U.S. patent application number 12/856204 was filed with the patent office on 2011-01-27 for drought and heat tolerance in plants.
Invention is credited to Cory Christensen, Wuyi Wang, Dennis Yang.
Application Number | 20110023193 12/856204 |
Document ID | / |
Family ID | 40957515 |
Filed Date | 2011-01-27 |
United States Patent
Application |
20110023193 |
Kind Code |
A1 |
Christensen; Cory ; et
al. |
January 27, 2011 |
DROUGHT AND HEAT TOLERANCE IN PLANTS
Abstract
Methods and materials for modulating heat and/or drought
tolerance in plants are disclosed. For example, nucleic acids
encoding heat and/or drought-tolerance polypeptides are disclosed
as well as methods for using such nucleic acids to transform plant
cells. Also disclosed are plants having increased heat and/or
drought tolerance and plant products produced from plants having
increased heat and/or drought tolerance.
Inventors: |
Christensen; Cory;
(Sherwood, OR) ; Wang; Wuyi; (Newbury Park,
CA) ; Yang; Dennis; (Ballwin, MO) |
Correspondence
Address: |
FISH & RICHARDSON P.C.
P.O. BOX 1022
MINNEAPOLIS
MN
55440-1022
US
|
Family ID: |
40957515 |
Appl. No.: |
12/856204 |
Filed: |
August 13, 2010 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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PCT/US2009/034068 |
Feb 13, 2009 |
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12856204 |
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61029048 |
Feb 15, 2008 |
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Current U.S.
Class: |
800/306 ;
435/419; 800/295; 800/298; 800/312; 800/314; 800/320; 800/320.1;
800/320.2; 800/320.3; 800/322 |
Current CPC
Class: |
C12N 15/8271 20130101;
C12N 15/8273 20130101; C07K 14/415 20130101 |
Class at
Publication: |
800/306 ;
435/419; 800/295; 800/320.1; 800/320; 800/320.2; 800/314; 800/322;
800/298; 800/312; 800/320.3 |
International
Class: |
A01H 5/00 20060101
A01H005/00; C12N 5/10 20060101 C12N005/10 |
Claims
1.-2. (canceled)
3. The method of claim 22, wherein the polypeptide comprises a
sulfotransferase 1 domain having 70 percent or greater sequence
identity to the sulfotransferase 1 domain of SEQ ID NO: 590, 591,
592, 593, 594, 595, 596, 597, 598, 599, 600, 602, 603, 604, 605,
607, 608, 610, 612, 614, 615, 616, 617, 618, 619, 620, 621, 623,
625, 627, 629, 631, 632, 634, 635, 636, 638, 640, 642, 644, 646,
647, 650, 651, 653, 654, 655, 656, 658, 660, 661, 662, 663, 664,
665, 666, 667, 668, 669, 671, 673, 674, 675, 676, 678, 680, 682,
684, 685, 687, 688, 690, 691, 692, 694, 696, 698, 700, 702, 971,
974, 975, 978, 979, 980, 981, 983, 985, 987, 995, or 997.
4. The method of claim 22, wherein the polypeptide comprises an
AN1-like zinc finger domain having 70 percent or greater sequence
identity to the AN1-like zinc finger domain of SEQ ID NO: 742, 743,
744, 745, 757, 759, 761, 763, 767, 771, 796, 798, 799, 801, 805,
807, 808, 809, 810, 811, 812, 813, 815, 816, 817, 819, 823, 825,
832, 834, 836, 839, 846, 848, 857, 859, 863, 885, 892, 900, 912,
918, 939, 944, 953, 955, 957, 973, 977, 988, 989, 990, 991, 992,
993, 999, 1000, 1010, 1242, 748, 749, 797, 803, 821, 1189, 1191,
1193, 1195, 1197, 1199, 1201, 1203, 1205, 1207, 1209, 1210, 1212,
1214, 1215, 1216, 1218, 1219, 1221, 1222, 1223, 1225, 1226, 1228,
1230, 1231, 1232, 1234, 1236, 1238, 1240, 1244, 1246, 1248, 1250,
1252, 1254, 1256, 1258, 1296, 1298, 1300, 1302, or 1304 and an
A20-like zinc finger domain having 70 percent or greater sequence
identity to the A20-like zinc finger domain of SEQ ID NO: 742, 743,
744, 745, 757, 759, 761, 763, 767, 771, 796, 798, 799, 801, 805,
807, 808, 809, 810, 811, 812, 813, 815, 816, 817, 819, 823, 825,
832, 834, 836, 839, 846, 848, 857, 859, 863, 885, 892, 900, 912,
918, 939, 944, 953, 955, 957, 973, 977, 988, 989, 990, 991, 992,
993, 999, 1000, 1010, 1242, 748, 749, 797, 803, 821, 1189, 1191,
1193, 1195, 1197, 1199, 1201, 1203, 1205, 1207, 1209, 1210, 1212,
1214, 1215, 1216, 1218, 1219, 1221, 1222, 1223, 1225, 1226, 1228,
1230, 1231, 1232, 1234, 1236, 1238, 1240, 1244, 1246, 1248, 1250,
1252, 1254, 1256, 1258, 1296, 1298, 1300, 1302, or 1304.
5. The method of claim 22, wherein the polypeptide comprises a
short chain dehydrogenase domain having 70 percent or greater
sequence identity to the short chain dehydrogenase domain of SEQ ID
NO: 922, 924, 929, 931, 935, 937, 941, 943, 947, 949, 951, 952,
958, 959, 960, 961, 962, 963, 964, 1012, 1013, 1014, 1015, 1016,
1018, 1020, 1022, 1023, 1025, 1027, 1028, 1030, 1031, 1033, 1035,
1037, 1039, 1041, 1045, 1048, 1050, 1051, 1053, 1055, 1056, 1058,
1061, 1062, 1064, 1066, 1068, 1071, 1073, 1075, 1077, 1079, 1080,
1082, 1084, 1086, 1088, 1090, 1092, 1093, 1094, 1096, 1098, 1100,
1102, 1104, 1106, 1108, 1110, 1112, 1113, 1114, 1116, 1117, 1118,
1119, 1121, 1123, 1125, or 1292.
6. The method of claim 22, wherein the polypeptide comprises a
trehalose-phosphatase domain having 70 percent or greater sequence
identity to the trehalose-phosphatase domain of SEQ ID NO: 63, 73,
96, 127, 139, 179, 180, 209, 467, 559, 648, 726, 741, 765, 773,
781, 827, 829, 840, 841, 842, 843, 844, 845, 849, 850, 851, 852,
853, 854, 861, 865, 867, 896, 916, 926, 932, 933, 1043, 1046, 1059,
1069, 1129, 1130, 1131, 1132, 1133, 1134, 1135, 1136, 1137, 1138,
1140, 1142, 1144, 1146, 1148, 1150, 1151, 1152, 1153, 1154, 1156,
1157, 1158, 1159, 1160, 1161, 1162, 1164, 1166, 1168, 1170, 1172,
1174, 1175, 1177, 1179, 1181, 1182, 1184, 1186, or 1294.
7. The method of claim 22, wherein the polypeptide comprises a
synaptobrevin-related polypeptide having 70 percent or greater
sequence identity to the synaptobrevin-related polypeptide of SEQ
ID NO: 160, 162, 164, 166, 168, 170, 172, 174, 176, 178, 182, 184,
186, 188, 190, 192, 194, 196, 198, 200, 202, 204, 206, 208, 212,
214, 216, 218, 220, 222, 224, 225, 228, 229, 231, 233, 234, 235,
237, 239, 240, 242, 244, 246, 248, 250, 252, 253, 254, 256, 258,
260, 262, 264, 266, 268, 269, 271, 273, 275, 277, 279, 280, 282,
284, 286, 288, 289, 290, 291, 292, 293, 294, 295, 296, 297, 298,
299, 300, 301, 302, 303, 304, 305, 307, 309, 998, 1001, 1003, 1005,
1007, 1188, 1262, 1264, 1265, 1267, 1269, 1271, 1273, 1274, 1276,
1277, 1279, 1281, 1283, 1285, 1286, 1287, 1288, or 1290.
8. The method of claim 22, wherein the polypeptide comprises an
HSF-type DNA-binding domain having 70 percent or greater sequence
identity to the HSF-type DNA-binding domain of SEQ ID NO: 364, 366,
368, 369, 370, 371, 373, 375, 377, 379, 381, 383, 385, 387, 389,
390, 392, 394, 396, 398, 400, 401, 402, 404, 405, 407, 408, 410,
412, 413, 414, 416, 418, 419, 420, 421, 422, 423, 424, 426, 428,
430, 431, 432, 433, 434, 435, 436, 438, 440, 753, 754, 755, 756,
760, 768, 769, 774, 775, 777, 779, 783, 785, 789, 791, 793, 945,
1293, 311, 312, 314, 315, 316, 317, 319, 320, 321, 322, 323, 325,
326, 327, 329, 330, 332, 334, 335, 336, 337, 338, 339, 340, 341,
343, 344, 346, 348, 349, 350, 351, 353, 355, 356, 358, 360, 362,
876, 878, 880, 882, 886, 887, 888, 889, 890, 891, 893, 894, 897,
898, or 901.
9. The method of claim 22, wherein the polypeptide comprises a DnaJ
domain having 70 percent or greater sequence identity to the DnaJ
domain of SEQ ID NO: 60, 62, 64, 65, 66, 67, 68, 69, 70, 71, 74,
76, 78, 80, 82, 84, 86, 88, 90, 92, 94, 98, 100, 102, 104, 106,
108, 110, 112, 114, 116, 118, 120, 122, 124, 126, 130, 132, 134,
136, 138, 142, 144, 146, 148, 150, 152, 154, 156, 158, or 915.
10. The method of claim 22, wherein the polypeptide comprises a
KH.sub.--2 domain having 70 percent or greater sequence identity to
the KH.sub.--2 domain of SEQ ID NO: 1311, 1313, 1315, 1317, 1319,
1321, 1323, 1324, 1325, 1326, 1327, 1329, 1331, 1333, 1334, 1336,
1338, 1340, 1342, 1343, 1345, 1347, 1349, 1350, 1351, 1353, 1354,
1356, 1358, 1360, or 1361.
11.-20. (canceled)
21. A plant cell comprising an exogenous nucleic acid, said
exogenous nucleic acid comprising a regulatory region operably
linked to a nucleotide sequence encoding a polypeptide, wherein the
HMM bit score of the amino acid sequence of said polypeptide is
greater than about 20, said HMM based on the amino acid sequences
depicted in one of FIGS. 1-16, and wherein said plant has a
difference in drought tolerance or heat tolerance as compared to a
control plant that does not comprise said nucleic acid.
22. A plant cell comprising an exogenous nucleic acid said
exogenous nucleic acid comprising a regulatory region operably
linked to a nucleotide sequence encoding a polypeptide having 80
percent or greater sequence identity to an amino acid sequence
selected from the group consisting of SEQ ID NO: 2, 3, 4, 5, 6, 7,
8, 9, 10, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37,
39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 60, 62, 63, 64, 65, 66, 67,
68, 69, 70, 71, 73, 74, 76, 78, 80, 82, 84, 86, 88, 90, 92, 94, 96,
98, 100, 102, 104, 106, 108, 110, 112, 114, 116, 118, 120, 122,
124, 126, 127, 130, 132, 134, 136, 138, 139, 142, 144, 146, 148,
150, 152, 154, 156, 158, 160, 162, 164, 166, 168, 170, 172, 174,
176, 178, 179, 180, 182, 184, 186, 188, 190, 192, 194, 196, 198,
200, 202, 204, 206, 208, 209, 212, 214, 216, 218, 220, 222, 224,
225, 228, 229, 231, 233, 234, 235, 237, 239, 240, 242, 244, 246,
248, 250, 252, 253, 254, 256, 258, 260, 262, 264, 266, 268, 269,
271, 273, 275, 277, 279, 280, 282, 284, 286, 288, 289, 290, 291,
292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304,
305, 307, 309, 311, 312, 314, 315, 316, 317, 319, 320, 321, 322,
323, 325, 326, 327, 329, 330, 332, 334, 335, 336, 337, 338, 339,
340, 341, 343, 344, 346, 348, 349, 350, 351, 353, 355, 356, 358,
360, 362, 364, 366, 368, 369, 370, 371, 373, 375, 377, 379, 381,
383, 385, 387, 389, 390, 392, 394, 396, 398, 400, 401, 402, 404,
405, 407, 408, 410, 412, 413, 414, 416, 418, 419, 420, 421, 422,
423, 424, 426, 428, 430, 431, 432, 433, 434, 435, 436, 438, 440,
442, 443, 445, 447, 449, 451, 453, 455, 457, 459, 461, 463, 465,
467, 469, 470, 472, 474, 476, 478, 480, 481, 483, 485, 487, 489,
490, 491, 492, 494, 496, 498, 500, 502, 504, 506, 508, 510, 512,
514, 515, 516, 518, 519, 520, 522, 524, 526, 528, 530, 531, 533,
535, 536, 538, 540, 541, 543, 545, 547, 548, 550, 552, 553, 555,
557, 558, 559, 561, 562, 564, 566, 568, 569, 570, 572, 574, 576,
578, 580, 582, 584, 586, 588, 590, 591, 592, 593, 594, 595, 596,
597, 598, 599, 600, 602, 603, 604, 605, 607, 608, 610, 612, 614,
615, 616, 617, 618, 619, 620, 621, 623, 625, 627, 629, 631, 632,
634, 635, 636, 638, 640, 642, 644, 646, 647, 648, 650, 651, 653,
654, 655, 656, 658, 660, 661, 662, 663, 664, 665, 666, 667, 668,
669, 671, 673, 674, 675, 676, 678, 680, 682, 684, 685, 687, 688,
690, 691, 692, 694, 696, 698, 700, 702, 705, 706, 707, 708, 709,
711, 712, 714, 715, 717, 719, 721, 723, 725, 726, 727, 729, 731,
733, 734, 735, 737, 738, 740, 741, 742, 743, 744, 745, 746, 747,
748, 749, 751, 752, 753, 754, 755, 756, 757, 759, 760, 761, 763,
765, 767, 768, 769, 771, 773, 774, 775, 777, 779, 781, 783, 785,
787, 789, 791, 793, 795, 796, 797, 798, 799, 801, 803, 805, 807,
808, 809, 810, 811, 812, 813, 815, 816, 817, 819, 821, 823, 825,
827, 829, 832, 834, 836, 839, 840, 841, 842, 843, 844, 845, 846,
848, 849, 850, 851, 852, 853, 854, 856, 857, 859, 861, 863, 865,
867, 869, 870, 871, 872, 873, 874, 876, 878, 880, 882, 885, 886,
887, 888, 889, 890, 891, 892, 893, 894, 896, 897, 898, 900, 901,
902, 904, 906, 908, 910, 912, 913, 914, 915, 916, 918, 920, 922,
924, 926, 929, 931, 932, 933, 935, 937, 939, 941, 943, 944, 945,
947, 949, 951, 952, 953, 955, 957, 958, 959, 960, 961, 962, 963,
964, 966, 968, 969, 970, 971, 973, 974, 975, 977, 978, 979, 980,
981, 983, 985, 987, 988, 989, 990, 991, 992, 993, 995, 997, 998,
999, 1000, 1001, 1003, 1005, 1007, 1010, 1012, 1013, 1014, 1015,
1016, 1018, 1020, 1022, 1023, 1025, 1027, 1028, 1030, 1031, 1033,
1035, 1037, 1039, 1041, 1043, 1045, 1046, 1048, 1050, 1051, 1053,
1055, 1056, 1058, 1059, 1061, 1062, 1064, 1066, 1068, 1069, 1071,
1073, 1075, 1077, 1079, 1080, 1082, 1084, 1086, 1088, 1090, 1092,
1093, 1094, 1096, 1098, 1100, 1102, 1104, 1106, 1108, 1110, 1112,
1113, 1114, 1116, 1117, 1118, 1119, 1121, 1123, 1125, 1129, 1130,
1131, 1132, 1133, 1134, 1135, 1136, 1137, 1138, 1140, 1142, 1144,
1146, 1148, 1150, 1151, 1152, 1153, 1154, 1156, 1157, 1158, 1159,
1160, 1161, 1162, 1164, 1166, 1168, 1170, 1172, 1174, 1175, 1177,
1179, 1181, 1182, 1184, 1186, 1188, 1189, 1191, 1193, 1195, 1197,
1199, 1201, 1203, 1205, 1207, 1209, 1210, 1212, 1214, 1215, 1216,
1218, 1219, 1221, 1222, 1223, 1225, 1226, 1228, 1230, 1231, 1232,
1234, 1236, 1238, 1240, 1242, 1244, 1246, 1248, 1250, 1252, 1254,
1256, 1258, 1262, 1264, 1265, 1267, 1269, 1271, 1273, 1274, 1276,
1277, 1279, 1281, 1283, 1285, 1286, 1287, 1288, 1290, 1292, 1293,
1294, 1296, 1298, 1300, 1302, 1304, 1306, 1311, 1313, 1315, 1317,
1319, 1321, 1323, 1324, 1325, 1326, 1327, 1329, 1331, 1333, 1334,
1336, 1338, 1340, 1342, 1343, 1345, 1347, 1349, 1350, 1351, 1353,
1354, 1356, 1358, 1360, and 1361 wherein a plant produced from said
plant cell has a difference in heat tolerance or drought tolerance
as compared to a control plant that does not comprise said nucleic
acid.
23. A transgenic plant comprising the plant cell of claim 21,
wherein said polypeptide is selected from the group consisting of
SEQ ID NO:442, SEQ ID NO:489, SEQ ID NO:568, SEQ ID NO:590, SEQ ID
NO:705, SEQ ID NO:711, SEQ ID NO:742, SEQ ID NO:1012, SEQ ID
NO:1129, SEQ ID NO:60, SEQ ID NO:364, SEQ ID NO:311, SEQ ID NO:160,
SEQ ID NO:2, SEQ ID NO:749, SEQ ID NO:1311, and SEQ ID NO:57.
24. A plant cell comprising an exogenous nucleic acid said
exogenous nucleic acid comprising a regulatory region operably
linked to a nucleotide sequence having 80 percent or greater
sequence identity to a nucleotide sequence selected from the group
consisting of SEQ ID NO: 1, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30,
32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 59, 61, 72,
75, 77, 79, 81, 83, 85, 87, 89, 91, 93, 95, 97, 99, 101, 103, 105,
107, 109, 111, 113, 115, 117, 119, 121, 123, 125, 128, 129, 131,
133, 135, 137, 140, 141, 143, 145, 147, 149, 151, 153, 155, 157,
159, 161, 163, 165, 167, 169, 171, 173, 175, 177, 181, 183, 185,
187, 189, 191, 193, 195, 197, 199, 201, 203, 205, 207, 210, 211,
213, 215, 217, 219, 221, 223, 226, 227, 230, 232, 236, 238, 241,
243, 245, 247, 249, 251, 255, 257, 259, 261, 263, 265, 267, 270,
272, 274, 276, 278, 281, 283, 285, 287, 306, 308, 310, 313, 318,
324, 328, 331, 333, 342, 345, 347, 352, 354, 357, 359, 361, 363,
365, 367, 372, 374, 376, 378, 380, 382, 384, 386, 388, 391, 393,
395, 397, 399, 403, 406, 409, 411, 415, 417, 425, 427, 429, 437,
439, 441, 444, 446, 448, 450, 452, 454, 456, 458, 460, 462, 464,
466, 468, 471, 473, 475, 477, 479, 482, 484, 486, 488, 493, 495,
497, 499, 501, 503, 505, 507, 509, 511, 513, 517, 521, 523, 525,
527, 529, 532, 534, 537, 539, 542, 544, 546, 549, 551, 554, 556,
560, 563, 565, 567, 571, 573, 575, 577, 579, 581, 583, 585, 587,
589, 601, 606, 609, 611, 613, 622, 624, 626, 628, 630, 633, 637,
639, 641, 643, 645, 649, 652, 657, 659, 670, 672, 677, 679, 681,
683, 686, 689, 693, 695, 697, 699, 701, 703, 704, 710, 713, 716,
718, 720, 722, 724, 728, 730, 732, 736, 739, 750, 758, 762, 764,
766, 770, 772, 776, 778, 780, 782, 784, 786, 788, 790, 792, 794,
800, 802, 804, 806, 814, 818, 820, 822, 824, 826, 828, 830, 831,
833, 835, 837, 838, 847, 855, 858, 860, 862, 864, 866, 868, 875,
877, 879, 881, 883, 884, 895, 899, 903, 905, 907, 909, 911, 917,
919, 921, 923, 925, 927, 928, 930, 934, 936, 938, 940, 942, 946,
948, 950, 954, 956, 965, 967, 972, 976, 982, 984, 986, 994, 996,
1002, 1004, 1006, 1008, 1009, 1011, 1017, 1019, 1021, 1024, 1026,
1029, 1032, 1034, 1036, 1038, 1040, 1042, 1044, 1047, 1049, 1052,
1054, 1057, 1060, 1063, 1065, 1067, 1070, 1072, 1074, 1076, 1078,
1081, 1083, 1085, 1087, 1089, 1091, 1095, 1097, 1099, 1101, 1103,
1105, 1107, 1109, 1111, 1115, 1120, 1122, 1124, 1126, 1127, 1128,
1139, 1141, 1143, 1145, 1147, 1149, 1155, 1163, 1165, 1167, 1169,
1171, 1173, 1176, 1178, 1180, 1183, 1185, 1187, 1190, 1192, 1194,
1196, 1198, 1200, 1202, 1204, 1206, 1208, 1211, 1213, 1217, 1220,
1224, 1227, 1229, 1233, 1235, 1237, 1239, 1241, 1243, 1245, 1247,
1249, 1251, 1253, 1255, 1257, 1259, 1260, 1261, 1263, 1266, 1268,
1270, 1272, 1275, 1278, 1280, 1282, 1284, 1289, 1291, 1295, 1297,
1299, 1301, 1303, 1305, 1312, 1314, 1316, 1318, 1320, 1322, 1328,
1330, 1332, 1335, 1337, 1339, 1341, 1344, 1346, 1348, 1352, 1355,
1357, 1359, and 1362, or a fragment thereof, wherein a plant
produced from said plant cell has a difference in drought tolerance
or heat tolerance as compared to a control plant that does not
comprise said nucleic acid.
25. A transgenic plant comprising the plant cell of claim 24.
26. The transgenic plant of claim 25, wherein said plant is a
member of a species selected from the group consisting of Panicum
virgatum (switchgrass), Sorghum bicolor (sorghum, sudangrass),
Miscanthus giganteus (miscanthus), Saccharum sp. (energycane),
Populus balsamifera (poplar), Zea mays (corn), Glycine max
(soybean), Brassica napus (canola), Triticum aestivum (wheat),
Gossypium hirsutum (cotton), Oryza sativa (rice), Helianthus annuus
(sunflower), Medicago sativa (alfalfa), Beta vulgaris (sugarbeet),
and Pennisetum glaucum (pearl millet).
27.-33. (canceled)
34. A transgenic plant comprising the plant cell of claim 21.
35. A transgenic plant comprising the plant cell of claim 22.
36. The transgenic plant of claim 34, wherein said plant is a
member of a species selected from the group consisting of Panicum
virgatum (switchgrass), Sorghum bicolor (sorghum, sudangrass),
Miscanthus giganteus (miscanthus), Saccharum sp. (energycane),
Populus balsamifera (poplar), Zea mays (corn), Glycine max
(soybean), Brassica napus (canola), Triticum aestivum (wheat),
Gossypium hirsutum (cotton), Oryza sativa (rice), Helianthus annuus
(sunflower), Medicago sativa (alfalfa), Beta vulgaris (sugarbeet),
and Pennisetum glaucum (pearl millet).
37. The transgenic plant of claim 34, wherein said plant is a
member of a species selected from the group consisting of Panicum
virgatum (switchgrass), Sorghum bicolor (sorghum, sudangrass),
Miscanthus giganteus (miscanthus), Saccharum sp. (energycane),
Populus balsamifera (poplar), Zea mays (corn), Glycine max
(soybean), Brassica napus (canola), Triticum aestivum (wheat),
Gossypium hirsutum (cotton), Oryza sativa (rice), Helianthus annuus
(sunflower), Medicago sativa (alfalfa), Beta vulgaris (sugarbeet),
and Pennisetum glaucum (pearl millet).
Description
INCORPORATION-BY-REFERENCE OF SEQUENCE LISTING OR TABLE
[0001] The material in the accompanying sequence listing is hereby
incorporated by reference into this application. The accompanying
file, named 11696-248WO1_sequence_listing was created on Feb. 5,
2009 and is 3090 KB. The file can be accessed using Microsoft Word
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TECHNICAL FIELD
[0002] This document relates to methods and materials involved in
modulating drought and/or heat tolerance in plants. For example,
this document provides plants having increased drought and/or heat
tolerance as well as materials and methods for making plants having
increased drought and/or heat tolerance.
BACKGROUND
[0003] Due to their sessile nature, plants are constantly under the
threat of temperature stress when they are subjected to a wide
range of temperature variation in different habitats and climates
during growing seasons and even diurnally. Most economically
valuable plants, including those used in agriculture, horticulture,
forestry, biomass for bioconversion, and other industries (e.g.,
the paper industry or pharmaceutical/chemical industries where
plants are used as production factories for proteins or other
compounds) are exposed to higher than optimal temperatures, or heat
stress, during some stages of their life cycle from seed
germination to seed maturation (Maestri et al. Plant Mol. Biol.
48:667-681 (2002)). Heat stress is one of the most common stresses
in crop production. Under heat stress, plants can succumb to a
variety of physiological and developmental damages, including
dehydration due to increased transpiration, impairment of
photosynthetic carbon assimilation, inhibition of translocation of
assimilates, increased respiration, reduced organ size due to a
decrease in the duration of developmental phases, disruption of
seed development, and a reduction in fertility (Berry and Bjorkman,
Ann. Rev. Plant Physiol. 31:491-543 (1980); Cheikh and Jones, Plant
Physiol. 106:45-51 (1994)). Thus, exposure to heat stress often
results in reduced yield and overall decreased crop quality
(Maestri et al. Plant Mol. Biol. 48:667-681 (2002)).
