgcacctc a 59194279DNAHomo sapiensmisc_feature(3)..(3)"n" refers to an undetermined base 94ttntgagttt tggcctgccc acagtctagc cctggacaga gaatccgagg ctcagccatg 60ctgcagcacc caggacactg catcccagca cctgcccgaa aatcagccca gggacccaaa 120ggaaagcagg ctccaagctc cccggaagcc aaggaaaata ggaaaacata tcctgccccg 180gggacacctt ctggaactat gaccacatgc acttgacctt ccggaacaat caccgcatgc 240acctgacctc ccggaactgt caccaccgcg cgcacctca 27995351DNAHomo sapiens 95cctttattat tgttaaacgt cacccagaaa acccttaact cttagacagc ggctctcatt 60aagcaaaagg ggaggcacat gaagctccag gcagggccgg gagggaaccg tgaagccaaa 120ggctctggga gcccccaggc acctgcgttt gcattttcat cctggaggag accaggcctc 180tggggctgct ccccggggtg cagagaggag gggtctttct tggtgtgtaa catactcatt 240gattcagtca cctgaccttt gactccatgt attttgttga gtctggatgt gtggtgtgct 300ctgcccagca gctgggatcc acatgagcac agacatggtc cccccgcggc a 35196171DNAHomo sapiens 96ttgagtgtcg cgtgaatacc taggggacac tcaggggaat gatggctccc ccgagaggta 60aagggtggaa agaaggggcc tcagcaggtt aggtcttgct gggtccttct gtagggcgtc 120tgggagatag atccgtgggg ctcctagggt cgcccctacc cggcgcgggc a 17197743DNAHomo sapiensmisc_feature(155)..(155)"n" refers to an undetermined base 97cctccctggc ccttgttccc aaggagcttc ccttgtccca gcctcttcgc cagtgacttc 60tcactggacc attcctttac aaggagcctg ttttttgtgt ttttttttta cacctttttt 120cttctatttc acagaaggaa caccggacgt ccctntgtga tggcagcagc catgctgcct 180ntgtttccgc tcaggggttc tntgccacct ccaattccac ccagtctntt ggcctcggct 240gggcttcggc tcccgcctnt gngccaaaaa ttgcaatgcc cgcggtcagg gcnctttgcg 300gagtctcacc gcctgcggag gcttgattcc ctcctcacag gcagcagcgt ttgatggccg 360gtgacncccc cctttccaag cacatntntc atggcccctg aatgccactt acagggcgtc 420cctccctgtg ctaagtgctg cctgganctt tgggtgtggc agcagcaaan acctctaccc 480ttgnggatgt tcgtttcggg gnggaaagac anatancaaa gttggtcgta aactgtaaag 540tgtgctggga ggaaactgag gcagggaggg cctggtgcca ctggggagcn ctgccccgac 600cccatgtgct tcccaggctc ccttggagcc acgtggatgg cgacttcctg accttggagg 660ccgnggncct cantcctcat gctcgatggc gtcanccccc tcttggggaa atccaancat 720tcctgacctg aaaatgcacc cnc 74398589DNAHomo sapiens 98ttgccgcgct gataaaggaa gcgtctagaa ggtctcccca gccttcatca tctgagactt 60ggctttcagc cccaaagcac taggccctgc tgttaacctt ccaccattaa cctttggtgc 120tcttcaatta gcagcagcca ggggtccttg gcaggtatga gaatttggaa ggacagcccc 180agggcatggc ccccggctgc agcaaaagtt ctaagtgttc ttctgttgga aggaagccca 240ggagatattg atcagctgca ggtgggggag gccccagatc ccacccttgc ctgcctccag 300gagaaggttc tccatgggcc aaaatggagg cagagtccca ctttgcctgg gcagctccct 360gagcatggct ccctgtggac ggagctgagt gacgtcatga ctctaggcct caacaaaaga 420gctttggaaa atcccgatga ttcgaattgt attaaatcaa caaacatcgg gttgcacagt 480tactagaaaa cggagatctg cgtcatcact tactagacac gtgaccttga acggcggctt 540ccccgtgtga aacagcaaag ttctgtaacc cccatgaacg cgcctctca 58999538DNAHomo sapiens 99tgccgcgtct gaccctactc tcacaaagac tttccaacta gcataattga gttaaatggt 60ccccccaact cccttaattc aagctaaact tgcagtttaa caactatagg agtgatatct 120acacattaat gccacacttt aacatgccta acactacaca tgaacacgct tccgggtgct 180gttacatccc gctctctccc aagcacgaga cacaggcagg atgctgacgt cctgcttctc 240tgctgcgggc gggaagtcaa gactccggat ttgctgcagg agttgccgtg gggatcctga 300cttcacgcag gagatggtcg gcctctggaa gtgcctggcc cgtttatcct tgaaatctac 360ctgtgcaggt ggtccttgcc tcagcccctc aggacaacac aggtctttcc taagttacag 420ggagaccatc agattgtcgt gtccgagccc cctgaagtgg aacccacagt ctccattcag 480tctgccctca gtttccctcc cctctgcagg gccattgctg ctgtggacgc gcctctca 538100486DNAHomo sapiens 100agaggtagaa aaaggagtta gaagcaaaga ggaaaaaata aataaacagg caacaaaaac 60ccaacccagc cagcctgagc catttgcatt agtgttcatt taggaaatta gcagacggga 120aacgctgggg agtggagtgg gccccggcct tggggactgc agagcccgct cagccctggg 180tggctgggcc cacatgggct gtgccccagg agcacaggag gacccagagg gtggccgaga 240gagcctcgcc gggctccggt atgggtcctg gcccctcaca ggtgcgagcc tggcccagtg 300actgtggacg ctgtgggaga gcaggcctcc gatacgcagg gctgggactg ctgacctgga 360aggtggtgcc gggcgtgtct ggtgaaggcg ccgttggcag ctagagagag acggcggatg 420gggtgacgcc attacccacg gtcccagttt tgaggcttga cggtgacgga aaaggacgtc 480ggcgca 486101450DNAHomo sapiens 101aattgaacca gggtgcacgg ccagcgccag acacagtgag cttcatggca actccagttt 60accggtgaga accatggggc cactcagaga ggcaaagagc ctcacccgag tgagtcctct 120ggcttctccc cacctgggcc gggccccagg ccgcgctgtg gttccctttc cagccgtcat 180ccctgggtga tgggaggtgg gcattctgtt caaccttgtg ggtcagggag ccagggccag 240tgtgcagatg agaagaggct gcggttactg gcgatgcgag ggactgtccc cttcgtgggc 300actttctctt ttgaggccag tgaaatgtgt tccctggggt tgtattcctg agaaggcctc 360atttaaaggg agccgccaaa ccaagtgggc ttagcaaaag cagtttgtca cctggcagca 420cgtgtgagcc tcgcccggac gcgcctctca 450102292DNAHomo sapiens 102agcgcggcct ggcagattgc ccattaatga aactcagtgg gcagaggctg ctgagggaca 60cggattccca ctccccgggg gagggggtgg aaatggcttc ctccctctgc ttccctacca 120ccagtaatgg ggagctcacc atgcttagaa gactcttcct tgcatggagt tcgggcctcc 180tccctgcacc taccacccta gtggccccaa gtcttaaggc tgaaggttaa tcctgtgtcc 240ttcagaagca aaggctgcaa ccgataccaa acagaggtgg ccagcgcggg ca 292103395DNAHomo sapiensmisc_feature(340)..(340)"n" refers to an undetermined base 103agagcttatc ccgcgagcac aagggagccg gggcctgggc cgccgtggga aggggctcct 60gccttccggg gacgcggtca gggaagtcca gccggggtgc tctctgcact gcgggtgccg 120ggctcggcag aggccaaccc ggcaaaacga gcaggatctc ccggccccac cctagtgggc 180tccgcctgcc ccaacaacca tcctgccatc ctccctgcga gacaggtgac tttcctctct 240gatgcggtgc atctgtcatc tgtctaacgg gcccattccc cagtgaaaca cccccaacca 300aagacacgaa ggggaaggcg caagcttcta ccaagctcan tttgcccatc tggtgcccac 360ctgcctngta tttggtgact tggaggatag gaagg 395