[0004] In the field, heat stress is often associated with other
stresses, such as drought and high light, which presents even
greater challenge to plants. Plants exposed to low water or drought
conditions typically have low yields of plant material, seeds,
fruit and other edible products. Some areas of the world
consistently have very low rainfall and therefore have problems
growing sufficient food crops for their population.
[0005] Thus, there is a need for methods of increasing drought
and/or heat tolerance in plants.
SUMMARY
[0006] This document provides methods and materials related to
plants having increased heat and/or drought tolerance. For example,
this document provides transgenic plants and plant cells having
increased heat and/or drought tolerance, nucleic acids used to
generate transgenic plants and plant cells having increased heat
and/or drought tolerance, and methods for making plants and plant
cells having increased heat and/or drought tolerance. Such plants
having increased heat and/or drought tolerance may be useful to
produce biomass which may be converted to a liquid fuel or other
chemicals and/or to produce crops with increased yield and/or
quality.
[0007] Methods of producing a plant are provided herein. In one
aspect, a method comprises growing a plant cell comprising an
exogenous nucleic acid. The exogenous nucleic acid comprises a
regulatory region operably linked to a nucleotide sequence encoding
a polypeptide. The Hidden Markov Model (HMM) bit score of the amino
acid sequence of the polypeptide is greater than about 20, using an
HMM generated from the amino acid sequences depicted in one of
FIGS. 1-16. The plant has a difference in drought tolerance or heat
tolerance as compared to a control plant that does not comprise the
exogenous nucleic acid.
[0008] In another aspect, a method comprises growing a plant cell
comprising an exogenous nucleic acid. The exogenous nucleic acid
comprises a regulatory region operably linked to a nucleotide
sequence encoding a polypeptide having 80 percent or greater
sequence identity to an amino acid sequence set forth in SEQ ID NO:
2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29,
31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 60, 62, 63,
64, 65, 66, 67, 68, 69, 70, 71, 73, 74, 76, 78, 80, 82, 84, 86, 88,
90, 92, 94, 96, 98, 100, 102, 104, 106, 108, 110, 112, 114, 116,
118, 120, 122, 124, 126, 127, 130, 132, 134, 136, 138, 139, 142,
144, 146, 148, 150, 152, 154, 156, 158, 160, 162, 164, 166, 168,
170, 172, 174, 176, 178, 179, 180, 182, 184, 186, 188, 190, 192,
194, 196, 198, 200, 202, 204, 206, 208, 209, 212, 214, 216, 218,
220, 222, 224, 225, 228, 229, 231, 233, 234, 235, 237, 239, 240,
242, 244, 246, 248, 250, 252, 253, 254, 256, 258, 260, 262, 264,
266, 268, 269, 271, 273, 275, 277, 279, 280, 282, 284, 286, 288,
289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301,
302, 303, 304, 305, 307, 309, 311, 312, 314, 315, 316, 317, 319,
320, 321, 322, 323, 325, 326, 327, 329, 330, 332, 334, 335, 336,
337, 338, 339, 340, 341, 343, 344, 346, 348, 349, 350, 351, 353,
355, 356, 358, 360, 362, 364, 366, 368, 369, 370, 371, 373, 375,
377, 379, 381, 383, 385, 387, 389, 390, 392, 394, 396, 398, 400,
401, 402, 404, 405, 407, 408, 410, 412, 413, 414, 416, 418, 419,
420, 421, 422, 423, 424, 426, 428, 430, 431, 432, 433, 434, 435,
436, 438, 440, 442, 443, 445, 447, 449, 451, 453, 455, 457, 459,
461, 463, 465, 467, 469, 470, 472, 474, 476, 478, 480, 481, 483,
485, 487, 489, 490, 491, 492, 494, 496, 498, 500, 502, 504, 506,
508, 510, 512, 514, 515, 516, 518, 519, 520, 522, 524, 526, 528,
530, 531, 533, 535, 536, 538, 540, 541, 543, 545, 547, 548, 550,
552, 553, 555, 557, 558, 559, 561, 562, 564, 566, 568, 569, 570,
572, 574, 576, 578, 580, 582, 584, 586, 588, 590, 591, 592, 593,
594, 595, 596, 597, 598, 599, 600, 602, 603, 604, 605, 607, 608,
610, 612, 614, 615, 616, 617, 618, 619, 620, 621, 623, 625, 627,
629, 631, 632, 634, 635, 636, 638, 640, 642, 644, 646, 647, 648,
650, 651, 653, 654, 655, 656, 658, 660, 661, 662, 663, 664, 665,
666, 667, 668, 669, 671, 673, 674, 675, 676, 678, 680, 682, 684,
685, 687, 688, 690, 691, 692, 694, 696, 698, 700, 702, 705, 706,
707, 708, 709, 711, 712, 714, 715, 717, 719, 721, 723, 725, 726,
727, 729, 731, 733, 734, 735, 737, 738, 740, 741, 742, 743, 744,
745, 746, 747, 748, 749, 751, 752, 753, 754, 755, 756, 757, 759,
760, 761, 763, 765, 767, 768, 769, 771, 773, 774, 775, 777, 779,
781, 783, 785, 787, 789, 791, 793, 795, 796, 797, 798, 799, 801,
803, 805, 807, 808, 809, 810, 811, 812, 813, 815, 816, 817, 819,
821, 823, 825, 827, 829, 832, 834, 836, 839, 840, 841, 842, 843,
844, 845, 846, 848, 849, 850, 851, 852, 853, 854, 856, 857, 859,
861, 863, 865, 867, 869, 870, 871, 872, 873, 874, 876, 878, 880,
882, 885, 886, 887, 888, 889, 890, 891, 892, 893, 894, 896, 897,
898, 900, 901, 902, 904, 906, 908, 910, 912, 913, 914, 915, 916,
918, 920, 922, 924, 926, 929, 931, 932, 933, 935, 937, 939, 941,
943, 944, 945, 947, 949, 951, 952, 953, 955, 957, 958, 959, 960,
961, 962, 963, 964, 966, 968, 969, 970, 971, 973, 974, 975, 977,
978, 979, 980, 981, 983, 985, 987, 988, 989, 990, 991, 992, 993,
995, 997, 998, 999, 1000, 1001, 1003, 1005, 1007, 1010, 1012, 1013,
1014, 1015, 1016, 1018, 1020, 1022, 1023, 1025, 1027, 1028, 1030,
1031, 1033, 1035, 1037, 1039, 1041, 1043, 1045, 1046, 1048, 1050,
1051, 1053, 1055, 1056, 1058, 1059, 1061, 1062, 1064, 1066, 1068,
1069, 1071, 1073, 1075, 1077, 1079, 1080, 1082, 1084, 1086, 1088,
1090, 1092, 1093, 1094, 1096, 1098, 1100, 1102, 1104, 1106, 1108,
1110, 1112, 1113, 1114, 1116, 1117, 1118, 1119, 1121, 1123, 1125,
1129, 1130, 1131, 1132, 1133, 1134, 1135, 1136, 1137, 1138, 1140,
1142, 1144, 1146, 1148, 1150, 1151, 1152, 1153, 1154, 1156, 1157,
1158, 1159, 1160, 1161, 1162, 1164, 1166, 1168, 1170, 1172, 1174,
1175, 1177, 1179, 1181, 1182, 1184, 1186, 1188, 1189, 1191, 1193,
1195, 1197, 1199, 1201, 1203, 1205, 1207, 1209, 1210, 1212, 1214,
1215, 1216, 1218, 1219, 1221, 1222, 1223, 1225, 1226, 1228, 1230,
1231, 1232, 1234, 1236, 1238, 1240, 1242, 1244, 1246, 1248, 1250,
1252, 1254, 1256, 1258, 1262, 1264, 1265, 1267, 1269, 1271, 1273,
1274, 1276, 1277, 1279, 1281, 1283, 1285, 1286, 1287, 1288, 1290,
1292, 1293, 1294, 1296, 1298, 1300, 1302, 1304, 1306, 1311, 1313,
1315, 1317, 1319, 1321, 1323, 1324, 1325, 1326, 1327, 1329, 1331,
1333, 1334, 1336, 1338, 1340, 1342, 1343, 1345, 1347, 1349, 1350,
1351, 1353, 1354, 1356, 1358, 1360, or 1361. A plant produced from
the plant cell has a difference in drought tolerance or heat
tolerance compared to a control plant that does not comprise the
exogenous nucleic acid.
[0009] In another aspect, the polypeptide can comprise a
sulfotransferase 1 domain having 70 percent or greater sequence
identity to the sulfotransferase 1 domain of SEQ ID NO:590 or
orthologs thereof such as those identified in the sequence listing;
an AN1-like zinc finger domain having 70 percent or greater
sequence identity to the AN1-like zinc finger domain of SEQ ID
NO:742 or SEQ ID NO:749 or orthologs thereof such as those
identified in the sequence listing and an A20-like zinc finger
domain having 70 percent or greater sequence identity to the
A20-like zinc finger domain of SEQ ID NO:742 or SEQ ID NO:749 or
orthologs thereof such as those identified in the sequence listing;
a short chain dehydrogenase domain having 70 percent or greater
sequence identity to the short chain dehydrogenase domain of SEQ ID
NO:1012 or orthologs thereof such as those identified in the
sequence listing; a trehalose-phosphatase domain having 70 percent
or greater sequence identity to the trehalose-phosphatase domain of
or SEQ ID NO:1129 or orthologs thereof such as those identified in
the sequence listing; a synaptobrevin-related polypeptide having 70
percent or greater sequence identity to the synaptobrevin-related
polypeptide of SEQ ID NO:160 or orthologs thereof such as those
identified in the sequence listing; an HSF-type DNA-binding domain
having 70 percent or greater sequence identity to the HSF-type
DNA-binding domain of SEQ ID NO:311 or SEQ ID NO:364 or orthologs
thereof such as those identified in the sequence listing; a DnaJ
domain having 70 percent or greater sequence identity to the DnaJ
domain of SEQ ID NO:60 or orthologs thereof such as those
identified in the sequence listing; or a KH.sub.--2 domain having
70 percent or greater sequence identity to the KH.sub.--2 domain of
SEQ ID NO:1311 or orthologs thereof such as those identified in the
sequence listing.
[0010] In another aspect, a method comprises growing a plant cell
comprising an exogenous nucleic acid. The exogenous nucleic acid
comprises a regulatory region operably linked to a nucleotide
sequence having 80 percent or greater sequence identity to a
nucleotide sequence, or a fragment thereof, set forth in SEQ ID NO:
1, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42,
44, 46, 48, 50, 52, 54, 56, 58, 59, 61, 72, 75, 77, 79, 81, 83, 85,
87, 89, 91, 93, 95, 97, 99, 101, 103, 105, 107, 109, 111, 113, 115,
117, 119, 121, 123, 125, 128, 129, 131, 133, 135, 137, 140, 141,
143, 145, 147, 149, 151, 153, 155, 157, 159, 161, 163, 165, 167,
169, 171, 173, 175, 177, 181, 183, 185, 187, 189, 191, 193, 195,
197, 199, 201, 203, 205, 207, 210, 211, 213, 215, 217, 219, 221,
223, 226, 227, 230, 232, 236, 238, 241, 243, 245, 247, 249, 251,
255, 257, 259, 261, 263, 265, 267, 270, 272, 274, 276, 278, 281,
283, 285, 287, 306, 308, 310, 313, 318, 324, 328, 331, 333, 342,
345, 347, 352, 354, 357, 359, 361, 363, 365, 367, 372, 374, 376,
378, 380, 382, 384, 386, 388, 391, 393, 395, 397, 399, 403, 406,
409, 411, 415, 417, 425, 427, 429, 437, 439, 441, 444, 446, 448,
450, 452, 454, 456, 458, 460, 462, 464, 466, 468, 471, 473, 475,
477, 479, 482, 484, 486, 488, 493, 495, 497, 499, 501, 503, 505,
507, 509, 511, 513, 517, 521, 523, 525, 527, 529, 532, 534, 537,
539, 542, 544, 546, 549, 551, 554, 556, 560, 563, 565, 567, 571,
573, 575, 577, 579, 581, 583, 585, 587, 589, 601, 606, 609, 611,
613, 622, 624, 626, 628, 630, 633, 637, 639, 641, 643, 645, 649,
652, 657, 659, 670, 672, 677, 679, 681, 683, 686, 689, 693, 695,
697, 699, 701, 703, 704, 710, 713, 716, 718, 720, 722, 724, 728,
730, 732, 736, 739, 750, 758, 762, 764, 766, 770, 772, 776, 778,
780, 782, 784, 786, 788, 790, 792, 794, 800, 802, 804, 806, 814,
818, 820, 822, 824, 826, 828, 830, 831, 833, 835, 837, 838, 847,
855, 858, 860, 862, 864, 866, 868, 875, 877, 879, 881, 883, 884,
895, 899, 903, 905, 907, 909, 911, 917, 919, 921, 923, 925, 927,
928, 930, 934, 936, 938, 940, 942, 946, 948, 950, 954, 956, 965,
967, 972, 976, 982, 984, 986, 994, 996, 1002, 1004, 1006, 1008,
1009, 1011, 1017, 1019, 1021, 1024, 1026, 1029, 1032, 1034, 1036,
1038, 1040, 1042, 1044, 1047, 1049, 1052, 1054, 1057, 1060, 1063,
1065, 1067, 1070, 1072, 1074, 1076, 1078, 1081, 1083, 1085, 1087,
1089, 1091, 1095, 1097, 1099, 1101, 1103, 1105, 1107, 1109, 1111,
1115, 1120, 1122, 1124, 1126, 1127, 1128, 1139, 1141, 1143, 1145,
1147, 1149, 1155, 1163, 1165, 1167, 1169, 1171, 1173, 1176, 1178,
1180, 1183, 1185, 1187, 1190, 1192, 1194, 1196, 1198, 1200, 1202,
1204, 1206, 1208, 1211, 1213, 1217, 1220, 1224, 1227, 1229, 1233,
1235, 1237, 1239, 1241, 1243, 1245, 1247, 1249, 1251, 1253, 1255,
1257, 1259, 1260, 1261, 1263, 1266, 1268, 1270, 1272, 1275, 1278,
1280, 1282, 1284, 1289, 1291, 1295, 1297, 1299, 1301, 1303, 1305,
1312, 1314, 1316, 1318, 1320, 1322, 1328, 1330, 1332, 1335, 1337,
1339, 1341, 1344, 1346, 1348, 1352, 1355, 1357, 1359 or 1362, or a
fragment thereof. A plant produced from the plant cell has a
difference in drought tolerance or heat tolerance as compared to a
control plant that does not comprise the exogenous nucleic
acid.
[0011] Methods of modulating drought in a plant are provided
herein. In one aspect, a method comprises introducing into a plant
cell an exogenous nucleic acid, that comprises a regulatory region
operably linked to a nucleotide sequence encoding a polypeptide.
The HMM bit score of the amino acid sequence of the polypeptide is
greater than about 20, using an HMM generated from the amino acid
sequences depicted in one of FIGS. 1-16. A plant produced from the
plant cell has a difference in drought tolerance as compared to a
control plant that does not comprise the exogenous nucleic
acid.
[0012] In another aspect, a method of modulating drought tolerance
comprises introducing into a plant cell an exogenous nucleic acid
that comprises a regulatory region operably linked to a nucleotide
sequence encoding a polypeptide having 80 percent or greater
sequence identity to an amino acid sequence set forth in SEQ ID NO:
442, 443, 445, 447, 449, 451, 453, 455, 457, 459, 461, 463, 465,
469, 470, 472, 474, 476, 478, 480, 481, 483, 485, 487, 966, 968,
969, 970, 902, 904, 906, 908, 910, 913, 914, 920, 748, 749, 797,
803, 821, 1189, 1191, 1193, 1195, 1197, 1199, 1201, 1203, 1205,
1207, 1209, 1210, 1212, 1214, 1215, 1216, 1218, 1219, 1221, 1222,
1223, 1225, 1226, 1228, 1230, 1231, 1232, 1234, 1236, 1238, 1240,
1244, 1246, 1248, 1250, 1252, 1254, 1256, 1258, 1296, 1298, 1300,
1302, 1304, 489, 490, 491, 492, 494, 496, 498, 500, 502, 504, 506,
508, 510, 512, 514, 515, 516, 518, 519, 520, 522, 524, 526, 528,
530, 531, 533, 535, 536, 538, 540, 541, 543, 545, 547, 548, 550,
552, 553, 555, 557, 558, 561, 562, 564, 566, 856, 869, 870, 871,
872, 873, 874, 568, 569, 570, 572, 574, 576, 578, 580, 582, 584,
586, 588, 795, 590, 591, 592, 593, 594, 595, 596, 597, 598, 599,
600, 602, 603, 604, 605, 607, 608, 610, 612, 614, 615, 616, 617,
618, 619, 620, 621, 623, 625, 627, 629, 631, 632, 634, 635, 636,
638, 640, 642, 644, 646, 647, 650, 651, 653, 654, 655, 656, 658,
660, 661, 662, 663, 664, 665, 666, 667, 668, 669, 671, 673, 674,
675, 676, 678, 680, 682, 684, 685, 687, 688, 690, 691, 692, 694,
696, 698, 700, 702, 971, 974, 975, 978, 979, 980, 981, 983, 985,
987, 995, 997, 705, 706, 707, 708, 709, 711, 712, 714, 715, 717,
719, 721, 723, 725, 727, 729, 731, 733, 734, 735, 737, 738, 740,
746, 747, 751, 752, 787, 742, 743, 744, 745, 757, 759, 761, 763,
767, 771, 796, 798, 799, 801, 805, 807, 808, 809, 810, 811, 812,
813, 815, 816, 817, 819, 823, 825, 832, 834, 836, 839, 846, 848,
857, 859, 863, 885, 892, 900, 912, 918, 939, 944, 953, 955, 957,
973, 977, 988, 989, 990, 991, 992, 993, 999, 1000, 1010, 1242, 922,
924, 929, 931, 935, 937, 941, 943, 947, 949, 951, 952, 958, 959,
960, 961, 962, 963, 964, 1012, 1013, 1014, 1015, 1016, 1018, 1020,
1022, 1023, 1025, 1027, 1028, 1030, 1031, 1033, 1035, 1037, 1039,
1041, 1045, 1048, 1050, 1051, 1053, 1055, 1056, 1058, 1061, 1062,
1064, 1066, 1068, 1071, 1073, 1075, 1077, 1079, 1080, 1082, 1084,
1086, 1088, 1090, 1092, 1093, 1094, 1096, 1098, 1100, 1102, 1104,
1106, 1108, 1110, 1112, 1113, 1114, 1116, 1117, 1118, 1119, 1121,
1123, 1125, 1292, 63, 73, 96, 127, 139, 179, 180, 209, 467, 559,
648, 726, 741, 765, 773, 781, 827, 829, 840, 841, 842, 843, 844,
845, 849, 850, 851, 852, 853, 854, 861, 865, 867, 896, 916, 926,
932, 933, 1043, 1046, 1059, 1069, 1129, 1130, 1131, 1132, 1133,
1134, 1135, 1136, 1137, 1138, 1140, 1142, 1144, 1146, 1148, 1150,
1151, 1152, 1153, 1154, 1156, 1157, 1158, 1159, 1160, 1161, 1162,
1164, 1166, 1168, 1170, 1172, 1174, 1175, 1177, 1179, 1181, 1182,
1184, 1186, 1294, 1311, 1313, 1315, 1317, 1319, 1321, 1323, 1324,
1325, 1326, 1327, 1329, 1331, 1333, 1334, 1336, 1338, 1340, 1342,
1343, 1345, 1347, 1349, 1350, 1351, 1353, 1354, 1356, 1358, 1360,
or 1361. A plant produced from the plant cell has a difference in
drought tolerance as compared to a control plant that does not
comprise the exogenous nucleic acid.
[0013] In another aspect, a method of modulating drought tolerance
comprises introducing into a plant cell an exogenous nucleic acid,
that comprises a regulatory region operably linked to a nucleotide
sequence having 80 percent or greater sequence identity to a
nucleotide sequence set forth in SEQ ID NO: 923, 994, 905, 921,
804, 837, 828, 930, 996, 928, 868, 645, 641, 695, 505, 1067, 501,
466, 718, 860, 503, 624, 628, 630, 639, 686, 806, 479, 764, 1060,
1052, 1120, 1105, 546, 780, 460, 814, 800, 499, 643, 670, 679,
1253, 927, 716, 1081, 128, 585, 581, 864, 1049, 1107, 523, 1076,
652, 1095, 1099, 1101, 497, 830, 507, 622, 140, 772, 532, 542, 549,
226, 1149, 210, 587, 583, 866, 527, 1097, 1103, 1141, 1019, 486,
1229, 1249, 739, 697, 701, 1011, 563, 1122, 1243, 1208, 1247, 1009,
1124, 699, 1206, 72, 565, 1245, 1021, 1008, 942, 948, 818, 794,
967, 986, 786, 936, 940, 984, 855, 946, 982, 1291, 919, 822, 824,
907, 750, 909, 934, 903, 826, 95, 965, 950, 611, 575, 1190, 484,
454, 1036, 722, 444, 456, 1192, 1034, 720, 831, 911, 838, 917, 468,
1257, 659, 1173, 1139, 1176, 938, 1063, 1032, 862, 758, 450, 509,
554, 1259, 458, 1217, 972, 477, 462, 954, 1163, 956, 525, 556,
1145, 1165, 1065, 677, 609, 762, 537, 1169, 884, 1198, 1085, 1109,
1211, 1029, 689, 1070, 1233, 770, 1047, 1194, 551, 446, 633, 1167,
1078, 1213, 730, 448, 766, 895, 511, 925, 495, 1239, 1235, 1057,
1074, 1202, 544, 1083, 1024, 1220, 1224, 1200, 613, 683, 672, 976,
517, 1227, 471, 1255, 1237, 473, 728, 521, 464, 1241, 1155, 681,
736, 539, 1017, 513, 835, 657, 534, 1044, 649, 1026, 560, 1171,
579, 1038, 529, 1091, 1147, 637, 452, 1072, 1180, 833, 1040, 1089,
482, 493, 732, 1115, 626, 899, 606, 1178, 1087, 1042, 1111, 577,
1251, 475, 858, 724, 1183, 693, 1143, 1185, 1054, 1187, 571, 713,
847, 1196, 573, 1204, 601, 441, 488, 567, 589, 703, 704, 710, 1260,
1126, 1127, 1128, 1295, 802, 1303, 1297, 1301, 1299, 820, 1312,
1314, 1316, 1318, 1320, 1322, 1328, 1330, 1332, 1335, 1337, 1339,
1341, 1344, 1346, 1348, 1352, 1355, 1357, 1359 or 1362, or a
fragment thereof. A plant produced from the plant cell has a
difference in drought tolerance as compared to a control plant that
does not comprise the exogenous nucleic acid.
[0014] Methods of modulating heat in a plant are provided herein.
In one aspect, a method comprises introducing into a plant cell an
exogenous nucleic acid, that comprises a regulatory region operably
linked to a nucleotide sequence encoding a polypeptide. The HMM bit
score of the amino acid sequence of the polypeptide is greater than
about 20, using an HMM generated from the amino acid sequences
depicted in one of FIGS. 10-14. A plant produced from the plant
cell has a difference in heat tolerance as compared to a control
plant that does not comprise the exogenous nucleic acid.
[0015] In another aspect, a method of modulating heat tolerance
comprises introducing into a plant cell an exogenous nucleic acid
that comprises a regulatory region operably linked to a nucleotide
sequence encoding a polypeptide having 80 percent or greater
sequence identity to an amino acid sequence set forth in SEQ ID NO:
60, 62, 64, 65, 66, 67, 68, 69, 70, 71, 74, 76, 78, 80, 82, 84, 86,
88, 90, 92, 94, 98, 100, 102, 104, 106, 108, 110, 112, 114, 116,
118, 120, 122, 124, 126, 130, 132, 134, 136, 138, 142, 144, 146,
148, 150, 152, 154, 156, 158, 915, 364, 366, 368, 369, 370, 371,
373, 375, 377, 379, 381, 383, 385, 387, 389, 390, 392, 394, 396,
398, 400, 401, 402, 404, 405, 407, 408, 410, 412, 413, 414, 416,
418, 419, 420, 421, 422, 423, 424, 426, 428, 430, 431, 432, 433,
434, 435, 436, 438, 440, 753, 754, 755, 756, 760, 768, 769, 774,
775, 777, 779, 783, 785, 789, 791, 793, 945, 1293, 311, 312, 314,
315, 316, 317, 319, 320, 321, 322, 323, 325, 326, 327, 329, 330,
332, 334, 335, 336, 337, 338, 339, 340, 341, 343, 344, 346, 348,
349, 350, 351, 353, 355, 356, 358, 360, 362, 876, 878, 880, 882,
886, 887, 888, 889, 890, 891, 893, 894, 897, 898, 901, 160, 162,
164, 166, 168, 170, 172, 174, 176, 178, 182, 184, 186, 188, 190,
192, 194, 196, 198, 200, 202, 204, 206, 208, 212, 214, 216, 218,
220, 222, 224, 225, 228, 229, 231, 233, 234, 235, 237, 239, 240,
242, 244, 246, 248, 250, 252, 253, 254, 256, 258, 260, 262, 264,
266, 268, 269, 271, 273, 275, 277, 279, 280, 282, 284, 286, 288,
289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301,
302, 303, 304, 305, 307, 309, 998, 1001, 1003, 1005, 1007, 1188,
1262, 1264, 1265, 1267, 1269, 1271, 1273, 1274, 1276, 1277, 1279,
1281, 1283, 1285, 1286, 1287, 1288, 1290, 2, 3, 4, 5, 6, 7, 8, 9,
10, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41,
43, 45, 47, 49, 51, 53, 55, 902, 904, 906, 908, 910, 913, 914, or
920. A plant produced from the plant cell has a difference in heat
tolerance as compared to a control plant that does not comprise the
exogenous nucleic acid.
[0016] In another aspect, a method of modulating heat tolerance
comprises introducing into a plant cell an exogenous nucleic acid,
that comprises a regulatory region operably linked to a nucleotide
sequence having 80 percent or greater sequence identity to a
nucleotide sequence set forth in SEQ ID NO: 877, 1263, 790, 1261,
875, 792, 201, 354, 85, 77, 75, 415, 46, 207, 333, 203, 382, 83,
411, 81, 425, 406, 189, 93, 223, 44, 345, 79, 199, 417, 42, 219,
352, 185, 193, 89, 205, 386, 365, 91, 211, 195, 217, 429, 213, 187,
221, 359, 40, 197, 357, 331, 215, 183, 191, 367, 318, 87, 378, 48,
157, 155, 306, 361, 439, 52, 50, 437, 308, 54, 153, 1002, 782,
1266, 1268, 1275, 881, 784, 1278, 1280, 1004, 778, 879, 1282, 1006,
1289, 1284, 1270, 788, 883, 1272, 776, 135, 259, 109, 245, 121, 28,
115, 261, 151, 145, 328, 313, 24, 20, 391, 99, 409, 272, 1, 342,
347, 374, 131, 372, 111, 103, 107, 388, 125, 117, 18, 380, 123,
129, 26, 399, 143, 283, 30, 167, 403, 287, 137, 281, 169, 161, 14,
165, 181, 12, 38, 163, 173, 324, 177, 34, 97, 175, 171, 101, 285,
36, 276, 274, 232, 56, 270, 376, 278, 58, 59, 16, 230, 141, 32,
267, 227, 159, 241, 133, 249, 397, 247, 263, 149, 393, 395, 257,
243, 251, 113, 265, 119, 384, 427, 147, 255, 61, 22, 238, 236, 105,
310, or 363, or a fragment thereof. A plant produced from the plant
cell has a difference in heat tolerance as compared to a control
plant that does not comprise the exogenous nucleic acid.
[0017] Plant cells comprising an exogenous nucleic acid are
provided herein. In one aspect, the exogenous nucleic acid
comprises a regulatory region operably linked to a nucleotide
sequence encoding a polypeptide. The HMM bit score of the amino
acid sequence of the polypeptide is greater than about 20, using an
HMM based on the amino acid sequences depicted in one of FIGS.
1-16. A plant produced from the cells has a difference in drought
tolerance or heat tolerance as compared to a control plant that
does not comprise the exogenous nucleic acid. In another aspect,
the exogenous nucleic acid comprises a regulatory region operably
linked to a nucleotide sequence encoding a polypeptide having 80
percent or greater sequence identity to an amino acid sequence
selected from the group consisting of SEQ ID NO: 2, 3, 4, 5, 6, 7,
8, 9, 10, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37,
39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 60, 62, 63, 64, 65, 66, 67,
68, 69, 70, 71, 73, 74, 76, 78, 80, 82, 84, 86, 88, 90, 92, 94, 96,
98, 100, 102, 104, 106, 108, 110, 112, 114, 116, 118, 120, 122,
124, 126, 127, 130, 132, 134, 136, 138, 139, 142, 144, 146, 148,
150, 152, 154, 156, 158, 160, 162, 164, 166, 168, 170, 172, 174,
176, 178, 179, 180, 182, 184, 186, 188, 190, 192, 194, 196, 198,
200, 202, 204, 206, 208, 209, 212, 214, 216, 218, 220, 222, 224,
225, 228, 229, 231, 233, 234, 235, 237, 239, 240, 242, 244, 246,
248, 250, 252, 253, 254, 256, 258, 260, 262, 264, 266, 268, 269,
271, 273, 275, 277, 279, 280, 282, 284, 286, 288, 289, 290, 291,
292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304,
305, 307, 309, 311, 312, 314, 315, 316, 317, 319, 320, 321, 322,
323, 325, 326, 327, 329, 330, 332, 334, 335, 336, 337, 338, 339,
340, 341, 343, 344, 346, 348, 349, 350, 351, 353, 355, 356, 358,
360, 362, 364, 366, 368, 369, 370, 371, 373, 375, 377, 379, 381,
383, 385, 387, 389, 390, 392, 394, 396, 398, 400, 401, 402, 404,
405, 407, 408, 410, 412, 413, 414, 416, 418, 419, 420, 421, 422,
423, 424, 426, 428, 430, 431, 432, 433, 434, 435, 436, 438, 440,
442, 443, 445, 447, 449, 451, 453, 455, 457, 459, 461, 463, 465,
467, 469, 470, 472, 474, 476, 478, 480, 481, 483, 485, 487, 489,
490, 491, 492, 494, 496, 498, 500, 502, 504, 506, 508, 510, 512,
514, 515, 516, 518, 519, 520, 522, 524, 526, 528, 530, 531, 533,
535, 536, 538, 540, 541, 543, 545, 547, 548, 550, 552, 553, 555,
557, 558, 559, 561, 562, 564, 566, 568, 569, 570, 572, 574, 576,
578, 580, 582, 584, 586, 588, 590, 591, 592, 593, 594, 595, 596,
597, 598, 599, 600, 602, 603, 604, 605, 607, 608, 610, 612, 614,
615, 616, 617, 618, 619, 620, 621, 623, 625, 627, 629, 631, 632,
634, 635, 636, 638, 640, 642, 644, 646, 647, 648, 650, 651, 653,
654, 655, 656, 658, 660, 661, 662, 663, 664, 665, 666, 667, 668,
669, 671, 673, 674, 675, 676, 678, 680, 682, 684, 685, 687, 688,
690, 691, 692, 694, 696, 698, 700, 702, 705, 706, 707, 708, 709,
711, 712, 714, 715, 717, 719, 721, 723, 725, 726, 727, 729, 731,
733, 734, 735, 737, 738, 740, 741, 742, 743, 744, 745, 746, 747,
748, 749, 751, 752, 753, 754, 755, 756, 757, 759, 760, 761, 763,
765, 767, 768, 769, 771, 773, 774, 775, 777, 779, 781, 783, 785,
787, 789, 791, 793, 795, 796, 797, 798, 799, 801, 803, 805, 807,
808, 809, 810, 811, 812, 813, 815, 816, 817, 819, 821, 823, 825,
827, 829, 832, 834, 836, 839, 840, 841, 842, 843, 844, 845, 846,
848, 849, 850, 851, 852, 853, 854, 856, 857, 859, 861, 863, 865,
867, 869, 870, 871, 872, 873, 874, 876, 878, 880, 882, 885, 886,
887, 888, 889, 890, 891, 892, 893, 894, 896, 897, 898, 900, 901,
902, 904, 906, 908, 910, 912, 913, 914, 915, 916, 918, 920, 922,
924, 926, 929, 931, 932, 933, 935, 937, 939, 941, 943, 944, 945,
947, 949, 951, 952, 953, 955, 957, 958, 959, 960, 961, 962, 963,
964, 966, 968, 969, 970, 971, 973, 974, 975, 977, 978, 979, 980,
981, 983, 985, 987, 988, 989, 990, 991, 992, 993, 995, 997, 998,
999, 1000, 1001, 1003, 1005, 1007, 1010, 1012, 1013, 1014, 1015,
1016, 1018, 1020, 1022, 1023, 1025, 1027, 1028, 1030, 1031, 1033,
1035, 1037, 1039, 1041, 1043, 1045, 1046, 1048, 1050, 1051, 1053,
1055, 1056, 1058, 1059, 1061, 1062, 1064, 1066, 1068, 1069, 1071,
1073, 1075, 1077, 1079, 1080, 1082, 1084, 1086, 1088, 1090, 1092,
1093, 1094, 1096, 1098, 1100, 1102, 1104, 1106, 1108, 1110, 1112,
1113, 1114, 1116, 1117, 1118, 1119, 1121, 1123, 1125, 1129, 1130,
1131, 1132, 1133, 1134, 1135, 1136, 1137, 1138, 1140, 1142, 1144,
1146, 1148, 1150, 1151, 1152, 1153, 1154, 1156, 1157, 1158, 1159,
1160, 1161, 1162, 1164, 1166, 1168, 1170, 1172, 1174, 1175, 1177,
1179, 1181, 1182, 1184, 1186, 1188, 1189, 1191, 1193, 1195, 1197,
1199, 1201, 1203, 1205, 1207, 1209, 1210, 1212, 1214, 1215, 1216,
1218, 1219, 1221, 1222, 1223, 1225, 1226, 1228, 1230, 1231, 1232,
1234, 1236, 1238, 1240, 1242, 1244, 1246, 1248, 1250, 1252, 1254,
1256, 1258, 1262, 1264, 1265, 1267, 1269, 1271, 1273, 1274, 1276,
1277, 1279, 1281, 1283, 1285, 1286, 1287, 1288, 1290, 1292, 1293,
1294, 1296, 1298, 1300, 1302, 1304, or 1311, 1313, 1315, 1317,
1319, 1321, 1323, 1324, 1325, 1326, 1327, 1329, 1331, 1333, 1334,
1336, 1338, 1340, 1342, 1343, 1345, 1347, 1349, 1350, 1351, 1353,
1354, 1356, 1358, 1360, or 1361. A plant produced from the plant
cell has a difference in drought tolerance or heat tolerance as
compared to a control plant that does not comprise the exogenous
nucleic acid.
[0018] In another aspect, the exogenous nucleic acid comprises a
regulatory region operably linked to a nucleotide sequence having
80 percent or greater sequence identity to a nucleotide sequence
selected from the group consisting of SEQ ID NO: 1, 12, 14, 16, 18,
20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52,
54, 56, 58, 59, 61, 72, 75, 77, 79, 81, 83, 85, 87, 89, 91, 93, 95,
97, 99, 101, 103, 105, 107, 109, 111, 113, 115, 117, 119, 121, 123,
125, 128, 129, 131, 133, 135, 137, 140, 141, 143, 145, 147, 149,
151, 153, 155, 157, 159, 161, 163, 165, 167, 169, 171, 173, 175,
177, 181, 183, 185, 187, 189, 191, 193, 195, 197, 199, 201, 203,
205, 207, 210, 211, 213, 215, 217, 219, 221, 223, 226, 227, 230,
232, 236, 238, 241, 243, 245, 247, 249, 251, 255, 257, 259, 261,
263, 265, 267, 270, 272, 274, 276, 278, 281, 283, 285, 287, 306,
308, 310, 313, 318, 324, 328, 331, 333, 342, 345, 347, 352, 354,
357, 359, 361, 363, 365, 367, 372, 374, 376, 378, 380, 382, 384,
386, 388, 391, 393, 395, 397, 399, 403, 406, 409, 411, 415, 417,
425, 427, 429, 437, 439, 441, 444, 446, 448, 450, 452, 454, 456,
458, 460, 462, 464, 466, 468, 471, 473, 475, 477, 479, 482, 484,
486, 488, 493, 495, 497, 499, 501, 503, 505, 507, 509, 511, 513,
517, 521, 523, 525, 527, 529, 532, 534, 537, 539, 542, 544, 546,
549, 551, 554, 556, 560, 563, 565, 567, 571, 573, 575, 577, 579,
581, 583, 585, 587, 589, 601, 606, 609, 611, 613, 622, 624, 626,
628, 630, 633, 637, 639, 641, 643, 645, 649, 652, 657, 659, 670,
672, 677, 679, 681, 683, 686, 689, 693, 695, 697, 699, 701, 703,
704, 710, 713, 716, 718, 720, 722, 724, 728, 730, 732, 736, 739,
750, 758, 762, 764, 766, 770, 772, 776, 778, 780, 782, 784, 786,
788, 790, 792, 794, 800, 802, 804, 806, 814, 818, 820, 822, 824,
826, 828, 830, 831, 833, 835, 837, 838, 847, 855, 858, 860, 862,
864, 866, 868, 875, 877, 879, 881, 883, 884, 895, 899, 903, 905,
907, 909, 911, 917, 919, 921, 923, 925, 927, 928, 930, 934, 936,
938, 940, 942, 946, 948, 950, 954, 956, 965, 967, 972, 976, 982,
984, 986, 994, 996, 1002, 1004, 1006, 1008, 1009, 1011, 1017, 1019,
1021, 1024, 1026, 1029, 1032, 1034, 1036, 1038, 1040, 1042, 1044,
1047, 1049, 1052, 1054, 1057, 1060, 1063, 1065, 1067, 1070, 1072,
1074, 1076, 1078, 1081, 1083, 1085, 1087, 1089, 1091, 1095, 1097,
1099, 1101, 1103, 1105, 1107, 1109, 1111, 1115, 1120, 1122, 1124,
1126, 1127, 1128, 1139, 1141, 1143, 1145, 1147, 1149, 1155, 1163,
1165, 1167, 1169, 1171, 1173, 1176, 1178, 1180, 1183, 1185, 1187,
1190, 1192, 1194, 1196, 1198, 1200, 1202, 1204, 1206, 1208, 1211,
1213, 1217, 1220, 1224, 1227, 1229, 1233, 1235, 1237, 1239, 1241,
1243, 1245, 1247, 1249, 1251, 1253, 1255, 1257, 1259, 1260, 1261,
1263, 1266, 1268, 1270, 1272, 1275, 1278, 1280, 1282, 1284, 1289,
1291, 1295, 1297, 1299, 1301, 1303, 1312, 1314, 1316, 1318, 1320,
1322, 1328, 1330, 1332, 1335, 1337, 1339, 1341, 1344, 1346, 1348,
1352, 1355, 1357, 1359 or 1362, or a fragment thereof. A plant
produced from the plant cell has a difference in drought or heat
tolerance as compared to a control plant that does not comprise the
exogenous nucleic acid. A transgenic plant comprising such a plant
cell is also provided.
[0019] Transgenic plants of any aspect provided herein can be a
member of a species selected from the group consisting of Panicum
virgatum (switchgrass), Sorghum bicolor (sorghum, sudangrass),
Miscanthus giganteus (miscanthus), Saccharum sp. (energycane),
Populus balsamifera (poplar), Zea mays (corn), Glycine max
(soybean), Brassica napus (canola), Triticum aestivum (wheat),
Gossypium hirsutum (cotton), Oryza sativa (rice), Helianthus annuus
(sunflower), Medicago sativa (alfalfa), Beta vulgaris (sugarbeet),
or Pennisetum glaucum (pearl millet).
[0020] Isolated nucleic acids are also provided. In one aspect, an
isolated nucleic acid comprises a nucleotide sequence having 80% or
greater sequence identity to the nucleotide sequence set forth in
SEQ ID NO: 1, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36,
38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 59, 61, 72, 75, 77, 79,
81, 83, 85, 87, 89, 91, 93, 95, 97, 99, 101, 103, 105, 107, 109,
111, 113, 115, 117, 119, 121, 123, 125, 128, 129, 131, 133, 135,
137, 140, 141, 143, 145, 147, 149, 151, 153, 155, 157, 159, 161,
163, 165, 167, 169, 171, 173, 175, 177, 181, 183, 185, 187, 189,
191, 193, 195, 197, 199, 201, 203, 205, 207, 210, 211, 213, 215,
217, 219, 221, 223, 226, 227, 230, 232, 236, 238, 241, 243, 245,
247, 249, 251, 255, 257, 259, 261, 263, 265, 267, 270, 272, 274,
276, 278, 281, 283, 285, 287, 306, 308, 310, 313, 318, 324, 328,
331, 333, 342, 345, 347, 352, 354, 357, 359, 361, 363, 365, 367,
372, 374, 376, 378, 380, 382, 384, 386, 388, 391, 393, 395, 397,
399, 403, 406, 409, 411, 415, 417, 425, 427, 429, 437, 439, 441,
444, 446, 448, 450, 452, 454, 456, 458, 460, 462, 464, 466, 468,
471, 473, 475, 477, 479, 482, 484, 486, 488, 493, 495, 497, 499,
501, 503, 505, 507, 509, 511, 513, 517, 521, 523, 525, 527, 529,
532, 534, 537, 539, 542, 544, 546, 549, 551, 554, 556, 560, 563,
565, 567, 571, 573, 575, 577, 579, 581, 583, 585, 587, 589, 601,
606, 609, 611, 613, 622, 624, 626, 628, 630, 633, 637, 639, 641,
643, 645, 649, 652, 657, 659, 670, 672, 677, 679, 681, 683, 686,
689, 693, 695, 697, 699, 701, 703, 704, 710, 713, 716, 718, 720,
722, 724, 728, 730, 732, 736, 739, 750, 758, 762, 764, 766, 770,
772, 776, 778, 780, 782, 784, 786, 788, 790, 792, 794, 800, 802,
804, 806, 814, 818, 820, 822, 824, 826, 828, 830, 831, 833, 835,
837, 838, 847, 855, 858, 860, 862, 864, 866, 868, 875, 877, 879,
881, 883, 884, 895, 899, 903, 905, 907, 909, 911, 917, 919, 921,
923, 925, 927, 928, 930, 934, 936, 938, 940, 942, 946, 948, 950,
954, 956, 965, 967, 972, 976, 982, 984, 986, 994, 996, 1002, 1004,
1006, 1008, 1009, 1011, 1017, 1019, 1021, 1024, 1026, 1029, 1032,
1034, 1036, 1038, 1040, 1042, 1044, 1047, 1049, 1052, 1054, 1057,
1060, 1063, 1065, 1067, 1070, 1072, 1074, 1076, 1078, 1081, 1083,
1085, 1087, 1089, 1091, 1095, 1097, 1099, 1101, 1103, 1105, 1107,
1109, 1111, 1115, 1120, 1122, 1124, 1126, 1127, 1128, 1139, 1141,
1143, 1145, 1147, 1149, 1155, 1163, 1165, 1167, 1169, 1171, 1173,
1176, 1178, 1180, 1183, 1185, 1187, 1190, 1192, 1194, 1196, 1198,
1200, 1202, 1204, 1206, 1208, 1211, 1213, 1217, 1220, 1224, 1227,
1229, 1233, 1235, 1237, 1239, 1241, 1243, 1245, 1247, 1249, 1251,
1253, 1255, 1257, 1259, 1260, 1261, 1263, 1266, 1268, 1270, 1272,
1275, 1278, 1280, 1282, 1284, 1289, 1291, 1295, 1297, 1299, 1301,
1303, 1312, 1314, 1316, 1318, 1320, 1322, 1328, 1330, 1332, 1335,
1337, 1339, 1341, 1344, 1346, 1348, 1352, 1355, 1357, 1359 or 1362,
or a fragment thereof.
[0021] In another aspect, an isolated nucleic acid comprises a
nucleotide sequence encoding a polypeptide having 80% or greater
sequence identity to the amino acid sequence set forth in SEQ ID
NO: 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 13, 15, 17, 19, 21, 23, 25, 27,
29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 60, 62,
63, 64, 65, 66, 67, 68, 69, 70, 71, 73, 74, 76, 78, 80, 82, 84, 86,
88, 90, 92, 94, 96, 98, 100, 102, 104, 106, 108, 110, 112, 114,
116, 118, 120, 122, 124, 126, 127, 130, 132, 134, 136, 138, 139,
142, 144, 146, 148, 150, 152, 154, 156, 158, 160, 162, 164, 166,
168, 170, 172, 174, 176, 178, 179, 180, 182, 184, 186, 188, 190,
192, 194, 196, 198, 200, 202, 204, 206, 208, 209, 212, 214, 216,
218, 220, 222, 224, 225, 228, 229, 231, 233, 234, 235, 237, 239,
240, 242, 244, 246, 248, 250, 252, 253, 254, 256, 258, 260, 262,
264, 266, 268, 269, 271, 273, 275, 277, 279, 280, 282, 284, 286,
288, 289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300,
301, 302, 303, 304, 305, 307, 309, 311, 312, 314, 315, 316, 317,
319, 320, 321, 322, 323, 325, 326, 327, 329, 330, 332, 334, 335,
336, 337, 338, 339, 340, 341, 343, 344, 346, 348, 349, 350, 351,
353, 355, 356, 358, 360, 362, 364, 366, 368, 369, 370, 371, 373,
375, 377, 379, 381, 383, 385, 387, 389, 390, 392, 394, 396, 398,
400, 401, 402, 404, 405, 407, 408, 410, 412, 413, 414, 416, 418,
419, 420, 421, 422, 423, 424, 426, 428, 430, 431, 432, 433, 434,
435, 436, 438, 440, 442, 443, 445, 447, 449, 451, 453, 455, 457,
459, 461, 463, 465, 467, 469, 470, 472, 474, 476, 478, 480, 481,
483, 485, 487, 489, 490, 491, 492, 494, 496, 498, 500, 502, 504,
506, 508, 510, 512, 514, 515, 516, 518, 519, 520, 522, 524, 526,
528, 530, 531, 533, 535, 536, 538, 540, 541, 543, 545, 547, 548,
550, 552, 553, 555, 557, 558, 559, 561, 562, 564, 566, 568, 569,
570, 572, 574, 576, 578, 580, 582, 584, 586, 588, 590, 591, 592,
593, 594, 595, 596, 597, 598, 599, 600, 602, 603, 604, 605, 607,
608, 610, 612, 614, 615, 616, 617, 618, 619, 620, 621, 623, 625,
627, 629, 631, 632, 634, 635, 636, 638, 640, 642, 644, 646, 647,
648, 650, 651, 653, 654, 655, 656, 658, 660, 661, 662, 663, 664,
665, 666, 667, 668, 669, 671, 673, 674, 675, 676, 678, 680, 682,
684, 685, 687, 688, 690, 691, 692, 694, 696, 698, 700, 702, 705,
706, 707, 708, 709, 711, 712, 714, 715, 717, 719, 721, 723, 725,
726, 727, 729, 731, 733, 734, 735, 737, 738, 740, 741, 742, 743,
744, 745, 746, 747, 748, 749, 751, 752, 753, 754, 755, 756, 757,
759, 760, 761, 763, 765, 767, 768, 769, 771, 773, 774, 775, 777,
779, 781, 783, 785, 787, 789, 791, 793, 795, 796, 797, 798, 799,
801, 803, 805, 807, 808, 809, 810, 811, 812, 813, 815, 816, 817,
819, 821, 823, 825, 827, 829, 832, 834, 836, 839, 840, 841, 842,
843, 844, 845, 846, 848, 849, 850, 851, 852, 853, 854, 856, 857,
859, 861, 863, 865, 867, 869, 870, 871, 872, 873, 874, 876, 878,
880, 882, 885, 886, 887, 888, 889, 890, 891, 892, 893, 894, 896,
897, 898, 900, 901, 902, 904, 906, 908, 910, 912, 913, 914, 915,
916, 918, 920, 922, 924, 926, 929, 931, 932, 933, 935, 937, 939,
941, 943, 944, 945, 947, 949, 951, 952, 953, 955, 957, 958, 959,
960, 961, 962, 963, 964, 966, 968, 969, 970, 971, 973, 974, 975,
977, 978, 979, 980, 981, 983, 985, 987, 988, 989, 990, 991, 992,
993, 995, 997, 998, 999, 1000, 1001, 1003, 1005, 1007, 1010, 1012,
1013, 1014, 1015, 1016, 1018, 1020, 1022, 1023, 1025, 1027, 1028,
1030, 1031, 1033, 1035, 1037, 1039, 1041, 1043, 1045, 1046, 1048,
1050, 1051, 1053, 1055, 1056, 1058, 1059, 1061, 1062, 1064, 1066,
1068, 1069, 1071, 1073, 1075, 1077, 1079, 1080, 1082, 1084, 1086,
1088, 1090, 1092, 1093, 1094, 1096, 1098, 1100, 1102, 1104, 1106,
1108, 1110, 1112, 1113, 1114, 1116, 1117, 1118, 1119, 1121, 1123,
1125, 1129, 1130, 1131, 1132, 1133, 1134, 1135, 1136, 1137, 1138,
1140, 1142, 1144, 1146, 1148, 1150, 1151, 1152, 1153, 1154, 1156,
1157, 1158, 1159, 1160, 1161, 1162, 1164, 1166, 1168, 1170, 1172,
1174, 1175, 1177, 1179, 1181, 1182, 1184, 1186, 1188, 1189, 1191,
1193, 1195, 1197, 1199, 1201, 1203, 1205, 1207, 1209, 1210, 1212,
1214, 1215, 1216, 1218, 1219, 1221, 1222, 1223, 1225, 1226, 1228,
1230, 1231, 1232, 1234, 1236, 1238, 1240, 1242, 1244, 1246, 1248,
1250, 1252, 1254, 1256, 1258, 1262, 1264, 1265, 1267, 1269, 1271,
1273, 1274, 1276, 1277, 1279, 1281, 1283, 1285, 1286, 1287, 1288,
1290, 1292, 1293, 1294, 1296, 1298, 1300, 1302, 1304, or 1311,
1313, 1315, 1317, 1319, 1321, 1323, 1324, 1325, 1326, 1327, 1329,
1331, 1333, 1334, 1336, 1338, 1340, 1342, 1343, 1345, 1347, 1349,
1350, 1351, 1353, 1354, 1356, 1358, 1360, or 1361.
[0022] In another aspect, methods of identifying a genetic
polymorphism associated with variation in drought tolerance or heat
tolerance are provided. The methods include providing a population
of plants, and determining whether one or more genetic
polymorphisms in the population are genetically linked to the locus
for a polypeptide selected from the group consisting of the
polypeptides depicted in FIGS. 1-16 and functional homologs
thereof. The correlation between variation in drought tolerance or
heat tolerance in plants of the population and the presence of the
one or more genetic polymorphisms in plants of the population is
measured, thereby permitting identification of whether or not the
one or more genetic polymorphisms are associated with such
variation. The population can be a population of switchgrass,
sorghum, sugar cane, or miscanthus plants.
[0023] In another aspect, methods of making a plant line are
provided. The methods include determining whether one or more
genetic polymorphisms in a population of plants is associated with
the locus for a polypeptide selected from the group consisting of
the polypeptides depicted in FIGS. 1-16 and functional homologs
thereof; identifying one or more plants in said population in which
the presence of at least one allele at the one or more genetic
polymorphisms is associated with variation in drought tolerance or
heat tolerance; crossing each of the one or more identified plants
with itself or a different plant to produce seed; crossing at least
one progeny plant grown from the seed with itself or a different
plant; and repeating the crossing steps for an additional 0-5
generations to make the plant line, where at least one allele is
present in the plant line. The population can be a population of
switchgrass, sorghum, sugar cane, or miscanthus plants.