Claims

1. A diagnostic or prognostic assay for cancer, comprising: (a) obtaining a tissue sample from a test tissue; (b) performing a methylation assay on DNA derived from the tissue sample, wherein the methylation assay determines the methylation state of a CpG dinucleotide within a DNA sequence of the DNA, and wherein the DNA sequence is a sequence selected from the group consisting of sequences of SEQ ID NOS:1-103, sequences having a nucleotide sequence at least 90% identical to sequences of SEQ ID NOS:1-103, CpG island sequences associated with sequences of SEQ ID NOS:1-103, CpG island sequences associated with sequences having a nucleotide sequence at least 90% identical to sequences of SEQ ID NOS:1-103, and combinations thereof, wherein the CpG island sequence associated with the sequence of the particular SEQ ID NO is that contiguous sequence of genomic DNA that encompasses at least one nucleotide of the particular SEQ ID NO sequence, and satisfies the criteria of having both a frequency of CpG dinucleotides corresponding to an Observed/Expected Ratio >0.6, and a GC Content >0.5; and (c) determining a diagnosis or prognosis based, at least in part, upon the methylation state of the CpG dinucleotide within the DNA sequence.

2. The diagnostic or prognostic assay of claim 1 wherein the DNA sequence is a sequence selected from the group consisting of CpG island sequences associated with sequences of SEQ ID NOS:1-103, CpG island sequences associated with sequences having a nucleotide sequence at least 90% identical to sequences of SEQ ID NOS:1-103, and combinations thereof.

3. The diagnostic or prognostic assay of claim 2 wherein the DNA sequence is a sequence selected from the group consisting of CpG island sequences associated with sequences of SEQ ID NOS: 2, 4, 6, 7, 9-16, 19, 20, 22-33, 35-43, 48, 51-55, 59, 60, 64, 71, 76, 78-81, 84 and 87-90, and combinations thereof.

4. The diagnostic or prognostic assay of claim 1 wherein the methylation assay procedure is selected from the group consisting of MethyLight, MS-SNuPE, MSP MCA, COBRA, and combinations thereof.

5. The diagnostic or prognostic assay of claim 1 wherein the methylation state of the CpG dinucleotide within the DNA sequence is that of hypermethylation, hypomethylation or normal methylation.

6. The diagnostic or prognostic assay of claim 1 wherein the cancer is selected from the group consisting of bladder cancer, prostate cancer, colon cancer, lung cancer, renal cancer, leukemia, breast cancer, uterine cancer, astrocytoma, glioblastoma, and neuroblastoma.

7. A kit useful for the detection of a methylated CpG-containing nucleic acid comprising a carrier means containing one or more containers comprising: (a) a container containing a probe or primer which hybridizes to any region of a sequence selected from the group consisting of SEQ ID NOS:1-103, and sequences having a nucleotide sequence at least 90% identical to sequences of SEQ ID NOS:1-103; and (b) additional standard methylation assay reagents required to affect detection of methylated CpG-containing nucleic acid based, at least in part, on the probe or primer.

8. The kit of claim 7, wherein the additional standard methylation assay reagents are standard reagents for performing a methylation assay from the group consisting of MethyLight, MS-SNuPE, MSP MCA, COBRA, and combinations thereof.

9. The kit of claim 7, wherein the probe or primer comprises at least about 12 to 15 nucleotides of a sequence selected from the group consisting of SEQ ID NOS:1-103, and sequences having a nucleotide sequence at least 90% identical to sequences of SEQ ID NOS:1-103.

10. An isolated nucleic acid molecule comprising a methylated or unmethylated polynucleotide sequence selected from the group consisting of SEQ ID NO:1, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:18, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:32, SEQ ID NO:34, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:44, SEQ ID NO:51, SEQ ID NO:52, SEQ ID NO:62, SEQ ID NO:64, SEQ ID NO:65, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:74, SEQ ID NO:76, SEQ ID NO:82, SEQ ID NO:83, SEQ ID NO:84, SEQ ID NO:86, SEQ ID NO:90, SEQ ID NO:92, SEQ ID NO:97, and SEQ ID NO:100.

11. The nucleic acid of claim 10, wherein the nucleic acid is methylated.

12. The nucleic acid of claim 10, wherein the nucleic acid is unmethylated.