[0024] Unless otherwise defined, all technical and scientific terms
used herein have the same meaning as commonly understood by one of
ordinary skill in the art to which this invention pertains.
Although methods and materials similar or equivalent to those
described herein can be used to practice the invention, suitable
methods and materials are described below. All publications, patent
applications, patents, and other references mentioned herein are
incorporated by reference in their entirety. In case of conflict,
the present specification, including definitions, will control. In
addition, the materials, methods, and examples are illustrative
only and not intended to be limiting.
[0025] The details of one or more embodiments of the invention are
set forth in the accompanying drawings and the description below.
Other features, objects, and advantages of the invention will be
apparent from the description and drawings, and from the claims.
The word "comprising" in the claims may be replaced by "consisting
essentially of or by "consisting of," according to standard
practice in patent law.
DESCRIPTION OF THE DRAWINGS
[0026] FIG. 1 is an alignment of Ceres SEEDLINE ID no. ME00029 (SEQ
ID NO:442) with homologous and/or orthologous sequences, including,
Public GI ID no. 72384401 (SEQ ID NO:443), Ceres CLONE ID no.
1079382 (SEQ ID NO:445), Ceres CLONE ID no. 1853461 (SEQ ID
NO:447), Ceres CLONE ID no. 1626485 (SEQ ID NO:451), Ceres CLONE ID
no. 1713920 (SEQ ID NO:459), Ceres CLONE ID no. 1772747 (SEQ ID
NO:463), Ceres CLONE ID no. 225960 (SEQ ID NO:465), Public GI ID
no. 115443807 (SEQ ID NO:470), and Ceres CLONE ID no. 569388 (SEQ
ID NO:483). In all the alignment figures shown herein, a dash in an
aligned sequence represents a gap, i.e., a lack of an amino acid at
that position. Identical amino acids or conserved amino acid
substitutions among aligned sequences are identified by boxes. FIG.
1 and the other alignment figures provided herein were generated
using the program MUSCLE version 3.52.
[0027] FIG. 2 is an alignment of Ceres SEEDLINE ID no. ME00045 (SEQ
ID NO:489) with homologous and/or orthologous sequences, including,
Ceres CLONE ID no. 571200 (SEQ ID NO:494), Ceres CLONE ID no.
1928532 (SEQ ID NO:496), Ceres ANNOT ID no. 1490637 (SEQ ID
NO:498), Ceres CLONE ID no. 295496 (SEQ ID NO:514), Public GI ID
no. 115463637 (SEQ ID NO:515), and Ceres CLONE ID no. 1999444 (SEQ
ID NO:518).
[0028] FIG. 3 is an alignment of Ceres SEEDLINE ID no. ME02190 (SEQ
ID NO:568) with homologous and/or orthologous sequences, including,
Ceres CLONE ID no. 695006 (SEQ ID NO:578) and Ceres ANNOT ID no.
1527488 (SEQ ID NO:584).
[0029] FIG. 4 is an alignment of Ceres SEEDLINE ID no. ME02549 (SEQ
ID NO:590) with homologous and/or orthologous sequences, including,
Ceres ANNOT ID no. 1501305 (SEQ ID NO:623), Ceres CLONE ID no.
607280 (SEQ ID NO:627), Public GI ID no. 92887174 (SEQ ID NO:632),
Ceres CLONE ID no. 1857162 (SEQ ID NO:634), Public GI ID no.
115477272 (SEQ ID NO:674), and Ceres CLONE ID no. 264002 (SEQ ID
NO:682).
[0030] FIG. 5 is an alignment of Ceres SEEDLINE ID no. ME02865 (SEQ
ID NO:705) with homologous and/or orthologous sequences, including,
Public GI ID no. 79320952 (SEQ ID NO:706) and Public GI ID no.
79320957 (SEQ ID NO:707).
[0031] FIG. 6 is an alignment of Ceres SEEDLINE ID no. ME03227 (SEQ
ID NO:711) with homologous and/or orthologous sequences, including,
Ceres CLONE ID no. 964616 (SEQ ID NO:714), Ceres CLONE ID no.
100009667 (SEQ ID NO:715), Ceres ANNOT ID no. 1444568 (SEQ ID
NO:719), Ceres CLONE ID no. 719489 (SEQ ID NO:725), Ceres CLONE ID
no. 587748 (SEQ ID NO:733), Public GI ID no. 125528114 (SEQ ID
NO:734), and Ceres CLONE ID no. 274172 (SEQ ID NO:737).
[0032] FIG. 7 is an alignment of Ceres SEEDLINE ID no. ME04477 (SEQ
ID NO:742) with homologous and/or orthologous sequences, including,
Ceres CLONE ID no. 1620215 (SEQ ID NO: 759); Public GI ID no.
38016527 (SEQ ID NO: 796); Ceres CLONE ID no. 1798756 (SEQ ID NO:
763); Public GI ID no. 75133829 (SEQ ID NO: 799); Ceres ANNOT ID
no. 1460527 (SEQ ID NO: 801); Public GI ID no. 119720772 (SEQ ID
NO: 857); Ceres CLONE ID no. 708446 (SEQ ID NO: 859); Public GI ID
no. 92896423 (SEQ ID NO: 892); Ceres CLONE ID no. 1387149 (SEQ ID
NO: 912); Public GI ID no. 5031281 (SEQ ID NO: 953); Ceres CLONE ID
no. 1775820 (SEQ ID NO: 955); Public GI ID no. 35187687 (SEQ ID NO:
988); Public GI ID no. 115468934 (SEQ ID NO: 991); Public GI ID no.
118424243 (SEQ ID NO: 1000); Ceres ANNOT ID no. 6063957 (SEQ ID NO:
1010); CeresClone: 1030374 (SEQ ID NO: 1306); Public GI ID no.
113196593 (SEQ ID NO: 798); Public GI ID no. 112819496 (SEQ ID NO:
999), and Public GI ID no. 169363 (SEQ ID NO: 1215).
[0033] FIG. 8 is an alignment of Ceres SEEDLINE ID no. ME18396 (SEQ
ID NO:1012) with homologous and/or orthologous sequences,
including, Ceres CLONE ID no. 287430 (SEQ ID NO:1018), Ceres ANNOT
ID no. 451889 (SEQ ID NO:1020), Ceres CLONE ID no. 936084 (SEQ ID
NO:1055), Ceres CLONE ID no. 1792501 (SEQ ID NO:1066), Ceres ANNOT
ID no. 1437875 (SEQ ID NO:1068), Public GI ID no. 1853968 (SEQ ID
NO:1093), Public GI ID no. 27530032 (SEQ ID NO:1094), Ceres CLONE
ID no. 1834483 (SEQ ID NO:1110), Public GI ID no. 84579418 (SEQ ID
NO:1113), Public GI ID no. 15077030 (SEQ ID NO:1117), and Public GI
ID no. 13752458 (SEQ ID NO:1118).
[0034] FIG. 9 is an alignment of Ceres SEEDLINE ID no. ME20095 (SEQ
ID NO:1129) with homologous and/or orthologous sequences,
including, Public GI ID no. 72255610 (SEQ ID NO:933), Ceres CLONE
ID no. 1927040 (SEQ ID NO:926), Ceres CLONE ID no. 1562633 (SEQ ID
NO:1140), Ceres ANNOT ID no. 1692728 (SEQ ID NO:1142), Ceres CLONE
ID no. 921110 (SEQ ID NO:1144), Ceres CLONE ID no. 1791180 (SEQ ID
NO:1146), Ceres CLONE ID no. 527891 (SEQ ID NO:1148), Ceres ANNOT
ID no. 1522414 (SEQ ID NO:1150), and Public GI ID no. 51458330 (SEQ
ID NO:1182).
[0035] FIG. 10 is an alignment of Ceres CLONE ID no. 31309 (SEQ ID
NO:60) with homologous and/or orthologous sequences, including,
Ceres CLONE ID no. 872030 (SEQ ID NO:62), Ceres CLONE ID no.
100029223 (SEQ ID NO:74), Ceres CLONE ID no. 1939845 (SEQ ID
NO:98), Ceres CLONE ID no. 2015383 (SEQ ID NO:102), Ceres CLONE ID
no. 1607893 (SEQ ID NO:104), Ceres CLONE ID no. 1075133 (SEQ ID
NO:152), and Ceres CLONE ID no. 1218065 (SEQ ID NO:915).
[0036] FIG. 11 is an alignment of Ceres LOCUS ID no.At2g26150 (SEQ
ID NO:364) with homologous and/or orthologous sequences, including,
Ceres ANNOT ID no. 1455221 (SEQ ID NO:407), Public GI ID no. 729774
(SEQ ID NO:408), Ceres CLONE ID no. 1414288 (SEQ ID NO:410), Public
GI ID no. 115482048 (SEQ ID NO:413), and Public GI ID no. 56117815
(SEQ ID NO:421).
[0037] FIG. 12 is an alignment of Ceres LOCUS ID no.At1g32330 (SEQ
ID NO:311) with homologous and/or orthologous sequences, including,
Ceres ANNOT ID no. 1538958 (SEQ ID NO:319), Public GI ID no.
115521213 (SEQ ID NO:320), Public GI ID no. 42415865 (SEQ ID
NO:321), Public GI ID no. 729775 (SEQ ID NO:322), Public GI ID no.
11386827 (SEQ ID NO:323), Public GI ID no. 115456675 (SEQ ID
NO:326), and Public GI ID no. 89274218 (SEQ ID NO:335).
[0038] FIG. 13 is an alignment of Ceres CLONE ID no. 41543 (SEQ ID
NO:160) with homologous and/or orthologous sequences, including,
Ceres CLONE ID no. 1837065 (SEQ ID NO:162), Ceres ANNOT ID no.
1531178 (SEQ ID NO:184), Ceres CLONE ID no. 470694 (SEQ ID NO:242),
Public GI ID no. 92867368 (SEQ ID NO:253), Ceres CLONE ID no.
859707 (SEQ ID NO:256), Ceres CLONE ID no. 392275 (SEQ ID NO:268),
Ceres CLONE ID no. 1828394 (SEQ ID NO:282), and Public GI ID no.
115466694 (SEQ ID NO:294).
[0039] FIG. 14 is an alignment of Ceres CLONE ID no. 14572 (SEQ ID
NO:2) with homologous and/or orthologous sequences, including,
Public GI ID no. 115470807 (SEQ ID NO:5), Ceres CLONE ID no.
1842931 (SEQ ID NO:15), Ceres CLONE ID no. 321308 (SEQ ID NO:17),
Ceres CLONE ID no. 1725811 (SEQ ID NO:19), Ceres CLONE ID no.
1357455 (SEQ ID NO:21), Ceres CLONE ID no. 943370 (SEQ ID NO:23),
Ceres CLONE ID no. 1327712 (SEQ ID NO:25), Ceres CLONE ID no.
1764692 (SEQ ID NO:27), and Ceres ANNOT ID no. 1512656 (SEQ ID
NO:41).
[0040] FIG. 15 is an alignment of Ceres SEEDLINE ID no. ME02401
(SEQ ID NO:749) with homologous and/or orthologous sequences,
including, Ceres CLONE ID no. 1847516 (SEQ ID NO:1195), Ceres CLONE
ID no. 1961986 (SEQ ID NO:1201), and Ceres ANNOT ID no. 6091930
(SEQ ID NO:1207).
[0041] FIG. 16 is an alignment of Ceres SEEDLINE ID no. ME004246
(SEQ ID NO:1311) with homologous and/or orthologous sequences,
including, CeresClone: 971761 (SEQ ID NO: 1313), CeresClone:
1946574 (SEQ ID NO: 1331), CeresClone: 2055551 (SEQ ID NO: 1340),
CeresClone: 100045499 (SEQ ID NO: 1347), CeresClone: 1465853 (SEQ
ID NO: 1353), and CeresClone: 753982 (SEQ ID NO: 1360).
DETAILED DESCRIPTION
[0042] The invention features methods and materials related to
increasing heat and/or drought tolerance in plants. In some
embodiments, the plants may have increased heat and drought
tolerance. The methods can include transforming a plant cell with a
nucleic acid encoding a heat and/or drought-tolerance polypeptide,
wherein expression of the polypeptide results in increased heat
and/or drought tolerance. Plant cells produced using such methods
can be grown to produce plants having an increased heat and/or
drought tolerance. Such plants, and the seeds of such plants, may
be used to produce, for example, may be used to produce biomass
and/or to produce crops with increased yield and/or quality.
I. DEFINITIONS
[0043] "Amino acid" refers to one of the twenty biologically
occurring amino acids and to synthetic amino acids, including D/L
optical isomers.
[0044] "Cell type-preferential promoter" or "tissue-preferential
promoter" refers to a promoter that drives expression
preferentially in a target cell type or tissue, respectively, but
may also lead to some transcription in other cell types or tissues
as well.
[0045] "Control plant" refers to a plant that does not contain the
exogenous nucleic acid present in a transgenic plant of interest,
but otherwise has the same or similar genetic background as such a
transgenic plant. A suitable control plant can be a non-transgenic
wild type plant, a non-transgenic segregant from a transformation
experiment, or a transgenic plant that contains an exogenous
nucleic acid other than the exogenous nucleic acid of interest.
[0046] "Domains" are groups of substantially contiguous amino acids
in a polypeptide that can be used to characterize protein families
and/or parts of proteins. Such domains have a "fingerprint" or
"signature" that can comprise conserved primary sequence, secondary
structure, and/or three-dimensional conformation. Generally,
domains are correlated with specific in vitro and/or in vivo
activities. A domain can have a length of from 10 amino acids to
400 amino acids, e.g., 10 to 50 amino acids, or 25 to 100 amino
acids, or 35 to 65 amino acids, or 35 to 55 amino acids, or 45 to
60 amino acids, or 200 to 300 amino acids, or 300 to 400 amino
acids.
[0047] "Exogenous" with respect to a nucleic acid indicates that
the nucleic acid is part of a recombinant nucleic acid construct,
or is not in its natural environment. For example, an exogenous
nucleic acid can be a sequence from one species introduced into
another species, i.e., a heterologous nucleic acid. Typically, such
an exogenous nucleic acid is introduced into the other species via
a recombinant nucleic acid construct. An exogenous nucleic acid can
also be a sequence that is native to an organism and that has been
reintroduced into cells of that organism. An exogenous nucleic acid
that includes a native sequence can often be distinguished from the
naturally occurring sequence by the presence of non-natural
sequences linked to the exogenous nucleic acid, e.g., non-native
regulatory sequences flanking a native sequence in a recombinant
nucleic acid construct. In addition, stably transformed exogenous
nucleic acids typically are integrated at positions other than the
position where the native sequence is found. It will be appreciated
that an exogenous nucleic acid may have been introduced into a
progenitor and not into the cell under consideration. For example,
a transgenic plant containing an exogenous nucleic acid can be the
progeny of a cross between a stably transformed plant and a
non-transgenic plant. Such progeny are considered to contain the
exogenous nucleic acid.
[0048] "Expression" refers to the process of converting genetic
information of a polynucleotide into RNA through transcription,
which is catalyzed by an enzyme, RNA polymerase, and into protein,
through translation of mRNA on ribosomes.
[0049] "Heterologous polypeptide" as used herein refers to a
polypeptide that is not a naturally occurring polypeptide in a
plant cell, e.g., a transgenic Panicum virgatum plant transformed
with and expressing the coding sequence for a nitrogen transporter
polypeptide from a Zea mays plant.
[0050] "Isolated nucleic acid" as used herein includes a
naturally-occurring nucleic acid, provided one or both of the
sequences immediately flanking that nucleic acid in its
naturally-occurring genome is removed or absent. Thus, an isolated
nucleic acid includes, without limitation, a nucleic acid that
exists as a purified molecule or a nucleic acid molecule that is
incorporated into a vector or a virus. A nucleic acid existing
among hundreds to millions of other nucleic acids within, for
example, cDNA libraries, genomic libraries, or gel slices
containing a genomic DNA restriction digest, is not to be
considered an isolated nucleic acid.
[0051] "Modulation" of the level of tolerance to a stimulus (e.g.,
drought conditions or heat shock conditions) refers to the change
in the level of tolerance of the indicated stimulus that is
observed as a result of expression of, or transcription from, an
exogenous nucleic acid in a plant cell. The change in level is
measured relative to the corresponding level in control plants.
[0052] "Nucleic acid" and "polynucleotide" are used interchangeably
herein, and refer to both RNA and DNA, including cDNA, genomic DNA,
synthetic DNA, and DNA or RNA containing nucleic acid analogs.
Polynucleotides can have any three-dimensional structure. A nucleic
acid can be double-stranded or single-stranded (i.e., a sense
strand or an antisense strand). Non-limiting examples of
polynucleotides include genes, gene fragments, exons, introns,
messenger RNA (mRNA), transfer RNA, ribosomal RNA, siRNA,
micro-RNA, ribozymes, cDNA, recombinant polynucleotides, branched
polynucleotides, nucleic acid probes and nucleic acid primers. A
polynucleotide may contain unconventional or modified
nucleotides.
[0053] "Operably linked" refers to the positioning of a regulatory
region and a sequence to be transcribed in a nucleic acid so that
the regulatory region is effective for regulating transcription or
translation of the sequence. For example, to operably link a coding
sequence and a regulatory region, the translation initiation site
of the translational reading frame of the coding sequence is
typically positioned between one and about fifty nucleotides
downstream of the regulatory region. A regulatory region can,
however, be positioned as much as about 5,000 nucleotides upstream
of the translation initiation site, or about 2,000 nucleotides
upstream of the transcription start site.
[0054] "Polypeptide" as used herein refers to a compound of two or
more subunit amino acids, amino acid analogs, or other
peptidomimetics, regardless of post-translational modification,
e.g., phosphorylation or glycosylation. The subunits may be linked
by peptide bonds or other bonds such as, for example, ester or
ether bonds. Full-length polypeptides, truncated polypeptides,
point mutants, insertion mutants, splice variants, chimeric
proteins, and fragments thereof are encompassed by this
definition.
[0055] "Progeny" includes descendants of a particular plant or
plant line. Progeny of an instant plant include seeds formed on
F.sub.1, F.sub.2, F.sub.3, F.sub.4, F.sub.5, F.sub.6 and subsequent
generation plants, or seeds formed on BC.sub.1, BC.sub.2, BC.sub.3,
and subsequent generation plants, or seeds formed on
F.sub.1BC.sub.1, F.sub.1BC.sub.2, F.sub.1BC.sub.3, and subsequent
generation plants. The designation F.sub.1 refers to the progeny of
a cross between two parents that are genetically distinct. The
designations F.sub.2, F.sub.3, F.sub.4, F.sub.5 and F.sub.6 refer
to subsequent generations of self- or sib-pollinated progeny of an
F.sub.1 plant.
[0056] "Regulatory region" refers to a nucleic acid having
nucleotide sequences that influence transcription or translation
initiation and rate, and stability and/or mobility of a
transcription or translation product. Regulatory regions include,
without limitation, promoter sequences, enhancer sequences,
response elements, protein recognition sites, inducible elements,
protein binding sequences, 5' and 3' untranslated regions (UTRs),
transcriptional start sites, termination sequences, polyadenylation
sequences, introns, and combinations thereof. A regulatory region
typically comprises at least a core (basal) promoter. A regulatory
region also may include at least one control element, such as an
enhancer sequence, an upstream element or an upstream activation
region (UAR). For example, a suitable enhancer is a cis-regulatory
element (-212 to -154) from the upstream region of the octopine
synthase (ocs) gene. Fromm et al., The Plant Cell, 1:977-984
(1989).
[0057] "Up-regulation" refers to regulation that increases the
level of an expression product (mRNA, polypeptide, or both)
relative to basal or native states.
[0058] "Vector" refers to a replicon, such as a plasmid, phage, or
cosmid, into which another DNA segment may be inserted so as to
bring about the replication of the inserted segment. Generally, a
vector is capable of replication when associated with the proper
control elements. The term "vector" includes cloning and expression
vectors, as well as viral vectors and integrating vectors. An
"expression vector" is a vector that includes a regulatory
region.
II. POLYPEPTIDES
[0059] Polypeptides described herein include heat and/or
drought-tolerance polypeptides. Heat and/or drought-tolerance
polypeptides can be effective to increase heat and/or
drought-tolerance when expressed in a plant or plant cell. Such
polypeptides typically contain at least one domain indicative of
heat and/or drought-tolerance polypeptides, as described in more
detail herein. Heat and/or drought-tolerance polypeptides typically
have an HMM bit score that is greater than 20 for an HMM model
based on one of the alignments set forth in FIGS. 1-16, as
described in more detail herein. In some embodiments, heat and/or
drought-tolerance polypeptides have greater than 80% identity to
SEQ ID NO:442, SEQ ID NO:489, SEQ ID NO:568, SEQ ID NO:590, SEQ ID
NO:705, SEQ ID NO:711, SEQ ID NO:742, SEQ ID NO:1012, SEQ ID
NO:1129, SEQ ID NO:60, SEQ ID NO:364, SEQ ID NO:311, SEQ ID NO:160,
SEQ ID NO:2, SEQ ID NO:749, SEQ ID NO:1311, or SEQ ID NO:57, as
described in more detail herein.
[0060] A. Domains Indicative of Heat and/or Drought-Tolerance
Polypeptides
[0061] A drought-tolerance polypeptide can be a RNA polymerase Rpb4
family member. The eukaryotic RNA polymerase subunit RPB4 forms a
heterodimer with subunit RPB7 that reversibly associates with the
RNA polymerase II core. SEQ ID NO:442 sets forth the amino acid
sequence of an Arabidopsis clone, identified herein as Ceres
SEEDLINE ID No. ME00029 (SEQ ID NO:441), that is predicted to
encode a RNA polymerase Rpb4 polypeptide.
[0062] A drought-tolerance polypeptide can contain a
sulfotransferase 1 domain characteristic of polypeptides belonging
to the sulfotransferase family. Members of the sulfotransferase
family catalyze the transfer of sulfate groups to specific
compounds. SEQ ID NO:590, which sets forth the amino acid sequence
of an Arabidopsis clone, identified herein as Ceres SEEDLINE ID No.
ME02549 (SEQ ID NO:589), is predicted to encode a polypeptide
containing a sulfotransferase 1 domain.
[0063] A drought-tolerance polypeptide can contain an AN1-like zinc
finger (zf-AN1) and an A20-like zinc finger (zf-A20) domain. The
AN1-like zinc finger domain was first identified as a zinc finger
at the C-terminus of An1, a ubiquitin-like protein in Xenopus
laevis. The AN1-like zinc finger domain is characterized by the
pattern, C-X2-C-X(9-12)-C-X(1-2)-C-X4-C-X2-H-X5-H-X-C Where X can
be any amino acid, and numbers in brackets indicate the number of
residues. In A20, the A20-like zinc finger domain mediates
self-association and IL-1-induced NF-kappa B activation. SEQ ID
NO:742, which sets forth the amino acid sequence of an Arabidopsis
clone, identified herein as Ceres Clone ID No. 24255 (SEQ ID
NO:1241), is predicted to encode a polypeptide containing an
AN1-like zinc finger domain and a A20-like zinc finger domain. SEQ
ID NO:749, which sets forth the amino acid sequence of an
Arabidopsis clone, identified herein as Ceres SEEDLINE ID No.
ME02401 (SEQ ID NO:1188), is also predicted to encode a polypeptide
containing an AN1-like zinc finger domain and a A20-like zinc
finger domain.
[0064] A drought-tolerance polypeptide can contain a short chain
dehydrogenase (adh_short) domain. Most polypeptides containing a
short chain dehydrogenase domain are polypeptides of about 250 to
300 amino acid residues, and are NAD- or NADP-dependent
oxidoreductases. SEQ ID NO:1012, which sets forth the amino acid
sequence of an Arabidopsis clone, identified herein as Ceres
SEEDLINE ID No. 18396 (SEQ ID NO:1260), is predicted to encode a
polypeptide containing a short chain dehydrogenase domain.
[0065] A drought-tolerance polypeptide can contain a
trehalose-phosphatase (Trehalose_PPase) domain characteristic of
trehalose phosphatase polypeptides. Trehalose phosphatases catalyse
the de-phosphorylation of trehalose-6-phosphate to trehalose and
orthophosphate. Trehalose is a common disaccharide in bacteria,
fungi, and invertebrates that appears to play a role in desiccation
tolerance. SEQ ID NO:1129, which sets forth the amino acid sequence
of an Arabidopsis clone, identified herein as Ceres SEEDLINE ID No.
ME20095 (SEQ ID NO:1126), is predicted to encode a polypeptide
containing a trehalose-phosphatase domain.
[0066] A heat-tolerance polypeptide can be a synaptobrevin-related
polypeptide. SEQ ID NO:160, which sets forth the amino acid
sequence of an Arabidopsis clone, identified herein as Ceres CLONE
ID no. 41543 (SEQ ID NO:159), is predicted to encode a
synaptobrevin-related polypeptide.
[0067] A heat-tolerance polypeptide can contain an HSF-type
DNA-binding domain, which is predicted to be characteristic of heat
shock factor transcription activator. Heat shock factor
transcription activators are often found associated with heat shock
protein promoters during heat shock. SEQ ID NO:311, which sets
forth the amino acid sequence of an Arabidopsis clone, identified
herein as Ceres LOCUS ID no.At1g32330 (SEQ ID NO:310), is predicted
to encode a polypeptide containing an HSF-type DNA-binding domain.
SEQ ID NO:364, which sets forth the amino acid sequence of an
Arabidopsis clone, identified herein as Ceres LOCUS ID no.At2g26150
(SEQ ID NO:363), is also predicted to encode a polypeptide
containing an HSF-type DNA-binding domain.
[0068] A heat-tolerance polypeptide can contain a DnaJ domain. The
prokaryotic heat shock protein DnaJ interacts with the chaperone
hsp70-like DnaK protein. Structurally, the DnaJ protein consists of
an N-terminal conserved domain (called `J` domain) of about 70
amino acids, a glycine-rich region (`G` domain`) of about 30
residues, a central domain containing four repeats of a CXXCXGXG
motif (`CRR` domain) and a C-terminal region of 120 to 170
residues. SEQ ID NO:60, which sets forth the amino acid sequence of
an Arabidopsis clone, identified herein as Ceres CLONE ID no. 31309
(SEQ ID NO:58), is predicted to encode an polypeptide containing a
DnaJ domain.
[0069] A drought-tolerance polypeptide can contain a KH.sub.--2
domain. Most polypeptides containing a KH.sub.--2 domain are
typically around 70 amino acids and are present in a wide variety
of quite diverse nucleic acid-binding proteins. It has been shown
to bind RNA. Like many other RNA-binding motifs, KH motifs are
found in one or multiple copies, each motif is necessary for in
vitro RNA binding activity, suggesting that they may function
cooperatively or, in the case of single KH motif proteins (for
example, Mer1p), independently. According to structural analysis
the KH domain can be separated in two groups. The first group or
type-1 contain a beta-alpha-alpha-beta-beta-alpha structure,
whereas in the type-2 the two last beta-sheet are located in the N
terminal part of the domain (alpha-beta-beta-alpha-alpha-beta).
Sequence similarity between these two folds are limited to a short
region (VIGXXGXXI) in the RNA binding motif. This motif is located
between helices 1 and 2 in type-1 and between helices 2 and 3 in
type-2. SEQ ID NO:1311, which sets forth the amino acid sequence of
an Arabidopsis clone, identified herein as Ceres SEEDLINE ID No.
ME04246 (SEQ ID NO:1362), is predicted to encode a polypeptide
containing a KH.sub.--2 domain.
[0070] B. Functional Homologs Identified by Reciprocal BLAST
[0071] In some embodiments, one or more functional homologs of a
reference heat and/or drought-tolerance polypeptide defined by one
or more of the Pfam descriptions indicated above are suitable for
use as heat and/or drought-tolerance polypeptides. A functional
homolog is a polypeptide that has sequence similarity to a
reference polypeptide, and that carries out one or more of the
biochemical or physiological function(s) of the reference
polypeptide. A functional homolog and the reference polypeptide may
be natural occurring polypeptides, and the sequence similarity may
be due to convergent or divergent evolutionary events. As such,
functional homologs are sometimes designated in the literature as
homologs, or orthologs, or paralogs. Variants of a naturally
occurring functional homolog, such as polypeptides encoded by
mutants of a wild type coding sequence, may themselves be
functional homologs. Functional homologs can also be created via
site-directed mutagenesis of the coding sequence for a heat and/or
drought-tolerance polypeptide, or by combining domains from the
coding sequences for different naturally-occurring heat and/or
drought-tolerance polypeptides ("domain swapping"). The term
"functional homolog" is sometimes applied to the nucleic acid that
encodes a functionally homologous polypeptide.
[0072] Functional homologs can be identified by analysis of
nucleotide and polypeptide sequence alignments. For example,
performing a query on a database of nucleotide or polypeptide
sequences can identify homologs of heat and/or drought-tolerance
polypeptides. Sequence analysis can involve BLAST, Reciprocal
BLAST, or PSI-BLAST analysis of nonredundant databases using a heat
and/or drought-tolerance polypeptide amino acid sequence as the
reference sequence. Amino acid sequence is, in some instances,
deduced from the nucleotide sequence. Those polypeptides in the
database that have greater than 40% sequence identity are
candidates for further evaluation for suitability as a heat and/or
drought-tolerance polypeptide. Amino acid sequence similarity
allows for conservative amino acid substitutions, such as
substitution of one hydrophobic residue for another or substitution
of one polar residue for another. If desired, manual inspection of
such candidates can be carried out in order to narrow the number of
candidates to be further evaluated. Manual inspection can be
performed by selecting those candidates that appear to have domains
present in heat and/or drought-tolerance polypeptides, e.g.,
conserved functional domains.
[0073] Conserved regions can be identified by locating a region
within the primary amino acid sequence of a heat and/or
drought-tolerance polypeptide that is a repeated sequence, forms
some secondary structure (e.g., helices and beta sheets),
establishes positively or negatively charged domains, or represents
a protein motif or domain. See, e.g., the Pfam web site describing
consensus sequences for a variety of protein motifs and domains on
the World Wide Web at sanger.ac.uk/Software/Pfam/ and
pfam.janelia.org/. A description of the information included at the
Pfam database is described in Sonnhammer et al., Nucl. Acids Res.,
26:320-322 (1998); Sonnhammer et al., Proteins, 28:405-420 (1997);
and Bateman et al., Nucl. Acids Res., 27:260-262 (1999). Conserved
regions also can be determined by aligning sequences of the same or
related polypeptides from closely related species. Closely related
species preferably are from the same family. In some embodiments,
alignment of sequences from two different species is adequate.
[0074] Typically, polypeptides that exhibit at least about 20%
amino acid sequence identity are useful to identify conserved
regions. Conserved regions of related polypeptides exhibit at least
45% amino acid sequence identity (e.g., at least 50%, at least 60%,
at least 70%, at least 80%, or at least 90% amino acid sequence
identity). In some embodiments, a conserved region exhibits at
least 92%, 94%, 96%, 98%, or 99% amino acid sequence identity.
[0075] Examples of amino acid sequences of functional homologs of
the polypeptide set forth in SEQ ID NO:442 are provided in FIG. 1
and in the sequence listing. Such functional homologs include
Public GI ID no. 72384401 (SEQ ID NO:443), Ceres CLONE ID no.
1079382 (SEQ ID NO:445), Ceres CLONE ID no. 1853461 (SEQ ID
NO:447), Ceres CLONE ID no. 1626485 (SEQ ID NO:451), Ceres CLONE ID
no. 1713920 (SEQ ID NO:459), Ceres CLONE ID no. 1772747 (SEQ ID
NO:463), Ceres CLONE ID no. 225960 (SEQ ID NO:465), Public GI ID
no. 115443807 (SEQ ID NO:470), and Ceres CLONE ID no. 569388 (SEQ
ID NO:483). Other functional homologs of SEQ ID NO:442 include
Ceres CLONE ID no. 1915549 (SEQ ID NO:449), Ceres CLONE ID no.
529871 (SEQ ID NO:453), Ceres CLONE ID no. 1067079 (SEQ ID NO:455),
Ceres CLONE ID no. 1079572 (SEQ ID NO:457), Ceres ANNOT ID no.
1456550 (SEQ ID NO:461), Ceres CLONE ID no. 1437889 (SEQ ID
NO:469), Ceres CLONE ID no. 2014249 (SEQ ID NO:472), Ceres CLONE ID
no. 2033133 (SEQ ID NO:474), Ceres CLONE ID no. 707404 (SEQ ID
NO:476), Ceres CLONE ID no. 1770680 (SEQ ID NO:478), Ceres ANNOT ID
no. 1450989 (SEQ ID NO:480), Public GI ID no. 72384445 (SEQ ID
NO:481), Ceres CLONE ID no. 1059299 (SEQ ID NO:485), and Ceres
ANNOT ID no. 6008086 (SEQ ID NO:487). In some cases, a functional
homolog of SEQ ID NO:442 has an amino acid sequence with at least
40% sequence identity, e.g., 50%, 52%, 56%, 59%, 61%, 65%, 70%,
75%, 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity, to the
amino acid sequence set forth in SEQ ID NO:442.
[0076] Examples of amino acid sequences of functional homologs of
the polypeptide set forth in SEQ ID NO:489 are provided in FIG. 2
and in the sequence listing. Such functional homologs include Ceres
CLONE ID no. 571200 (SEQ ID NO:494), Ceres CLONE ID no. 1928532
(SEQ ID NO:496), Ceres ANNOT ID no. 1490637 (SEQ ID NO:498), Ceres
CLONE ID no. 295496 (SEQ ID NO:514), Public GI ID no. 115463637
(SEQ ID NO:515), and Ceres CLONE ID no. 1999444 (SEQ ID NO:518).
Other functional homologs of SEQ ID NO:489 include Public GI ID no.
147844794 (SEQ ID NO:490), Public GI ID no. 147842279 (SEQ ID
NO:491), Public GI ID no. 145355441 (SEQ ID NO:492), Ceres ANNOT ID
no. 1461381 (SEQ ID NO:500), Ceres ANNOT ID no. 1440313 (SEQ ID
NO:502), Ceres ANNOT ID no. 1448275 (SEQ ID NO:504), Ceres ANNOT ID
no. 1437838 (SEQ ID NO:506), Ceres ANNOT ID no. 1501275 (SEQ ID
NO:508), Ceres CLONE ID no. 1644562 (SEQ ID NO:510), Ceres CLONE ID
no. 1925967 (SEQ ID NO:512), Public GI ID no. 115435904 (SEQ ID
NO:516), Public GI ID no. 125552168 (SEQ ID NO:519), Public GI ID
no. 125594093 (SEQ ID NO:520), Ceres CLONE ID no. 221188 (SEQ ID
NO:522), Ceres ANNOT ID no. 1477714 (SEQ ID NO:524), Ceres CLONE ID
no. 1787953 (SEQ ID NO:526), Ceres ANNOT ID no. 1531210 (SEQ ID
NO:528), Ceres CLONE ID no. 521176 (SEQ ID NO:530), Public GI ID
no. 22327055 (SEQ ID NO:531), Ceres ANNOT ID no. 1508824 (SEQ ID
NO:533), Ceres CLONE ID no. 38879 (SEQ ID NO:535), Public GI ID no.
42569309 (SEQ ID NO:536), Ceres CLONE ID no. 1817784 (SEQ ID
NO:538), Ceres CLONE ID no. 284637 (SEQ ID NO:540), Public GI ID
no. 125596251 (SEQ ID NO:541), Public GI ID no. 125554300 (SEQ ID
NO:543), Ceres CLONE ID no. 1935437 (SEQ ID NO:545), Ceres ANNOT ID
no. 1455622 (SEQ ID NO:547), Public GI ID no. 55771354 (SEQ ID
NO:548), Ceres ANNOT ID no. 1514655 (SEQ ID NO:550), Ceres CLONE ID
no. 1848736 (SEQ ID NO:552), Public GI ID no. 125569872 (SEQ ID
NO:553), Ceres CLONE ID no. 1645078 (SEQ ID NO:555), Ceres CLONE ID
no. 1790573 (SEQ ID NO:557), Public GI ID no. 4567251 (SEQ ID
NO:558), Ceres CLONE ID no. 444113 (SEQ ID NO:561), Public GI ID
no. 125525355 (SEQ ID NO:562), Ceres ANNOT ID no. 6028854 (SEQ ID
NO:564), and Ceres ANNOT ID no. 6115356 (SEQ ID NO:566). In some
cases, a functional homolog of SEQ ID NO:489 has an amino acid
sequence with at least 40% sequence identity, e.g., 50%, 52%, 56%,
59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 97%, 98%, or 99%
sequence identity, to the amino acid sequence set forth in SEQ ID
NO:489.
[0077] Examples of amino acid sequences of functional homologs of
the polypeptide set forth in SEQ ID NO:568 are provided in FIG. 3
and in the sequence listing. Such functional homologs include Ceres
CLONE ID no. 695006 (SEQ ID NO:578) and Ceres ANNOT ID no. 1527488
(SEQ ID NO:584). Other functional homologs of SEQ ID NO:568 include
Public GI ID no. 79318519 (SEQ ID NO:569), Public GI ID no.
79318537 (SEQ ID NO:570), Ceres CLONE ID no. 956998 (SEQ ID
NO:572), Ceres CLONE ID no. 978154 (SEQ ID NO:574), Ceres CLONE ID
no. 1035628 (SEQ ID NO:576), Ceres CLONE ID no. 464169 (SEQ ID
NO:580), Ceres ANNOT ID no. 1474075 (SEQ ID NO:582), Ceres ANNOT ID
no. 1474073 (SEQ ID NO:586), and Ceres ANNOT ID no. 1527486 (SEQ ID
NO:588). In some cases, a functional homolog of SEQ ID NO:568 has
an amino acid sequence with at least 40% sequence identity, e.g.,
50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 97%,
98%, or 99% sequence identity, to the amino acid sequence set forth
in SEQ ID NO:568.
[0078] Examples of amino acid sequences of functional homologs of
the polypeptide set forth in SEQ ID NO:590 are provided in FIG. 4
and in the sequence listing. Such functional homologs include Ceres
ANNOT ID no. 1501305 (SEQ ID NO:623), Ceres CLONE ID no. 607280
(SEQ ID NO:627), Public GI ID no. 92887174 (SEQ ID NO:632), Ceres
CLONE ID no. 1857162 (SEQ ID NO:634), Public GI ID no. 115477272
(SEQ ID NO:674), and Ceres CLONE ID no. 264002 (SEQ ID NO:682).
Other functional homologs of SEQ ID NO:590 include Public GI ID no.
147822456 (SEQ ID NO:591), Public GI ID no. 144923935 (SEQ ID
NO:592), Public GI ID no. 140038730 (SEQ ID NO:593), Public GI ID
no. 78708014 (SEQ ID NO:594), Public GI ID no. 115481362 (SEQ ID
NO:595), Public GI ID no. 125531350 (SEQ ID NO:596), Public GI ID
no. 125525598 (SEQ ID NO:597), Public GI ID no. 77548630 (SEQ ID
NO:598), Public GI ID no. 46798895 (SEQ ID NO:599), Public GI ID
no. 125564653 (SEQ ID NO:600), Ceres CLONE ID no. 998865 (SEQ ID
NO:602), Public GI ID no. 125556140 (SEQ ID NO:603), Public GI ID
no. 125534482 (SEQ ID NO:604), Public GI ID no. 125550135 (SEQ ID
NO:605), Ceres CLONE ID no. 639816 (SEQ ID NO:607), Public GI ID
no. 125562170 (SEQ ID NO:608), Ceres CLONE ID no. 1797059 (SEQ ID
NO:610), Ceres CLONE ID no. 1031510 (SEQ ID NO:612), Ceres CLONE ID
no. 1973081 (SEQ ID NO:614), Public GI ID no. 125561002 (SEQ ID
NO:615), Public GI ID no. 125541732 (SEQ ID NO:616), Public GI ID
no. 125560677 (SEQ ID NO:617), Public GI ID no. 125559115 (SEQ ID
NO:618), Public GI ID no. 15809903 (SEQ ID NO:619), Public GI ID
no. 30681703 (SEQ ID NO:620), Public GI ID no. 9759556 (SEQ ID
NO:621), Ceres ANNOT ID no. 1448303 (SEQ ID NO:625), Ceres ANNOT ID
no. 1448305 (SEQ ID NO:629), Ceres ANNOT ID no. 1448307 (SEQ ID
NO:631), Public GI ID no. 92888243 (SEQ ID NO:635), Public GI ID
no. 92879395 (SEQ ID NO:636), Ceres CLONE ID no. 528876 (SEQ ID
NO:638), Ceres ANNOT ID no. 1448352 (SEQ ID NO:640), Ceres ANNOT ID
no. 1437745 (SEQ ID NO:642), Ceres ANNOT ID no. 1464146 (SEQ ID
NO:644), Ceres ANNOT ID no. 1437744 (SEQ ID NO:646), Public GI ID
no. 92894684 (SEQ ID NO:647), Ceres CLONE ID no. 3964 (SEQ ID
NO:650), Public GI ID no. 13272389 (SEQ ID NO:651), Ceres ANNOT ID
no. 1481203 (SEQ ID NO:653), Public GI ID no. 15227699 (SEQ ID
NO:654), Public GI ID no. 92886084 (SEQ ID NO:655), Public GI ID
no. 15239947 (SEQ ID NO:656), Ceres CLONE ID no. 34878 (SEQ ID
NO:658), Ceres CLONE ID no. 150484 (SEQ ID NO:660), Public GI ID
no. 21553545 (SEQ ID NO:661), Public GI ID no. 15222843 (SEQ ID
NO:662), Public GI ID no. 38230552 (SEQ ID NO:663), Public GI ID
no. 3420008 (SEQ ID NO:664), Public GI ID no. 15230602 (SEQ ID
NO:665), Public GI ID no. 3420004 (SEQ ID NO:666), Public GI ID no.
2129586 (SEQ ID NO:667), Public GI ID no. 15217849 (SEQ ID NO:668),
Public GI ID no. 15227704 (SEQ ID NO:669), Ceres ANNOT ID no.
1465750 (SEQ ID NO:671), Ceres CLONE ID no. 1983975 (SEQ ID
NO:673), Public GI ID no. 15226028 (SEQ ID NO:675), Public GI ID
no. 115459524 (SEQ ID NO:676), Ceres CLONE ID no. 1793353 (SEQ ID
NO:678), Ceres ANNOT ID no. 1467399 (SEQ ID NO:680), Ceres CLONE ID
no. 1982930 (SEQ ID NO:684), Public GI ID no. 125540700 (SEQ ID
NO:685), Ceres ANNOT ID no. 1448743 (SEQ ID NO:687), Public GI ID
no. 50251910 (SEQ ID NO:688), Ceres CLONE ID no. 1836748 (SEQ ID
NO:690), Public GI ID no. 3420006 (SEQ ID NO:691), Public GI ID no.
92879376 (SEQ ID NO:692), Ceres CLONE ID no. 838941 (SEQ ID
NO:694), Ceres ANNOT ID no. 1437746 (SEQ ID NO:696), Ceres ANNOT ID
no. 6017241 (SEQ ID NO:698), Ceres ANNOT ID no. 6085947 (SEQ ID
NO:700), and Ceres ANNOT ID no. 6017242 (SEQ ID NO:702). In some
cases, a functional homolog of SEQ ID NO:590 has an amino acid
sequence with at least 40% sequence identity, e.g., 50%, 52%, 56%,
59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 97%, 98%, or 99%
sequence identity, to the amino acid sequence set forth in SEQ ID
NO:590.
[0079] Examples of amino acid sequences of functional homologs of
the polypeptide set forth in SEQ ID NO:705 are provided in FIG. 5
and in the sequence listing. Such functional homologs include
Public GI ID no. 79320952 (SEQ ID NO:706) and Public GI ID no.
79320957 (SEQ ID NO:707). Other functional homologs of SEQ ID
NO:705 include Public GI ID no. 6692094 (SEQ ID NO:708) and Public
GI ID no. 145323049 (SEQ ID NO:709). In some cases, a functional
homolog of SEQ ID NO:705 has an amino acid sequence with at least
40% sequence identity, e.g., 50%, 52%, 56%, 59%, 61%, 65%, 70%,
75%, 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity, to the
amino acid sequence set forth in SEQ ID NO:705.
[0080] Examples of amino acid sequences of functional homologs of
the polypeptide set forth in SEQ ID NO:711 are provided in FIG. 6
and in the sequence listing. Such functional homologs include Ceres
CLONE ID no. 964616 (SEQ ID NO:714), Ceres CLONE ID no. 100009667
(SEQ ID NO:715), Ceres ANNOT ID no. 1444568 (SEQ ID NO:719), Ceres
CLONE ID no. 719489 (SEQ ID NO:725), Ceres CLONE ID no. 587748 (SEQ
ID NO:733), Public GI ID no. 125528114 (SEQ ID NO:734), and Ceres
CLONE ID no. 274172 (SEQ ID NO:737). Other functional homologs of
SEQ ID NO:711 include Public GI ID no. 144923134 (SEQ ID NO:712),
Ceres ANNOT ID no. 1471437 (SEQ ID NO:717), Ceres CLONE ID no.
1270484 (SEQ ID NO:721), Ceres CLONE ID no. 1075098 (SEQ ID
NO:723), Public GI ID no. 18412211 (SEQ ID NO:727), Ceres CLONE ID
no. 20358 (SEQ ID NO:729), Ceres CLONE ID no. 1915503 (SEQ ID
NO:731), Public GI ID no. 115440619 (SEQ ID NO:735), Public GI ID
no. 125572387 (SEQ ID NO:738), and Ceres ANNOT ID no. 6015812 (SEQ
ID NO:740). In some cases, a functional homolog of SEQ ID NO:711
has an amino acid sequence with at least 40% sequence identity,
e.g., 50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%, 95%,
97%, 98%, or 99% sequence identity, to the amino acid sequence set
forth in SEQ ID NO:711.
[0081] Examples of amino acid sequences of functional homologs of
the polypeptide set forth in SEQ ID NO:742 are provided in FIG. 7
and in the sequence listing. Such functional homologs include
Public GI ID no. 147783026 (SEQ ID NO: 743), Public GI ID no.
119367488 (SEQ ID NO: 744), Public GI ID no. 147860340 (SEQ ID NO:
745), Public GI ID no. 115477170 (SEQ ID NO: 757), Ceres CLONE ID
no. 1620215 (SEQ ID NO: 759), Ceres CLONE ID no. 1931889 (SEQ ID
NO: 761), Ceres CLONE ID no. 1798756 (SEQ ID NO: 763), Ceres CLONE
ID no. 1918424 (SEQ ID NO: 767), Ceres CLONE ID no. 1845154 (SEQ ID
NO: 771), Public GI ID no. 38016527 (SEQ ID NO: 796), Ceres Clone
ID no. 1084216 (SEQ ID NO: 797), Public GI ID no. 113196593 (SEQ ID
NO: 798), Public GI ID no. 75133829 (SEQ ID NO: 799), Ceres ANNOT
ID no. 1460527 (SEQ ID NO: 801), Ceres Annot ID no. 8644540 (SEQ ID
NO: 805), Ceres ANNOT ID no. 1450673 (SEQ ID NO: 807), Public GI ID
no. 116778802 (SEQ ID NO: 808), Public GI ID no. 116778893 (SEQ ID
NO: 809), Public GI ID no. 116778998 (SEQ ID NO: 810), Public GI ID
no. 157849766 (SEQ ID NO: 811), Public GI ID no. 159474166 (SEQ ID
NO: 812), Public GI ID no. 168036656 (SEQ ID NO: 813), Ceres ANNOT
ID no. 1456578 (SEQ ID NO: 815), Public GI ID no. 168053490 (SEQ ID
NO: 816), Public GI ID no. 193237563 (SEQ ID NO: 817), Ceres Clone
ID no. 100879386 (SEQ ID NO: 819), Ceres Clone ID no. 2055733 (SEQ
ID NO: 823), Ceres Clone ID no. 2056478 (SEQ ID NO: 825), Ceres
CLONE ID no. 13007 (SEQ ID NO: 832), Ceres CLONE ID no. 5522 (SEQ
ID NO: 834), Ceres CLONE ID no. 30543 (SEQ ID NO: 836), Ceres CLONE
ID no. 14203 (SEQ ID NO: 839), Ceres CLONE ID no. 975913 (SEQ ID
NO: 846), Ceres CLONE ID no. 967417 (SEQ ID NO: 848), Public GI ID
no. 119720772 (SEQ ID NO: 857), Ceres CLONE ID no. 708446 (SEQ ID
NO: 859), Ceres CLONE ID no. 1614593 (SEQ ID NO: 863), Ceres CLONE
ID no. 2025938 (SEQ ID NO: 885), Public GI ID no. 92896423 (SEQ ID
NO: 892), Ceres CLONE ID no. 634261 (SEQ ID NO: 900), Ceres CLONE
ID no. 1387149 (SEQ ID NO: 912), Ceres CLONE ID no. 1423851 (SEQ ID
NO: 918), Ceres CLONE ID no. 1589047 (SEQ ID NO: 939), Ceres CLONE
ID no. 1748922 (SEQ ID NO: 944), Public GI ID no. 5031281 (SEQ ID
NO: 953), Ceres CLONE ID no. 1775820 (SEQ ID NO: 955), Ceres CLONE
ID no. 1787151 (SEQ ID NO: 957), Ceres CLONE ID no. 1765871 (SEQ ID
NO: 973), Ceres CLONE ID no. 1990071 (SEQ ID NO: 977), Public GI ID
no. 35187687 (SEQ ID NO: 988), Public GI ID no. 125556051 (SEQ ID
NO: 989), Public GI ID no. 125561658 (SEQ ID NO: 990), Public GI ID
no. 115468934 (SEQ ID NO: 991), Public GI ID no. 115470773 (SEQ ID
NO: 992), Public GI ID no. 115444813 (SEQ ID NO: 993), Public GI ID
no. 112819496 (SEQ ID NO: 999), Public GI ID no. 118424243 (SEQ ID
NO: 1000), Ceres ANNOT ID no. 6063957 (SEQ ID NO: 1010), Public GI
ID no. 169363 (SEQ ID NO:1215), Ceres CLONE ID no. 24255 (SEQ ID
NO: 1242), Ceres Clone ID no. 1030374 (SEQ ID NO: 1306), Ceres
Clone ID no. 1738028 (SEQ ID NO: 1308), Public GI ID no. 115455855
(SEQ ID NO: 1309), and Public GI ID no. 193237563 (SEQ ID NO:
1310). In some cases, a functional homolog of SEQ ID NO:742 has an
amino acid sequence with at least 40% sequence identity, e.g., 50%,
52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 97%, 98%, or
99% sequence identity, to the amino acid sequence set forth in SEQ
ID NO:742.
[0082] Examples of amino acid sequences of functional homologs of
the polypeptide set forth in SEQ ID NO:1012 are provided in FIG. 8
and in the sequence listing. Such functional homologs include Ceres
CLONE ID no. 287430 (SEQ ID NO:1018), Ceres ANNOT ID no. 451889
(SEQ ID NO:1020), Ceres CLONE ID no. 936084 (SEQ ID NO:1055), Ceres
CLONE ID no. 1792501 (SEQ ID NO:1066), Ceres ANNOT ID no. 1437875
(SEQ ID NO:1068), Public GI ID no. 1853968 (SEQ ID NO:1093), Public
GI ID no. 27530032 (SEQ ID NO:1094), Ceres CLONE ID no. 1834483
(SEQ ID NO:1110), Public GI ID no. 84579418 (SEQ ID NO:1113),
Public GI ID no. 15077030 (SEQ ID NO:1117), and Public GI ID no.
13752458 (SEQ ID NO:1118). Other functional homologs of SEQ ID
NO:1012 include Public GI ID no. 147865685 (SEQ ID NO:1013), Public
GI ID no. 119503368 (SEQ ID NO:1014), Public GI ID no. 72161874
(SEQ ID NO:1015), Public GI ID no. 91780661 (SEQ ID NO:1016), Ceres
ANNOT ID no. 835908 (SEQ ID NO:1022), Public GI ID no. 38326750
(SEQ ID NO:1023), Ceres CLONE ID no. 1939396 (SEQ ID NO:1025),
Ceres CLONE ID no. 403637 (SEQ ID NO:1027), Public GI ID no.
42539907 (SEQ ID NO:1028), Ceres CLONE ID no. 1836494 (SEQ ID
NO:1030), Public GI ID no. 15192945 (SEQ ID NO:1031), Ceres CLONE
ID no. 1607947 (SEQ ID NO:1033), Ceres CLONE ID no. 115880 (SEQ ID
NO:1035), Ceres CLONE ID no. 1074009 (SEQ ID NO:1037), Ceres CLONE
ID no. 476073 (SEQ ID NO:1039), Ceres CLONE ID no. 554053 (SEQ ID
NO:1041), Ceres CLONE ID no. 391449 (SEQ ID NO:1045), Ceres CLONE
ID no. 1846400 (SEQ ID NO:1048), Ceres ANNOT ID no. 1475185 (SEQ ID
NO:1050), Public GI ID no. 115457148 (SEQ ID NO:1051), Ceres ANNOT
ID no. 1454960 (SEQ ID NO:1053), Public GI ID no. 50346893 (SEQ ID
NO:1056), Ceres CLONE ID no. 1931526 (SEQ ID NO:1058), Ceres ANNOT
ID no. 1454260 (SEQ ID NO:1061), Public GI ID no. 115456131 (SEQ ID
NO:1062), Ceres CLONE ID no. 159151 (SEQ ID NO:1064), Ceres CLONE
ID no. 1842801 (SEQ ID NO:1071), Ceres CLONE ID no. 533030 (SEQ ID
NO:1073), Ceres CLONE ID no. 1931881 (SEQ ID NO:1075), Ceres ANNOT
ID no. 1480006 (SEQ ID NO:1077), Ceres CLONE ID no. 1895007 (SEQ ID
NO:1079), Public GI ID no. 3598863 (SEQ ID NO:1080), Ceres ANNOT ID
no. 1471735 (SEQ ID NO:1082), Ceres CLONE ID no. 1937530 (SEQ ID
NO:1084), Ceres CLONE ID no. 1833050 (SEQ ID NO:1086), Ceres CLONE
ID no. 644213 (SEQ ID NO:1088), Ceres CLONE ID no. 568154 (SEQ ID
NO:1090), Ceres CLONE ID no. 527598 (SEQ ID NO:1092), Ceres ANNOT
ID no. 1487614 (SEQ ID NO:1096), Ceres ANNOT ID no. 1541881 (SEQ ID
NO:1098), Ceres ANNOT ID no. 1488465 (SEQ ID NO:1100), Ceres ANNOT
ID no. 1488468 (SEQ ID NO:1102), Ceres ANNOT ID no. 1541884 (SEQ ID
NO:1104), Ceres ANNOT ID no. 1454972 (SEQ ID NO:1106), Ceres ANNOT
ID no. 1475186 (SEQ ID NO:1108), Ceres CLONE ID no. 686198 (SEQ ID
NO:1112), Public GI ID no. 84579420 (SEQ ID NO:1114), Ceres CLONE
ID no. 605144 (SEQ ID NO:1116), Public GI ID no. 15077028 (SEQ ID
NO:1119), Ceres ANNOT ID no. 1454963 (SEQ ID NO:1121), Ceres ANNOT
ID no. 6030208 (SEQ ID NO:1123), and Ceres ANNOT ID no. 6076767
(SEQ ID NO:1125). In some cases, a functional homolog of SEQ ID
NO:1012 has an amino acid sequence with at least 40% sequence
identity, e.g., 50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%,
90%, 95%, 97%, 98%, or 99% sequence identity, to the amino acid
sequence set forth in SEQ ID NO:1012.
[0083] Examples of amino acid sequences of functional homologs of
the polypeptide set forth in SEQ ID NO:1129 are provided in FIG. 9
and in the sequence listing. Such functional homologs include
Public GI ID no. 72255610 (SEQ ID NO:933), Ceres SEEDLINE ID no.
ME20095 (SEQ ID NO:1129), Ceres CLONE ID no. 1562633 (SEQ ID
NO:1140), Ceres ANNOT ID no. 1692728 (SEQ ID NO:1142), Ceres CLONE
ID no. 921110 (SEQ ID NO:1144), Ceres CLONE ID no. 1791180 (SEQ ID
NO:1146), Ceres CLONE ID no. 527891 (SEQ ID NO:1148), Ceres ANNOT
ID no. 1522414 (SEQ ID NO:1150), and Public GI ID no. 51458330 (SEQ
ID NO:1182). Other functional homologs of SEQ ID NO:1129 include
Public GI ID no. 18399940 (SEQ ID NO:63), Ceres ANNOT ID no.
6107929 (SEQ ID NO:73), Ceres ANNOT ID no. 1473094 (SEQ ID NO:96),
Ceres ANNOT ID no. 1503955 (SEQ ID NO:127), Ceres ANNOT ID no.
1526501 (SEQ ID NO:139), Public GI ID no. 15235713 (SEQ ID NO:179),
Public GI ID no. 15234552 (SEQ ID NO:180), Ceres ANNOT ID no.
1520801 (SEQ ID NO:209), Ceres ANNOT ID no. 1443434 (SEQ ID
NO:467), Public GI ID no. 30696058 (SEQ ID NO:559), Ceres ANNOT ID
no. 1509601 (SEQ ID NO:648), Ceres ANNOT ID no. 1451912 (SEQ ID
NO:726), Ceres ANNOT ID no. 6017545 (SEQ ID NO:741), Ceres ANNOT ID
no. 1504999 (SEQ ID NO:765), Ceres ANNOT ID no. 1456402 (SEQ ID
NO:773), Ceres ANNOT ID no. 1496359 (SEQ ID NO:781), Ceres ANNOT ID
no. 1447260 (SEQ ID NO:842), Ceres ANNOT ID no. 1469023 (SEQ ID
NO:861), Ceres ANNOT ID no. 1474186 (SEQ ID NO:865), Ceres ANNOT ID
no. 1527596 (SEQ ID NO:867), Ceres CLONE ID no. 1919624 (SEQ ID
NO:896), Ceres CLONE ID no. 677797 (SEQ ID NO:916), Public GI ID
no. 7960729 (SEQ ID NO:932), Ceres CLONE ID no. 941845 (SEQ ID
NO:1043), Public GI ID no. 30683008 (SEQ ID NO:1046), Public GI ID
no. 42568786 (SEQ ID NO:1059), Ceres CLONE ID no. 255364 (SEQ ID
NO:1069), Public GI ID no. 147771549 (SEQ ID NO:1130), Public GI ID
no. 144924741 (SEQ ID NO:1131), Public GI ID no. 121594508 (SEQ ID
NO:1132), Public GI ID no. 91202290 (SEQ ID NO:1133), Public GI ID
no. 89900939 (SEQ ID NO:1134), Public GI ID no. 77917672 (SEQ ID
NO:1135), Public GI ID no. 77165252 (SEQ ID NO:1136), Public GI ID
no. 115375374 (SEQ ID NO:1137), Public GI ID no. 110602449 (SEQ ID
NO:1138), Public GI ID no. 125540573 (SEQ ID NO:1151), Public GI ID
no. 125561519 (SEQ ID NO:1152), Public GI ID no. 125528345 (SEQ ID
NO:1153), Public GI ID no. 116310408 (SEQ ID NO:1154), Public GI ID
no. 115483332 (SEQ ID NO:1156), Public GI ID no. 125575640 (SEQ ID
NO:1157), Public GI ID no. 115476368 (SEQ ID NO:1158), Public GI ID
no. 115478841 (SEQ ID NO:1159), Public GI ID no. 115467158 (SEQ ID
NO:1160), Public GI ID no. 72384477 (SEQ ID NO:1161), Public GI ID
no. 116054703 (SEQ ID NO:1162), Ceres CLONE ID no. 1786317 (SEQ ID
NO:1164), Ceres CLONE ID no. 1791336 (SEQ ID NO:1166), Ceres CLONE
ID no. 1877752 (SEQ ID NO:1168), Ceres CLONE ID no. 1821191 (SEQ ID
NO:1170), Ceres CLONE ID no. 446838 (SEQ ID NO:1172), Ceres CLONE
ID no. 1556915 (SEQ ID NO:1174), Public GI ID no. 90200725 (SEQ ID
NO:1175), Ceres CLONE ID no. 1580247 (SEQ ID NO:1177), Ceres CLONE
ID no. 644201 (SEQ ID NO:1179), Ceres CLONE ID no. 538689 (SEQ ID
NO:1181), Ceres CLONE ID no. 817225 (SEQ ID NO:1184), and Ceres
CLONE ID no. 923842 (SEQ ID NO:1186). In some cases, a functional
homolog of SEQ ID NO:1129 has an amino acid sequence with at least
40% sequence identity, e.g., 50%, 52%, 56%, 59%, 61%, 65%, 70%,
75%, 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity, to the
amino acid sequence set forth in SEQ ID NO:1129.
[0084] Examples of amino acid sequences of functional homologs of
the polypeptide set forth in SEQ ID NO:60 are provided in FIG. 10
and in the sequence listing. Such functional homologs include Ceres
CLONE ID no. 872030 (SEQ ID NO:62), Ceres CLONE ID no. 100029223
(SEQ ID NO:74), Ceres CLONE ID no. 1939845 (SEQ ID NO:98), Ceres
CLONE ID no. 2015383 (SEQ ID NO:102), Ceres CLONE ID no. 1607893
(SEQ ID NO:104), Ceres CLONE ID no. 1075133 (SEQ ID NO:152), and
Ceres CLONE ID no. 1218065 (SEQ ID NO:915). Other functional
homologs of SEQ ID NO:60 include Public GI ID no. 15227919 (SEQ ID
NO:64), Public GI ID no. 9843641 (SEQ ID NO:65), Public GI ID no.
124294789 (SEQ ID NO:66), Public GI ID no. 15233446 (SEQ ID NO:67),
Public GI ID no. 115477679 (SEQ ID NO:68), Public GI ID no.
42407552 (SEQ ID NO:69), Public GI ID no. 125562450 (SEQ ID NO:70),
Public GI ID no. 115469980 (SEQ ID NO:71), Ceres ANNOT ID no.
1442539 (SEQ ID NO:76), Ceres ANNOT ID no. 1442538 (SEQ ID NO:78),
Ceres ANNOT ID no. 1460661 (SEQ ID NO:80), Ceres ANNOT ID no.
1452884 (SEQ ID NO:82), Ceres ANNOT ID no. 1450523 (SEQ ID NO:84),
Ceres ANNOT ID no. 1442257 (SEQ ID NO:86), Ceres ANNOT ID no.
1541121 (SEQ ID NO:88), Ceres ANNOT ID no. 1480481 (SEQ ID NO:90),
Ceres ANNOT ID no. 1487713 (SEQ ID NO:92), Ceres ANNOT ID no.
1456204 (SEQ ID NO:94), Ceres CLONE ID no. 1387402 (SEQ ID NO:100),
Ceres CLONE ID no. 972919 (SEQ ID NO:106), Ceres CLONE ID no.
1645860 (SEQ ID NO:108), Ceres CLONE ID no. 1042804 (SEQ ID
NO:110), Ceres CLONE ID no. 1606678 (SEQ ID NO:112), Ceres CLONE ID
no. 684496 (SEQ ID NO:114), Ceres CLONE ID no. 1062366 (SEQ ID
NO:116), Ceres CLONE ID no. 1722931 (SEQ ID NO:118), Ceres CLONE ID
no. 746169 (SEQ ID NO:120), Ceres CLONE ID no. 1050475 (SEQ ID
NO:122), Ceres CLONE ID no. 1728904 (SEQ ID NO:124), Ceres CLONE ID
no. 1698538 (SEQ ID NO:126), Ceres CLONE ID no. 1762328 (SEQ ID
NO:130), Ceres CLONE ID no. 1590072 (SEQ ID NO:132), Ceres CLONE ID
no. 471593 (SEQ ID NO:134), Ceres CLONE ID no. 1031667 (SEQ ID
NO:136), Ceres CLONE ID no. 1826085 (SEQ ID NO:138), Ceres CLONE ID
no. 373932 (SEQ ID NO:142), Ceres CLONE ID no. 1797836 (SEQ ID
NO:144), Ceres CLONE ID no. 1102774 (SEQ ID NO:146), Ceres CLONE ID
no. 842287 (SEQ ID NO:148), Ceres CLONE ID no. 568205 (SEQ ID
NO:150), Ceres ANNOT ID no. 6108997 (SEQ ID NO:154), Ceres ANNOT ID
no. 6023609 (SEQ ID NO:156), and Ceres ANNOT ID no. 6017906 (SEQ ID
NO:158). In some cases, a functional homolog of SEQ ID NO:60 has an
amino acid sequence with at least 40% sequence identity, e.g., 50%,
52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 97%, 98%, or
99% sequence identity, to the amino acid sequence set forth in SEQ
ID NO:60.
[0085] Examples of amino acid sequences of functional homologs of
the polypeptide set forth in SEQ ID NO:364 are provided in FIG. 11
and in the sequence listing. Such functional homologs include Ceres
ANNOT ID no. 1455221 (SEQ ID NO:407), Public GI ID no. 729774 (SEQ
ID NO:408), Ceres CLONE ID no. 1414288 (SEQ ID NO:410), Public GI
ID no. 115482048 (SEQ ID NO:413), and Public GI ID no. 56117815
(SEQ ID NO:421). Other functional homologs of SEQ ID NO:364 include
Ceres ANNOT ID no. 1485538 (SEQ ID NO:366), Ceres ANNOT ID no.
1538505 (SEQ ID NO:368), Public GI ID no. 8347238 (SEQ ID NO:369),
Public GI ID no. 125542955 (SEQ ID NO:370), Public GI ID no.
115446219 (SEQ ID NO:371), Ceres CLONE ID no. 1597127 (SEQ ID
NO:373), Ceres CLONE ID no. 1584568 (SEQ ID NO:375), Ceres CLONE ID
no. 286056 (SEQ ID NO:377), Ceres ANNOT ID no. 1543432 (SEQ ID
NO:379), Ceres CLONE ID no. 1728062 (SEQ ID NO:381), Ceres ANNOT ID
no. 1448950 (SEQ ID NO:383), Ceres CLONE ID no. 771500 (SEQ ID
NO:385), Ceres ANNOT ID no. 1485096 (SEQ ID NO:387), Ceres CLONE ID
no. 1646104 (SEQ ID NO:389), Public GI ID no. 111184724 (SEQ ID
NO:390), Ceres CLONE ID no. 1362475 (SEQ ID NO:392), Ceres CLONE ID
no. 597906 (SEQ ID NO:394), Ceres CLONE ID no. 615781 (SEQ ID
NO:396), Ceres CLONE ID no. 538713 (SEQ ID NO:398), Ceres CLONE ID
no. 1794141 (SEQ ID NO:400), Public GI ID no. 5821138 (SEQ ID
NO:401), Public GI ID no. 886742 (SEQ ID NO:402), Ceres CLONE ID
no. 1814498 (SEQ ID NO:404), Public GI ID no. 7158882 (SEQ ID
NO:405), Ceres ANNOT ID no. 1452564 (SEQ ID NO:412), Public GI ID
no. 125557431 (SEQ ID NO:414), Ceres ANNOT ID no. 1442880 (SEQ ID
NO:416), Ceres ANNOT ID no. 1463437 (SEQ ID NO:418), Public GI ID
no. 42415865 (SEQ ID NO:419), Public GI ID no. 33087081 (SEQ ID
NO:420), Public GI ID no. 115521217 (SEQ ID NO:422), Public GI ID
no. 115521211 (SEQ ID NO:423), Public GI ID no. 115521215 (SEQ ID
NO:424), Ceres ANNOT ID no. 1454376 (SEQ ID NO:426), Ceres CLONE ID
no. 835571 (SEQ ID NO:428), Ceres ANNOT ID no. 1500046 (SEQ ID
NO:430), Public GI ID no. 110738569 (SEQ ID NO:431), Public GI ID
no. 125542510 (SEQ ID NO:432), Public GI ID no. 30686034 (SEQ ID
NO:433), Public GI ID no. 125570883 (SEQ ID NO:434), Public GI ID
no. 42562463 (SEQ ID NO:435), Ceres CLONE ID no. 100043265 (SEQ ID
NO:436), Ceres ANNOT ID no. 6089790 (SEQ ID NO:438), and Ceres
ANNOT ID no. 6043635 (SEQ ID NO:440). In some cases, a functional
homolog of SEQ ID NO:364 has an amino acid sequence with at least
40% sequence identity, e.g., 50%, 52%, 56%, 59%, 61%, 65%, 70%,
75%, 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity, to the
amino acid sequence set forth in SEQ ID NO:364.
[0086] Examples of amino acid sequences of functional homologs of
the polypeptide set forth in SEQ ID NO:311 are provided in FIG. 12
and in the sequence listing. Such functional homologs include Ceres
ANNOT ID no. 1538958 (SEQ ID NO:319), Public GI ID no. 115521213
(SEQ ID NO:320), Public GI ID no. 42415865 (SEQ ID NO:321), Public
GI ID no. 729775 (SEQ ID NO:322), Public GI ID no. 11386827 (SEQ ID
NO:323), Public GI ID no. 115456675 (SEQ ID NO:326), and Public GI
ID no. 89274218 (SEQ ID NO:335). Other functional homologs of SEQ
ID NO:311 include Public GI ID no. 125546035 (SEQ ID NO:312), Ceres
CLONE ID no. 125228 (SEQ ID NO:314), Public GI ID no. 115465055
(SEQ ID NO:315), Public GI ID no. 125553080 (SEQ ID NO:316), Public
GI ID no. 8920606 (SEQ ID NO:317), Ceres CLONE ID no. 1919643 (SEQ
ID NO:325), Public GI ID no. 429155 (SEQ ID NO:327), Ceres CLONE ID
no. 1194199 (SEQ ID NO:329), Public GI ID no. 15236631 (SEQ ID
NO:330), Ceres ANNOT ID no. 1523420 (SEQ ID NO:332), Ceres ANNOT ID
no. 1447046 (SEQ ID NO:334), Public GI ID no. 25296101 (SEQ ID
NO:336), Public GI ID no. 15228440 (SEQ ID NO:337), Public GI ID
no. 6624612 (SEQ ID NO:338), Public GI ID no. 125588688 (SEQ ID
NO:339), Public GI ID no. 115470859 (SEQ ID NO:340), Public GI ID
no. 15228865 (SEQ ID NO:341), Ceres CLONE ID no. 1571069 (SEQ ID
NO:343), Public GI ID no. 115455903 (SEQ ID NO:344), Ceres ANNOT ID
no. 1459422 (SEQ ID NO:346), Ceres CLONE ID no. 1577511 (SEQ ID
NO:348), Public GI ID no. 115451657 (SEQ ID NO:349), Public GI ID
no. 15225255 (SEQ ID NO:350), Public GI ID no. 125545695 (SEQ ID
NO:351), Ceres ANNOT ID no. 1471817 (SEQ ID NO:353), Ceres ANNOT ID
no. 1440896 (SEQ ID NO:355), Public GI ID no. 119393868 (SEQ ID
NO:356), Ceres ANNOT ID no. 1517025 (SEQ ID NO:358), Ceres ANNOT ID
no. 1512410 (SEQ ID NO:360), and Ceres ANNOT ID no. 6035498 (SEQ ID
NO:362). In some cases, a functional homolog of SEQ ID NO:311 has
an amino acid sequence with at least 40% sequence identity, e.g.,
50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 97%,
98%, or 99% sequence identity, to the amino acid sequence set forth
in SEQ ID NO:311.
[0087] Examples of amino acid sequences of functional homologs of
the polypeptide set forth in SEQ ID NO:160 are provided in FIG. 13
and in the sequence listing. Such functional homologs include Ceres
CLONE ID no. 1837065 (SEQ ID NO:162), Ceres ANNOT ID no. 1531178
(SEQ ID NO:184), Ceres CLONE ID no. 470694 (SEQ ID NO:242), Public
GI ID no. 92867368 (SEQ ID NO:253), Ceres CLONE ID no. 859707 (SEQ
ID NO:256), Ceres CLONE ID no. 392275 (SEQ ID NO:268), Ceres CLONE
ID no. 1828394 (SEQ ID NO:282), and Public GI ID no. 115466694 (SEQ
ID NO:294). Other functional homologs of SEQ ID NO:160 include
Ceres CLONE ID no. 1895763 (SEQ ID NO:164), Ceres CLONE ID no.
1850460 (SEQ ID NO:166), Ceres CLONE ID no. 1808074 (SEQ ID
NO:168), Ceres CLONE ID no. 1832882 (SEQ ID NO:170), Ceres CLONE ID
no. 1978977 (SEQ ID NO:172), Ceres CLONE ID no. 1918742 (SEQ ID
NO:174), Ceres CLONE ID no. 1940023 (SEQ ID NO:176), Ceres CLONE ID
no. 1923495 (SEQ ID NO:178), Ceres CLONE ID no. 1851388 (SEQ ID
NO:182), Ceres ANNOT ID no. 1477681 (SEQ ID NO:186), Ceres ANNOT ID
no. 1508800 (SEQ ID NO:188), Ceres ANNOT ID no. 1455597 (SEQ ID
NO:190), Ceres ANNOT ID no. 1532016 (SEQ ID NO:192), Ceres ANNOT ID
no. 1478526 (SEQ ID NO:194), Ceres ANNOT ID no. 1492567 (SEQ ID
NO:196), Ceres ANNOT ID no. 1515273 (SEQ ID NO:198), Ceres ANNOT ID
no. 1461979 (SEQ ID NO:200), Ceres ANNOT ID no. 1439702 (SEQ ID
NO:202), Ceres ANNOT ID no. 1448044 (SEQ ID NO:204), Ceres ANNOT ID
no. 1483370 (SEQ ID NO:206), Ceres ANNOT ID no. 1445651 (SEQ ID
NO:208), Ceres ANNOT ID no. 1490863 (SEQ ID NO:212), Ceres ANNOT ID
no. 1501043 (SEQ ID NO:214), Ceres ANNOT ID no. 1527507 (SEQ ID
NO:216), Ceres ANNOT ID no. 1498633 (SEQ ID NO:218), Ceres ANNOT ID
no. 1465403 (SEQ ID NO:220), Ceres ANNOT ID no. 1510956 (SEQ ID
NO:222), Ceres ANNOT ID no. 1457735 (SEQ ID NO:224), Public GI ID
no. 15224711 (SEQ ID NO:225), Ceres CLONE ID no. 40062 (SEQ ID
NO:228), Public GI ID no. 15220315 (SEQ ID NO:229), Ceres CLONE ID
no. 36480 (SEQ ID NO:231), Ceres CLONE ID no. 2443 (SEQ ID NO:233),
Public GI ID no. 30685375 (SEQ ID NO:234), Public GI ID no.
126009434 (SEQ ID NO:235), Ceres CLONE ID no. 965753 (SEQ ID
NO:237), Ceres CLONE ID no. 948834 (SEQ ID NO:239), Public GI ID
no. 33347409 (SEQ ID NO:240), Ceres CLONE ID no. 643614 (SEQ ID
NO:244), Ceres CLONE ID no. 1049362 (SEQ ID NO:246), Ceres CLONE ID
no. 546675 (SEQ ID NO:248), Ceres CLONE ID no. 522046 (SEQ ID
NO:250), Ceres CLONE ID no. 651076 (SEQ ID NO:252), Public GI ID
no. 92897590 (SEQ ID NO:254), Ceres CLONE ID no. 634176 (SEQ ID
NO:258), Ceres CLONE ID no. 1041432 (SEQ ID NO:260), Ceres CLONE ID
no. 1065203 (SEQ ID NO:262), Ceres CLONE ID no. 555361 (SEQ ID
NO:264), Ceres CLONE ID no. 704227 (SEQ ID NO:266), Ceres CLONE ID
no. 100819945 (SEQ ID NO:269), Ceres CLONE ID no. 282584 (SEQ ID
NO:271), Ceres CLONE ID no. 1448469 (SEQ ID NO:273), Ceres CLONE ID
no. 241246 (SEQ ID NO:275), Ceres CLONE ID no. 234443 (SEQ ID
NO:277), Ceres CLONE ID no. 293549 (SEQ ID NO:279), Public GI ID
no. 109450926 (SEQ ID NO:280), Ceres CLONE ID no. 1803923 (SEQ ID
NO:284), Ceres CLONE ID no. 2024557 (SEQ ID NO:286), Ceres CLONE ID
no. 1820364 (SEQ ID NO:288), Public GI ID no. 125554274 (SEQ ID
NO:289), Public GI ID no. 125531165 (SEQ ID NO:290), Public GI ID
no. 125541514 (SEQ ID NO:291), Public GI ID no. 125557559 (SEQ ID
NO:292), Public GI ID no. 125557844 (SEQ ID NO:293), Public GI ID
no. 115449295 (SEQ ID NO:295), Public GI ID no. 115481182 (SEQ ID
NO:296), Public GI ID no. 20177639 (SEQ ID NO:297), Public GI ID
no. 115471021 (SEQ ID NO:298), Public GI ID no. 115456011 (SEQ ID
NO:299), Public GI ID no. 125588290 (SEQ ID NO:300), Public GI ID
no. 115477799 (SEQ ID NO:301), Public GI ID no. 115471385 (SEQ ID
NO:302), Public GI ID no. 125599705 (SEQ ID NO:303), Public GI ID
no. 115489810 (SEQ ID NO:304), Public GI ID no. 55276718 (SEQ ID
NO:305), Ceres ANNOT ID no. 6025568 (SEQ ID NO:307), and Ceres
ANNOT ID no. 6091128 (SEQ ID NO:309). In some cases, a functional
homolog of SEQ ID NO:160 has an amino acid sequence with at least
40% sequence identity, e.g., 50%, 52%, 56%, 59%, 61%, 65%, 70%,
75%, 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity, to the
amino acid sequence set forth in SEQ ID NO:160.
[0088] Examples of amino acid sequences of functional homologs of
the polypeptide set forth in SEQ ID NO:2 are provided in FIG. 14
and in the sequence listing. Such functional homologs include
Public GI ID no. 115470807 (SEQ ID NO:5), Ceres CLONE ID no.
1842931 (SEQ ID NO:15), Ceres CLONE ID no. 321308 (SEQ ID NO:17),
Ceres CLONE ID no. 1725811 (SEQ ID NO:19), Ceres CLONE ID no.
1357455 (SEQ ID NO:21), Ceres CLONE ID no. 943370 (SEQ ID NO:23),
Ceres CLONE ID no. 1327712 (SEQ ID NO:25), Ceres CLONE ID no.
1764692 (SEQ ID NO:27), and Ceres ANNOT ID no. 1512656 (SEQ ID
NO:41). Other functional homologs of SEQ ID NO:2 include Public GI
ID no. 18403425 (SEQ ID NO:3), Public GI ID no. 11994285 (SEQ ID
NO:4), Public GI ID no. 108706298 (SEQ ID NO:6), Public GI ID no.
15233585 (SEQ ID NO:7), Public GI ID no. 125599279 (SEQ ID NO:8),
Public GI ID no. 115450865 (SEQ ID NO:9), Public GI ID no. 22773244
(SEQ ID NO:10), Public GI ID no. 92874749 (SEQ ID NO:11), Ceres
CLONE ID no. 1857034 (SEQ ID NO:13), Ceres CLONE ID no. 1059300
(SEQ ID NO:29), Ceres CLONE ID no. 1804869 (SEQ ID NO:31), Ceres
CLONE ID no. 378863 (SEQ ID NO:33), Ceres CLONE ID no. 1930271 (SEQ
ID NO:35), Ceres CLONE ID no. 2034728 (SEQ ID NO:37), Ceres CLONE
ID no. 1866421 (SEQ ID NO:39), Ceres ANNOT ID no. 1463492 (SEQ ID
NO:43), Ceres ANNOT ID no. 1459391 (SEQ ID NO:45), Ceres ANNOT ID
no. 1442920 (SEQ ID NO:47), Ceres ANNOT ID no. 6011447 (SEQ ID
NO:49), Ceres ANNOT ID no. 6068218 (SEQ ID NO:51), Ceres ANNOT ID
no. 6044121 (SEQ ID NO:53), and Ceres ANNOT ID no. 6100755 (SEQ ID
NO:55). In some cases, a functional homolog of SEQ ID NO:2 has an
amino acid sequence with at least 40% sequence identity, e.g., 50%,
52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 97%, 98%, or
99% sequence identity, to the amino acid sequence set forth in SEQ
ID NO:2.
[0089] Examples of amino acid sequences of functional homologs of
the polypeptide set forth in SEQ ID NO:749 are provided in FIG. 15
and in the sequence listing. Such functional homologs include Ceres
CLONE ID no. 1847516 (SEQ ID NO:1195), Ceres CLONE ID no. 1961986
(SEQ ID NO:1201), and Ceres ANNOT ID no. 6091930 (SEQ ID NO:1207).
Other functional homologs of SEQ ID NO:749 include Ceres CLONE ID
no. 2025938 (SEQ ID NO:885), Ceres CLONE ID no. 1821214 (SEQ ID
NO:1189), Ceres CLONE ID no. 1040399 (SEQ ID NO:1191), Ceres CLONE
ID no. 1093691 (SEQ ID NO:1193), Ceres CLONE ID no. 974539 (SEQ ID
NO:1197), Ceres CLONE ID no. 1832340 (SEQ ID NO:1199), Ceres CLONE
ID no. 1933211 (SEQ ID NO:1203), Ceres CLONE ID no. 997558 (SEQ ID
NO:1205), Ceres ANNOT ID no. 6041596 (SEQ ID NO:1209), Public GI ID
no. 125564176 (SEQ ID NO:1210), Ceres CLONE ID no. 1836064 (SEQ ID
NO:1212), Ceres CLONE ID no. 1909693 (SEQ ID NO:1214), Public GI ID
no. 40287480 (SEQ ID NO:1216), Ceres CLONE ID no. 1765346 (SEQ ID
NO:1218), Public GI ID no. 125546008 (SEQ ID NO:1219), Ceres CLONE
ID no. 1950900 (SEQ ID NO:1221), Public GI ID no. 41350259 (SEQ ID
NO:1222), Public GI ID no. 125588210 (SEQ ID NO:1223), Ceres CLONE
ID no. 1954395 (SEQ ID NO:1225), Public GI ID no. 18403408 (SEQ ID
NO:1226), Ceres CLONE ID no. 2010121 (SEQ ID NO:1228), Ceres ANNOT
ID no. 6011486 (SEQ ID NO:1230), Public GI ID no. 25082726 (SEQ ID
NO:1231), Public GI ID no. 113196593 (SEQ ID NO:1232), Ceres CLONE
ID no. 1843021 (SEQ ID NO:1234), Ceres CLONE ID no. 1931194 (SEQ ID
NO:1236), Ceres CLONE ID no. 1652996 (SEQ ID NO:1238), Ceres CLONE
ID no. 1930044 (SEQ ID NO:1240), Ceres CLONE ID no. 24255 (SEQ ID
NO:1242), Ceres ANNOT ID no. 6034955 (SEQ ID NO:1244), Ceres ANNOT
ID no. 6119444 (SEQ ID NO:1246), Ceres ANNOT ID no. 6063956 (SEQ ID
NO:1248), Ceres ANNOT ID no. 6015461 (SEQ ID NO:1250), Ceres CLONE
ID no. 696244 (SEQ ID NO:1252), Ceres ANNOT ID no. 1468973 (SEQ ID
NO:1254), Ceres CLONE ID no. 2019529 (SEQ ID NO:1256), and Ceres
CLONE ID no. 1492169 (SEQ ID NO:1258). In some cases, a functional
homolog of SEQ ID NO:749 has an amino acid sequence with at least
40% sequence identity, e.g., 50%, 52%, 56%, 59%, 61%, 65%, 70%,
75%, 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity, to the
amino acid sequence set forth in SEQ ID NO:749.
[0090] Examples of amino acid sequences of functional homologs of
the polypeptide set forth in SEQ ID NO:1311 are provided in FIG. 16
and in the sequence listing. Such functional homologs include
CeresClone: 971761 (SEQ ID NO: 1313), CeresClone: 1946574 (SEQ ID
NO: 1331), CeresClone: 2055551 (SEQ ID NO: 1340), CeresClone:
100045499 (SEQ ID NO: 1347), CeresClone: 1465853 (SEQ ID NO: 1353),
CeresClone: 753982 (SEQ ID NO: 1360), CeresClone: 1076093 (SEQ ID
NO: 1315), CeresClone: 1627875 (SEQ ID NO: 1317), CeresAnnot:
1508362 (SEQ ID NO: 1319), CeresAnnot: 1526950 (SEQ ID NO: 1321),
CeresClone: 1832333 (SEQ ID NO: 1323), GI: 74272607 (SEQ ID NO:
1324), GI: 192910782 (SEQ ID NO: 1325), GI: 157342563 (SEQ ID NO:
1326), GI: 159483497 (SEQ ID NO: 1327), CeresClone: 1839999 (SEQ ID
NO: 1329), CeresClone: 467335 (SEQ ID NO: 1333), GI: 168010087 (SEQ
ID NO: 1334), CeresClone: 1728202 (SEQ ID NO: 1336), CeresClone:
1994239 (SEQ ID NO: 1338), CeresAnnot: 8704704 (SEQ ID NO: 1342),
GI: 115453877 (SEQ ID NO: 1343), CeresClone: 1646411 (SEQ ID NO:
1345), CeresClone: 100920260 (SEQ ID NO: 1349), GI: 116786147 (SEQ
ID NO: 1350), GI: 116783944 (SEQ ID NO: 1351), GI: 82623397 (SEQ ID
NO: 1354), CeresAnnot: 8656662 (SEQ ID NO: 1356), CeresClone:
1417803 (SEQ ID NO: 1358), and GI: 145345927 (SEQ ID NO: 1361). In
some cases, a functional homolog of SEQ ID NO:1311 has an amino
acid sequence with at least 40% sequence identity, e.g., 50%, 52%,
56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 97%, 98%, or 99%
sequence identity, to the amino acid sequence set forth in SEQ ID
NO:1311.
[0091] The identification of conserved regions in a heat and/or
drought-tolerance polypeptide facilitates production of variants of
heat and/or drought-tolerance polypeptides. Variants of heat and/or
drought-tolerance polypeptides typically have 10 or fewer
conservative amino acid substitutions within the primary amino acid
sequence, e.g., 7 or fewer conservative amino acid substitutions, 5
or fewer conservative amino acid substitutions, or between 1 and 5
conservative substitutions. A useful variant polypeptide can be
constructed based on one of the alignments set forth in FIGS. 1-16.
Such a polypeptide includes the conserved regions, arranged in the
order depicted in the Figure from amino-terminal end to
carboxy-terminal end. Such a polypeptide may also include zero,
one, or more than one amino acid in positions marked by dashes.
When no amino acids are present at positions marked by dashes, the
length of such a polypeptide is the sum of the amino acid residues
in all conserved regions. When amino acids are present at all
positions marked by dashes, such a polypeptide has a length that is
the sum of the amino acid residues in all conserved regions and all
dashes.
[0092] C. Functional Homologs Identified by HMMER
[0093] In some embodiments, useful heat and/or drought-tolerance
polypeptides include those that fit a Hidden Markov Model based on
the polypeptides set forth in any one of FIGS. 1-16. A Hidden
Markov Model (HMM) is a statistical model of a consensus sequence
for a group of functional homologs. See, Durbin et al., Biological
Sequence Analysis: Probabilistic Models of Proteins and Nucleic
Acids, Cambridge University Press, Cambridge, UK (1998). An HMM is
generated by the program HMMER 2.3.2 with default program
parameters, using the sequences of the group of functional homologs
as input. The multiple sequence alignment is generated by ProbCons
(Do et al., Genome Res., 15(2):330-40 (2005)) version 1.11 using a
set of default parameters: -c, --consistency REPS of 2; -ir,
--iterative-refinement REPS of 100; -pre, --pre-training REPS of 0.
ProbCons is a public domain software program provided by Stanford
University.
[0094] The default parameters for building an HMM (hmmbuild) are as
follows: the default "architecture prior" (archpri) used by MAP
architecture construction is 0.85, and the default cutoff threshold
(idlevel) used to determine the effective sequence number is 0.62.
HMMER 2.3.2 was released Oct. 3, 2003 under a GNU general public
license, and is available from various sources on the World Wide
Web such as hmmer.janelia.org; hmmer.wustl.edu; and
fr.com/hmmer232/. Hmmbuild outputs the model as a text file.
[0095] The HMM for a group of functional homologs can be used to
determine the likelihood that a candidate heat and/or
drought-tolerance polypeptide sequence is a better fit to that
particular HMM than to a null HMM generated using a group of
sequences that are not structurally or functionally related. The
likelihood that a candidate polypeptide sequence is a better fit to
an HMM than to a null HMM is indicated by the HMM bit score, a
number generated when the candidate sequence is fitted to the HMM
profile using the HMMER hmmsearch program. The following default
parameters are used when running hmmsearch: the default E-value
cutoff (E) is 10, the default bit score cutoff (T) is negative
infinity, the default number of sequences in a database (Z) is the
real number of sequences in the database, the default E-value
cutoff for the per-domain ranked hit list (domE) is infinity, and
the default bit score cutoff for the per-domain ranked hit list
(domT) is negative infinity. A high HMM bit score indicates a
greater likelihood that the candidate sequence carries out one or
more of the biochemical or physiological function(s) of the
polypeptides used to generate the HMM. A high HMM bit score is at
least 20, and often is higher. Slight variations in the HMM bit
score of a particular sequence can occur due to factors such as the
order in which sequences are processed for alignment by multiple
sequence alignment algorithms such as the ProbCons program.
Nevertheless, such HMM bit score variation is minor.
[0096] The heat and/or drought-tolerance polypeptides discussed
below fit the indicated HMM with an HMM bit score greater than 20
(e.g., greater than 30, 40, 50, 60, 70, 80, 90, 100, 200, 300, 400,
or 500). In some embodiments, the HMM bit score of a heat and/or
drought-tolerance polypeptide discussed below is about 50%, 60%,
70%, 80%, 90%, or 95% of the HMM bit score of a functional homolog
provided in Sequence Listing. In some embodiments, a heat and/or
drought-tolerance polypeptide discussed below fits the indicated
HMM with an HMM bit score greater than 20, and has a domain
indicative of a heat and/or drought-tolerance polypeptide. In some
embodiments, a heat and/or drought-tolerance polypeptide discussed
below fits the indicated HMM with an HMM bit score greater than 20,
and has 70% or greater sequence identity (e.g., 75%, 80%, 85%, 90%,
95%, or 100% sequence identity) to an amino acid sequence shown in
any one of FIGS. 1-16.
[0097] Polypeptides are shown in the sequence listing that have HMM
bit scores greater than 270 when fitted to an HMM generated from
the amino acid sequences set forth in FIG. 1. Such polypeptides
include Ceres SEEDLINE ID no. ME00029 (SEQ ID NO:442), Public GI ID
no. 72384401 (SEQ ID NO:443), Ceres CLONE ID no. 1079382 (SEQ ID
NO:445), Ceres CLONE ID no. 1853461 (SEQ ID NO:447), Ceres CLONE ID
no. 1626485 (SEQ ID NO:451), Ceres CLONE ID no. 1713920 (SEQ ID
NO:459), Ceres CLONE ID no. 1772747 (SEQ ID NO:463), Ceres CLONE ID
no. 225960 (SEQ ID NO:465), Public GI ID no. 115443807 (SEQ ID
NO:470), Ceres CLONE ID no. 569388 (SEQ ID NO:483), Ceres CLONE ID
no. 1915549 (SEQ ID NO:449), Ceres CLONE ID no. 529871 (SEQ ID
NO:453), Ceres CLONE ID no. 1067079 (SEQ ID NO:455), Ceres CLONE ID
no. 1079572 (SEQ ID NO:457), Ceres ANNOT ID no. 1456550 (SEQ ID
NO:461), Ceres CLONE ID no. 1437889 (SEQ ID NO:469), Ceres CLONE ID
no. 2014249 (SEQ ID NO:472), Ceres CLONE ID no. 2033133 (SEQ ID
NO:474), Ceres CLONE ID no. 707404 (SEQ ID NO:476), Ceres CLONE ID
no. 1770680 (SEQ ID NO:478), Ceres ANNOT ID no. 1450989 (SEQ ID
NO:480), Public GI ID no. 72384445 (SEQ ID NO:481), Ceres CLONE ID
no. 1059299 (SEQ ID NO:485), and Ceres ANNOT ID no. 6008086 (SEQ ID
NO:487.
[0098] Polypeptides are shown in the sequence listing that have HMM
bit scores greater than 100 when fitted to an HMM generated from
the amino acid sequences set forth in FIG. 2. Such polypeptides
include Ceres SEEDLINE ID no. ME00045 (SEQ ID NO:489), Ceres CLONE
ID no. 571200 (SEQ ID NO:494), Ceres CLONE ID no. 1928532 (SEQ ID
NO:496), Ceres ANNOT ID no. 1490637 (SEQ ID NO:498), Ceres CLONE ID
no. 295496 (SEQ ID NO:514), Public GI ID no. 115463637 (SEQ ID
NO:515), Ceres CLONE ID no. 1999444 (SEQ ID NO:518), Public GI ID
no. 147844794 (SEQ ID NO:490), Public GI ID no. 147842279 (SEQ ID
NO:491), Public GI ID no. 145355441 (SEQ ID NO:492), Ceres ANNOT ID
no. 1461381 (SEQ ID NO:500), Ceres ANNOT ID no. 1440313 (SEQ ID
NO:502), Ceres ANNOT ID no. 1448275 (SEQ ID NO:504), Ceres ANNOT ID
no. 1437838 (SEQ ID NO:506), Ceres ANNOT ID no. 1501275 (SEQ ID
NO:508), Ceres CLONE ID no. 1644562 (SEQ ID NO:510), Ceres CLONE ID
no. 1925967 (SEQ ID NO:512), Public GI ID no. 115435904 (SEQ ID
NO:516), Public GI ID no. 125552168 (SEQ ID NO:519), Public GI ID
no. 125594093 (SEQ ID NO:520), Ceres CLONE ID no. 221188 (SEQ ID
NO:522), Ceres ANNOT ID no. 1477714 (SEQ ID NO:524), Ceres CLONE ID
no. 1787953 (SEQ ID NO:526), Ceres ANNOT ID no. 1531210 (SEQ ID
NO:528), Ceres CLONE ID no. 521176 (SEQ ID NO:530), Public GI ID
no. 22327055 (SEQ ID NO:531), Ceres ANNOT ID no. 1508824 (SEQ ID
NO:533), Ceres CLONE ID no. 38879 (SEQ ID NO:535), Public GI ID no.
42569309 (SEQ ID NO:536), Ceres CLONE ID no. 1817784 (SEQ ID
NO:538), Ceres CLONE ID no. 284637 (SEQ ID NO:540), Public GI ID
no. 125596251 (SEQ ID NO:541), Public GI ID no. 125554300 (SEQ ID
NO:543), Ceres CLONE ID no. 1935437 (SEQ ID NO:545), Ceres ANNOT ID
no. 1455622 (SEQ ID NO:547), Public GI ID no. 55771354 (SEQ ID
NO:548), Ceres ANNOT ID no. 1514655 (SEQ ID NO:550), Ceres CLONE ID
no. 1848736 (SEQ ID NO:552), Public GI ID no. 125569872 (SEQ ID
NO:553), Ceres CLONE ID no. 1645078 (SEQ ID NO:555), Ceres CLONE ID
no. 1790573 (SEQ ID NO:557), Public GI ID no. 4567251 (SEQ ID
NO:558), Ceres CLONE ID no. 444113 (SEQ ID NO:561), Public GI ID
no. 125525355 (SEQ ID NO:562), Ceres ANNOT ID no. 6028854 (SEQ ID
NO:564), and Ceres ANNOT ID no. 6115356 (SEQ ID NO:566).
[0099] Polypeptides are shown in the sequence listing that have HMM
bit scores greater than 150 when fitted to an HMM generated from
the amino acid sequences set forth in FIG. 3. Such polypeptides
include Ceres SEEDLINE ID no. ME02190 (SEQ ID NO:568), Ceres CLONE
ID no. 695006 (SEQ ID NO:578), Ceres ANNOT ID no. 1527488 (SEQ ID
NO:584), Public GI ID no. 79318519 (SEQ ID NO:569), Public GI ID
no. 79318537 (SEQ ID NO:570), Ceres CLONE ID no. 956998 (SEQ ID
NO:572), Ceres CLONE ID no. 978154 (SEQ ID NO:574), Ceres CLONE ID
no. 1035628 (SEQ ID NO:576), Ceres CLONE ID no. 464169 (SEQ ID
NO:580), Ceres ANNOT ID no. 1474075 (SEQ ID NO:582), Ceres ANNOT ID
no. 1474073 (SEQ ID NO:586), and Ceres ANNOT ID no. 1527486 (SEQ ID
NO:588).
[0100] Polypeptides are shown in the sequence listing that have HMM
bit scores greater than 120 when fitted to an HMM generated from
the amino acid sequences set forth in FIG. 4. Such polypeptides
include Ceres SEEDLINE ID no. ME02549 (SEQ ID NO:590), Ceres ANNOT
ID no. 1501305 (SEQ ID NO:623), Ceres CLONE ID no. 607280 (SEQ ID
NO:627), Public GI ID no. 92887174 (SEQ ID NO:632), Ceres CLONE ID
no. 1857162 (SEQ ID NO:634), Public GI ID no. 115477272 (SEQ ID
NO:674), Ceres CLONE ID no. 264002 (SEQ ID NO:682), Public GI ID
no. 147822456 (SEQ ID NO:591), Public GI ID no. 144923935 (SEQ ID
NO:592), Public GI ID no. 140038730 (SEQ ID NO:593), Public GI ID
no. 78708014 (SEQ ID NO:594), Public GI ID no. 115481362 (SEQ ID
NO:595), Public GI ID no. 125531350 (SEQ ID NO:596), Public GI ID
no. 125525598 (SEQ ID NO:597), Public GI ID no. 77548630 (SEQ ID
NO:598), Public GI ID no. 46798895 (SEQ ID NO:599), Public GI ID
no. 125564653 (SEQ ID NO:600), Ceres CLONE ID no. 998865 (SEQ ID
NO:602), Public GI ID no. 125556140 (SEQ ID NO:603), Public GI ID
no. 125534482 (SEQ ID NO:604), Public GI ID no. 125550135 (SEQ ID
NO:605), Ceres CLONE ID no. 639816 (SEQ ID NO:607), Public GI ID
no. 125562170 (SEQ ID NO:608), Ceres CLONE ID no. 1797059 (SEQ ID
NO:610), Ceres CLONE ID no. 1031510 (SEQ ID NO:612), Ceres CLONE ID
no. 1973081 (SEQ ID NO:614), Public GI ID no. 125561002 (SEQ ID
NO:615), Public GI ID no. 125541732 (SEQ ID NO:616), Public GI ID
no. 125560677 (SEQ ID NO:617), Public GI ID no. 125559115 (SEQ ID
NO:618), Public GI ID no. 15809903 (SEQ ID NO:619), Public GI ID
no. 30681703 (SEQ ID NO:620), Public GI ID no. 9759556 (SEQ ID
NO:621), Ceres ANNOT ID no. 1448303 (SEQ ID NO:625), Ceres ANNOT ID
no. 1448305 (SEQ ID NO:629), Ceres ANNOT ID no. 1448307 (SEQ ID
NO:631), Public GI ID no. 92888243 (SEQ ID NO:635), Public GI ID
no. 92879395 (SEQ ID NO:636), Ceres CLONE ID no. 528876 (SEQ ID
NO:638), Ceres ANNOT ID no. 1448352 (SEQ ID NO:640), Ceres ANNOT ID
no. 1437745 (SEQ ID NO:642), Ceres ANNOT ID no. 1464146 (SEQ ID
NO:644), Ceres ANNOT ID no. 1437744 (SEQ ID NO:646), Public GI ID
no. 92894684 (SEQ ID NO:647), Ceres CLONE ID no. 3964 (SEQ ID
NO:650), Public GI ID no. 13272389 (SEQ ID NO:651), Ceres ANNOT ID
no. 1481203 (SEQ ID NO:653), Public GI ID no. 15227699 (SEQ ID
NO:654), Public GI ID no. 92886084 (SEQ ID NO:655), Public GI ID
no. 15239947 (SEQ ID NO:656), Ceres CLONE ID no. 34878 (SEQ ID
NO:658), Ceres CLONE ID no. 150484 (SEQ ID NO:660), Public GI ID
no. 21553545 (SEQ ID NO:661), Public GI ID no. 15222843 (SEQ ID
NO:662), Public GI ID no. 38230552 (SEQ ID NO:663), Public GI ID
no. 3420008 (SEQ ID NO:664), Public GI ID no. 15230602 (SEQ ID
NO:665), Public GI ID no. 3420004 (SEQ ID NO:666), Public GI ID no.
2129586 (SEQ ID NO:667), Public GI ID no. 15217849 (SEQ ID NO:668),
Public GI ID no. 15227704 (SEQ ID NO:669), Ceres ANNOT ID no.
1465750 (SEQ ID NO:671), Ceres CLONE ID no. 1983975 (SEQ ID
NO:673), Public GI ID no. 15226028 (SEQ ID NO:675), Public GI ID
no. 115459524 (SEQ ID NO:676), Ceres CLONE ID no. 1793353 (SEQ ID
NO:678), Ceres ANNOT ID no. 1467399 (SEQ ID NO:680), Ceres CLONE ID
no. 1982930 (SEQ ID NO:684), Public GI ID no. 125540700 (SEQ ID
NO:685), Ceres ANNOT ID no. 1448743 (SEQ ID NO:687), Public GI ID
no. 50251910 (SEQ ID NO:688), Ceres CLONE ID no. 1836748 (SEQ ID
NO:690), Public GI ID no. 3420006 (SEQ ID NO:691), Public GI ID no.
92879376 (SEQ ID NO:692), Ceres CLONE ID no. 838941 (SEQ ID
NO:694), Ceres ANNOT ID no. 1437746 (SEQ ID NO:696), Ceres ANNOT ID
no. 6017241 (SEQ ID NO:698), Ceres ANNOT ID no. 6085947 (SEQ ID
NO:700), and Ceres ANNOT ID no. 6017242 (SEQ ID NO:702).
[0101] Polypeptides are shown in the sequence listing that have HMM
bit scores greater than 55 when fitted to an HMM generated from the
amino acid sequences set forth in FIG. 5. Such polypeptides include
Ceres SEEDLINE ID no. ME02865 (SEQ ID NO:705), Public GI ID no.
79320952 (SEQ ID NO:706), Public GI ID no. 79320957 (SEQ ID
NO:707), Public GI ID no. 6692094 (SEQ ID NO:708), and Public GI ID
no. 145323049 (SEQ ID NO:709).
[0102] Polypeptides are shown in the sequence listing that have HMM
bit scores greater than 185 when fitted to an HMM generated from
the amino acid sequences set forth in FIG. 6. Such polypeptides
include Ceres SEEDLINE ID no. ME03227 (SEQ ID NO:711), Ceres CLONE
ID no. 964616 (SEQ ID NO:714), Ceres CLONE ID no. 100009667 (SEQ ID
NO:715), Ceres ANNOT ID no. 1444568 (SEQ ID NO:719), Ceres CLONE ID
no. 719489 (SEQ ID NO:725), Ceres CLONE ID no. 587748 (SEQ ID
NO:733), Public GI ID no. 125528114 (SEQ ID NO:734), Ceres CLONE ID
no. 274172 (SEQ ID NO:737), Public GI ID no. 144923134 (SEQ ID
NO:712), Ceres ANNOT ID no. 1471437 (SEQ ID NO:717), Ceres CLONE ID
no. 1270484 (SEQ ID NO:721), Ceres CLONE ID no. 1075098 (SEQ ID
NO:723), Public GI ID no. 18412211 (SEQ ID NO:727), Ceres CLONE ID
no. 20358 (SEQ ID NO:729), Ceres CLONE ID no. 1915503 (SEQ ID
NO:731), Public GI ID no. 115440619 (SEQ ID NO:735), Public GI ID
no. 125572387 (SEQ ID NO:738), and Ceres ANNOT ID no. 6015812 (SEQ
ID NO:740).
[0103] Polypeptides are shown in the sequence listing that have HMM
bit scores greater than 110 when fitted to an HMM generated from
the amino acid sequences set forth in FIG. 7. Such polypeptides
include Ceres SEEDLINE ID no. ME04477 (SEQ ID NO:742), Ceres CLONE
ID no. 1620215 (SEQ ID NO: 759); Ceres CLONE ID no. 1798756 (SEQ ID
NO: 763); Public GI ID no. 38016527 (SEQ ID NO: 796); Public GI ID
no. 75133829 (SEQ ID NO: 799); Ceres ANNOT ID no. 1460527 (SEQ ID
NO: 801); Public GI ID no. 119720772 (SEQ ID NO: 857); Ceres CLONE
ID no. 708446 (SEQ ID NO: 859); Public GI ID no. 92896423 (SEQ ID
NO: 892); Ceres CLONE ID no. 1387149 (SEQ ID NO: 912); Public GI ID
no. 5031281 (SEQ ID NO: 953); Ceres CLONE ID no. 1775820 (SEQ ID
NO: 955); Public GI ID no. 35187687 (SEQ ID NO: 988); Public GI ID
no. 115468934 (SEQ ID NO: 991); Public GI ID no. 118424243 (SEQ ID
NO: 1000); Ceres ANNOT ID no. 6063957 (SEQ ID NO: 1010); Public GI
ID no. 113196593 (SEQ ID NO: 798); Public GI ID no. 112819496 (SEQ
ID NO: 999); Public GI ID no. 147783026 (SEQ ID NO: 743); Public GI
ID no. 119367488 (SEQ ID NO: 744); Public GI ID no. 147860340 (SEQ
ID NO: 745); Public GI ID no. 115477170 (SEQ ID NO: 757); Ceres
CLONE ID no. 1931889 (SEQ ID NO: 761); Ceres CLONE ID no. 1918424
(SEQ ID NO: 767); Ceres CLONE ID no. 1845154 (SEQ ID NO: 771);
Ceres Clone ID no. 1084216 (SEQ ID NO: 797); Ceres Annot ID no.
8644540 (SEQ ID NO: 805); Ceres ANNOT ID no. 1450673 (SEQ ID NO:
807); Public GI ID no. 116778802 (SEQ ID NO: 808); Public GI ID no.
116778893 (SEQ ID NO: 809); and Public GI ID no. 116778998 (SEQ ID
NO: 810); Public GI ID no. 157849766 (SEQ ID NO: 811); Public GI ID
no. 159474166 (SEQ ID NO: 812); Public GI ID no. 168036656 (SEQ ID
NO: 813); Ceres ANNOT ID no. 1456578 (SEQ ID NO: 815); Public GI ID
no. 168053490 (SEQ ID NO: 816); Public GI ID no. 193237563 (SEQ ID
NO: 817); Ceres Clone ID no. 100879386 (SEQ ID NO: 819); Ceres
Clone ID no. 2055733 (SEQ ID NO: 823); Ceres Clone ID no. 2056478
(SEQ ID NO: 825); Ceres CLONE ID no. 13007 (SEQ ID NO: 832); Ceres
CLONE ID no. 5522 (SEQ ID NO: 834); Ceres CLONE ID no. 30543 (SEQ
ID NO: 836); Ceres CLONE ID no. 14203 (SEQ ID NO: 839); Ceres CLONE
ID no. 975913 (SEQ ID NO: 846); Ceres CLONE ID no. 967417 (SEQ ID
NO: 848); Ceres CLONE ID no. 1614593 (SEQ ID NO: 863); Ceres CLONE
ID no. 2025938 (SEQ ID NO: 885); Ceres CLONE ID no. 634261 (SEQ ID
NO: 900); Ceres CLONE ID no. 1423851 (SEQ ID NO: 918); Ceres CLONE
ID no. 1589047 (SEQ ID NO: 939); Ceres CLONE ID no. 1748922 (SEQ ID
NO: 944); Ceres CLONE ID no. 1787151 (SEQ ID NO: 957); Ceres CLONE
ID no. 1765871 (SEQ ID NO: 973); Ceres CLONE ID no. 1990071 (SEQ ID
NO: 977); Public GI ID no. 125556051 (SEQ ID NO: 989); Public GI ID
no. 125561658 (SEQ ID NO: 990); Public GI ID no. 115470773 (SEQ ID
NO: 992); Public GI ID no. 115444813 (SEQ ID NO: 993); GI ID no.
169363 (SEQ ID NO: 1215) and Ceres CLONE ID no. 24255 (SEQ ID NO:
1242).
[0104] Polypeptides are shown in the sequence listing that have HMM
bit scores greater than 215 when fitted to an HMM generated from
the amino acid sequences set forth in FIG. 8. Such polypeptides
include Ceres SEEDLINE ID no. ME18396 (SEQ ID NO:1012), Ceres CLONE
ID no. 287430 (SEQ ID NO:1018), Ceres ANNOT ID no. 451889 (SEQ ID
NO:1020), Ceres CLONE ID no. 936084 (SEQ ID NO:1055), Ceres CLONE
ID no. 1792501 (SEQ ID NO:1066), Ceres ANNOT ID no. 1437875 (SEQ ID
NO:1068), Public GI ID no. 1853968 (SEQ ID NO:1093), Public GI ID
no. 27530032 (SEQ ID NO:1094), Ceres CLONE ID no. 1834483 (SEQ ID
NO:1110), Public GI ID no. 84579418 (SEQ ID NO:1113), Public GI ID
no. 15077030 (SEQ ID NO:1117), Public GI ID no. 13752458 (SEQ ID
NO:1118), Public GI ID no. 147865685 (SEQ ID NO:1013), Public GI ID
no. 119503368 (SEQ ID NO:1014), Public GI ID no. 72161874 (SEQ ID
NO:1015), Public GI ID no. 91780661 (SEQ ID NO:1016), Ceres ANNOT
ID no. 835908 (SEQ ID NO:1022), Public GI ID no. 38326750 (SEQ ID
NO:1023), Ceres CLONE ID no. 1939396 (SEQ ID NO:1025), Ceres CLONE
ID no. 403637 (SEQ ID NO:1027), Public GI ID no. 42539907 (SEQ ID
NO:1028), Ceres CLONE ID no. 1836494 (SEQ ID NO:1030), Public GI ID
no. 15192945 (SEQ ID NO:1031), Ceres CLONE ID no. 1607947 (SEQ ID
NO:1033), Ceres CLONE ID no. 115880 (SEQ ID NO:1035), Ceres CLONE
ID no. 1074009 (SEQ ID NO:1037), Ceres CLONE ID no. 476073 (SEQ ID
NO:1039), Ceres CLONE ID no. 554053 (SEQ ID NO:1041), Ceres CLONE
ID no. 391449 (SEQ ID NO:1045), Ceres CLONE ID no. 1846400 (SEQ ID
NO:1048), Ceres ANNOT ID no. 1475185 (SEQ ID NO:1050), Public GI ID
no. 115457148 (SEQ ID NO:1051), Ceres ANNOT ID no. 1454960 (SEQ ID
NO:1053), Public GI ID no. 50346893 (SEQ ID NO:1056), Ceres CLONE
ID no. 1931526 (SEQ ID NO:1058), Ceres ANNOT ID no. 1454260 (SEQ ID
NO:1061), Public GI ID no. 115456131 (SEQ ID NO:1062), Ceres CLONE
ID no. 159151 (SEQ ID NO:1064), Ceres CLONE ID no. 1842801 (SEQ ID
NO:1071), Ceres CLONE ID no. 533030 (SEQ ID NO:1073), Ceres CLONE
ID no. 1931881 (SEQ ID NO:1075), Ceres ANNOT ID no. 1480006 (SEQ ID
NO:1077), Ceres CLONE ID no. 1895007 (SEQ ID NO:1079), Public GI ID
no. 3598863 (SEQ ID NO:1080), Ceres ANNOT ID no. 1471735 (SEQ ID
NO:1082), Ceres CLONE ID no. 1937530 (SEQ ID NO:1084), Ceres CLONE
ID no. 1833050 (SEQ ID NO:1086), Ceres CLONE ID no. 644213 (SEQ ID
NO:1088), Ceres CLONE ID no. 568154 (SEQ ID NO:1090), Ceres CLONE
ID no. 527598 (SEQ ID NO:1092), Ceres ANNOT ID no. 1487614 (SEQ ID
NO:1096), Ceres ANNOT ID no. 1541881 (SEQ ID NO:1098), Ceres ANNOT
ID no. 1488465 (SEQ ID NO:1100), Ceres ANNOT ID no. 1488468 (SEQ ID
NO:1102), Ceres ANNOT ID no. 1541884 (SEQ ID NO:1104), Ceres ANNOT
ID no. 1454972 (SEQ ID NO:1106), Ceres ANNOT ID no. 1475186 (SEQ ID
NO:1108), Ceres CLONE ID no. 686198 (SEQ ID NO:1112), Public GI ID
no. 84579420 (SEQ ID NO:1114), Ceres CLONE ID no. 605144 (SEQ ID
NO:1116), Public GI ID no. 15077028 (SEQ ID NO:1119), Ceres ANNOT
ID no. 1454963 (SEQ ID NO:1121), Ceres ANNOT ID no. 6030208 (SEQ ID
NO:1123), and Ceres ANNOT ID no. 6076767 (SEQ ID NO:1125).
[0105] Polypeptides are shown in the sequence listing that have HMM
bit scores greater than 105 when fitted to an HMM generated from
the amino acid sequences set forth in FIG. 9. Such polypeptides
include Ceres SEEDLINE ID no. ME20095 (SEQ ID NO:1129), Public GI
ID no. 72255610 (SEQ ID NO:933), Ceres SEEDLINE ID no. ME20095 (SEQ
ID NO:1129), Ceres CLONE ID no. 1562633 (SEQ ID NO:1140), Ceres
ANNOT ID no. 1692728 (SEQ ID NO:1142), Ceres CLONE ID no. 921110
(SEQ ID NO:1144), Ceres CLONE ID no. 1791180 (SEQ ID NO:1146),
Ceres CLONE ID no. 527891 (SEQ ID NO:1148), Ceres ANNOT ID no.
1522414 (SEQ ID NO:1150), Public GI ID no. 51458330 (SEQ ID
NO:1182), Public GI ID no. 18399940 (SEQ ID NO:63), Ceres ANNOT ID
no. 6107929 (SEQ ID NO:73), Ceres ANNOT ID no. 1473094 (SEQ ID
NO:96), Ceres ANNOT ID no. 1503955 (SEQ ID NO:127), Ceres ANNOT ID
no. 1526501 (SEQ ID NO:139), Public GI ID no. 15235713 (SEQ ID
NO:179), Public GI ID no. 15234552 (SEQ ID NO:180), Ceres ANNOT ID
no. 1520801 (SEQ ID NO:209), Ceres ANNOT ID no. 1443434 (SEQ ID
NO:467), Public GI ID no. 30696058 (SEQ ID NO:559), Ceres ANNOT ID
no. 1509601 (SEQ ID NO:648), Ceres ANNOT ID no. 1451912 (SEQ ID
NO:726), Ceres ANNOT ID no. 6017545 (SEQ ID NO:741), Ceres ANNOT ID
no. 1504999 (SEQ ID NO:765), Ceres ANNOT ID no. 1456402 (SEQ ID
NO:773), Ceres ANNOT ID no. 1496359 (SEQ ID NO:781), Ceres ANNOT ID
no. 1447260 (SEQ ID NO:842), Ceres ANNOT ID no. 1469023 (SEQ ID
NO:861), Ceres ANNOT ID no. 1474186 (SEQ ID NO:865), Ceres ANNOT ID
no. 1527596 (SEQ ID NO:867), Ceres CLONE ID no. 1919624 (SEQ ID
NO:896), Ceres CLONE ID no. 677797 (SEQ ID NO:916), Public GI ID
no. 7960729 (SEQ ID NO:932), Ceres CLONE ID no. 941845 (SEQ ID
NO:1043), Public GI ID no. 30683008 (SEQ ID NO:1046), Public GI ID
no. 42568786 (SEQ ID NO:1059), Ceres CLONE ID no. 255364 (SEQ ID
NO:1069), Public GI ID no. 147771549 (SEQ ID NO:1130), Public GI ID
no. 144924741 (SEQ ID NO:1131), Public GI ID no. 121594508 (SEQ ID
NO:1132), Public GI ID no. 91202290 (SEQ ID NO:1133), Public GI ID
no. 89900939 (SEQ ID NO:1134), Public GI ID no. 77917672 (SEQ ID
NO:1135), Public GI ID no. 77165252 (SEQ ID NO:1136), Public GI ID
no. 115375374 (SEQ ID NO:1137), Public GI ID no. 110602449 (SEQ ID
NO:1138), Public GI ID no. 125540573 (SEQ ID NO:1151), Public GI ID
no. 125561519 (SEQ ID NO:1152), Public GI ID no. 125528345 (SEQ ID
NO:1153), Public GI ID no. 116310408 (SEQ ID NO:1154), Public GI ID
no. 115483332 (SEQ ID NO:1156), Public GI ID no. 125575640 (SEQ ID
NO:1157), Public GI ID no. 115476368 (SEQ ID NO:1158), Public GI ID
no. 115478841 (SEQ ID NO:1159), Public GI ID no. 115467158 (SEQ ID
NO:1160), Public GI ID no. 72384477 (SEQ ID NO:1161), Public GI ID
no. 116054703 (SEQ ID NO:1162), Ceres CLONE ID no. 1786317 (SEQ ID
NO:1164), Ceres CLONE ID no. 1791336 (SEQ ID NO:1166), Ceres CLONE
ID no. 1877752 (SEQ ID NO:1168), Ceres CLONE ID no. 1821191 (SEQ ID
NO:1170), Ceres CLONE ID no. 446838 (SEQ ID NO:1172), Ceres CLONE
ID no. 1556915 (SEQ ID NO:1174), Public GI ID no. 90200725 (SEQ ID
NO:1175), Ceres CLONE ID no. 1580247 (SEQ ID NO:1177), Ceres CLONE
ID no. 644201 (SEQ ID NO:1179), Ceres CLONE ID no. 538689 (SEQ ID
NO:1181), Ceres CLONE ID no. 817225 (SEQ ID NO:1184), and Ceres
CLONE ID no. 923842 (SEQ ID NO:1186).
[0106] Polypeptides are shown in the sequence listing that have HMM
bit scores greater than 55 when fitted to an HMM generated from the
amino acid sequences set forth in FIG. 10. Such polypeptides
include Ceres CLONE ID no. 31309 (SEQ ID NO:60), Ceres CLONE ID no.
872030 (SEQ ID NO:62), Ceres CLONE ID no. 100029223 (SEQ ID NO:74),
Ceres CLONE ID no. 1939845 (SEQ ID NO:98), Ceres CLONE ID no.
2015383 (SEQ ID NO:102), Ceres CLONE ID no. 1607893 (SEQ ID
NO:104), Ceres CLONE ID no. 1075133 (SEQ ID NO:152), Ceres CLONE ID
no. 1218065 (SEQ ID NO:915), Public GI ID no. 15227919 (SEQ ID
NO:64), Public GI ID no. 9843641 (SEQ ID NO:65), Public GI ID no.
124294789 (SEQ ID NO:66), Public GI ID no. 15233446 (SEQ ID NO:67),
Public GI ID no. 115477679 (SEQ ID NO:68), Public GI ID no.
42407552 (SEQ ID NO:69), Public GI ID no. 125562450 (SEQ ID NO:70),
Public GI ID no. 115469980 (SEQ ID NO:71), Ceres ANNOT ID no.
1442539 (SEQ ID NO:76), Ceres ANNOT ID no. 1442538 (SEQ ID NO:78),
Ceres ANNOT ID no. 1460661 (SEQ ID NO:80), Ceres ANNOT ID no.
1452884 (SEQ ID NO:82), Ceres ANNOT ID no. 1450523 (SEQ ID NO:84),
Ceres ANNOT ID no. 1442257 (SEQ ID NO:86), Ceres ANNOT ID no.
1541121 (SEQ ID NO:88), Ceres ANNOT ID no. 1480481 (SEQ ID NO:90),
Ceres ANNOT ID no. 1487713 (SEQ ID NO:92), Ceres ANNOT ID no.
1456204 (SEQ ID NO:94), Ceres CLONE ID no. 1387402 (SEQ ID NO:100),
Ceres CLONE ID no. 972919 (SEQ ID NO:106), Ceres CLONE ID no.
1645860 (SEQ ID NO:108), Ceres CLONE ID no. 1042804 (SEQ ID
NO:110), Ceres CLONE ID no. 1606678 (SEQ ID NO:112), Ceres CLONE ID
no. 684496 (SEQ ID NO:114), Ceres CLONE ID no. 1062366 (SEQ ID
NO:116), Ceres CLONE ID no. 1722931 (SEQ ID NO:118), Ceres CLONE ID
no. 746169 (SEQ ID NO:120), Ceres CLONE ID no. 1050475 (SEQ ID
NO:122), Ceres CLONE ID no. 1728904 (SEQ ID NO:124), Ceres CLONE ID
no. 1698538 (SEQ ID NO:126), Ceres CLONE ID no. 1762328 (SEQ ID
NO:130), Ceres CLONE ID no. 1590072 (SEQ ID NO:132), Ceres CLONE ID
no. 471593 (SEQ ID NO:134), Ceres CLONE ID no. 1031667 (SEQ ID
NO:136), Ceres CLONE ID no. 1826085 (SEQ ID NO:138), Ceres CLONE ID
no. 373932 (SEQ ID NO:142), Ceres CLONE ID no. 1797836 (SEQ ID
NO:144), Ceres CLONE ID no. 1102774 (SEQ ID NO:146), Ceres CLONE ID
no. 842287 (SEQ ID NO:148), Ceres CLONE ID no. 568205 (SEQ ID
NO:150), Ceres ANNOT ID no. 6108997 (SEQ ID NO:154), Ceres ANNOT ID
no. 6023609 (SEQ ID NO:156), and Ceres ANNOT ID no. 6017906 (SEQ ID
NO:158).
[0107] Polypeptides are shown in the sequence listing that have HMM
bit scores greater than 55 when fitted to an HMM generated from the
amino acid sequences set forth in FIG. 11. Such polypeptides
include Ceres LOCUS ID no.At2g26150 (SEQ ID NO:364), Ceres ANNOT ID
no. 1455221 (SEQ ID NO:407), Public GI ID no. 729774 (SEQ ID
NO:408), Ceres CLONE ID no. 1414288 (SEQ ID NO:410), Public GI ID
no. 115482048 (SEQ ID NO:413), Public GI ID no. 56117815 (SEQ ID
NO:421), Ceres ANNOT ID no. 1485538 (SEQ ID NO:366), Ceres ANNOT ID
no. 1538505 (SEQ ID NO:368), Public GI ID no. 8347238 (SEQ ID
NO:369), Public GI ID no. 125542955 (SEQ ID NO:370), Public GI ID
no. 115446219 (SEQ ID NO:371), Ceres CLONE ID no. 1597127 (SEQ ID
NO:373), Ceres CLONE ID no. 1584568 (SEQ ID NO:375), Ceres CLONE ID
no. 286056 (SEQ ID NO:377), Ceres ANNOT ID no. 1543432 (SEQ ID
NO:379), Ceres CLONE ID no. 1728062 (SEQ ID NO:381), Ceres ANNOT ID
no. 1448950 (SEQ ID NO:383), Ceres CLONE ID no. 771500 (SEQ ID
NO:385), Ceres ANNOT ID no. 1485096 (SEQ ID NO:387), Ceres CLONE ID
no. 1646104 (SEQ ID NO:389), Public GI ID no. 111184724 (SEQ ID
NO:390), Ceres CLONE ID no. 1362475 (SEQ ID NO:392), Ceres CLONE ID
no. 597906 (SEQ ID NO:394), Ceres CLONE ID no. 615781 (SEQ ID
NO:396), Ceres CLONE ID no. 538713 (SEQ ID NO:398), Ceres CLONE ID
no. 1794141 (SEQ ID NO:400), Public GI ID no. 5821138 (SEQ ID
NO:401), Public GI ID no. 886742 (SEQ ID NO:402), Ceres CLONE ID
no. 1814498 (SEQ ID NO:404), Public GI ID no. 7158882 (SEQ ID
NO:405), Ceres ANNOT ID no. 1452564 (SEQ ID NO:412), Public GI ID
no. 125557431 (SEQ ID NO:414), Ceres ANNOT ID no. 1442880 (SEQ ID
NO:416), Ceres ANNOT ID no. 1463437 (SEQ ID NO:418), Public GI ID
no. 42415865 (SEQ ID NO:419), Public GI ID no. 33087081 (SEQ ID
NO:420), Public GI ID no. 115521217 (SEQ ID NO:422), Public GI ID
no. 115521211 (SEQ ID NO:423), Public GI ID no. 115521215 (SEQ ID
NO:424), Ceres ANNOT ID no. 1454376 (SEQ ID NO:426), Ceres CLONE ID
no. 835571 (SEQ ID NO:428), Ceres ANNOT ID no. 1500046 (SEQ ID
NO:430), Public GI ID no. 110738569 (SEQ ID NO:431), Public GI ID
no. 125542510 (SEQ ID NO:432), Public GI ID no. 30686034 (SEQ ID
NO:433), Public GI ID no. 125570883 (SEQ ID NO:434), Public GI ID
no. 42562463 (SEQ ID NO:435), Ceres CLONE ID no. 100043265 (SEQ ID
NO:436), Ceres ANNOT ID no. 6089790 (SEQ ID NO:438), and Ceres
ANNOT ID no. 6043635 (SEQ ID NO:440).
[0108] Polypeptides are shown in the sequence listing that have HMM
bit scores greater than 50 when fitted to an HMM generated from the
amino acid sequences set forth in FIG. 12. Such polypeptides
include Ceres LOCUS ID no.At1g32330 (SEQ ID NO:311), Ceres ANNOT ID
no. 1538958 (SEQ ID NO:319), Public GI ID no. 115521213 (SEQ ID
NO:320), Public GI ID no. 42415865 (SEQ ID NO:321), Public GI ID
no. 729775 (SEQ ID NO:322), Public GI ID no. 11386827 (SEQ ID
NO:323), Public GI ID no. 115456675 (SEQ ID NO:326), Public GI ID
no. 89274218 (SEQ ID NO:335), Public GI ID no. 125546035 (SEQ ID
NO:312), Ceres CLONE ID no. 125228 (SEQ ID NO:314), Public GI ID
no. 115465055 (SEQ ID NO:315), Public GI ID no. 125553080 (SEQ ID
NO:316), Public GI ID no. 8920606 (SEQ ID NO:317), Ceres CLONE ID
no. 1919643 (SEQ ID NO:325), Public GI ID no. 429155 (SEQ ID
NO:327), Ceres CLONE ID no. 1194199 (SEQ ID NO:329), Public GI ID
no. 15236631 (SEQ ID NO:330), Ceres ANNOT ID no. 1523420 (SEQ ID
NO:332), Ceres ANNOT ID no. 1447046 (SEQ ID NO:334), Public GI ID
no. 25296101 (SEQ ID NO:336), Public GI ID no. 15228440 (SEQ ID
NO:337), Public GI ID no. 6624612 (SEQ ID NO:338), Public GI ID no.
125588688 (SEQ ID NO:339), Public GI ID no. 115470859 (SEQ ID
NO:340), Public GI ID no. 15228865 (SEQ ID NO:341), Ceres CLONE ID
no. 1571069 (SEQ ID NO:343), Public GI ID no. 115455903 (SEQ ID
NO:344), Ceres ANNOT ID no. 1459422 (SEQ ID NO:346), Ceres CLONE ID
no. 1577511 (SEQ ID NO:348), Public GI ID no. 115451657 (SEQ ID
NO:349), Public GI ID no. 15225255 (SEQ ID NO:350), Public GI ID
no. 125545695 (SEQ ID NO:351), Ceres ANNOT ID no. 1471817 (SEQ ID
NO:353), Ceres ANNOT ID no. 1440896 (SEQ ID NO:355), Public GI ID
no. 119393868 (SEQ ID NO:356), Ceres ANNOT ID no. 1517025 (SEQ ID
NO:358), Ceres ANNOT ID no. 1512410 (SEQ ID NO:360), and Ceres
ANNOT ID no. 6035498 (SEQ ID NO:362).
[0109] Polypeptides are shown in the sequence listing that have HMM
bit scores greater than 25 when fitted to an HMM generated from the
amino acid sequences set forth in FIG. 13. Such polypeptides
include Ceres CLONE ID no. 41543 (SEQ ID NO:160) Ceres CLONE ID no.
1837065 (SEQ ID NO:162), Ceres ANNOT ID no. 1531178 (SEQ ID
NO:184), Ceres CLONE ID no. 470694 (SEQ ID NO:242), Public GI ID
no. 92867368 (SEQ ID NO:253), Ceres CLONE ID no. 859707 (SEQ ID
NO:256), Ceres CLONE ID no. 392275 (SEQ ID NO:268), Ceres CLONE ID
no. 1828394 (SEQ ID NO:282), Public GI ID no. 115466694 (SEQ ID
NO:294), Ceres CLONE ID no. 1895763 (SEQ ID NO:164), Ceres CLONE ID
no. 1850460 (SEQ ID NO:166), Ceres CLONE ID no. 1808074 (SEQ ID
NO:168), Ceres CLONE ID no. 1832882 (SEQ ID NO:170), Ceres CLONE ID
no. 1978977 (SEQ ID NO:172), Ceres CLONE ID no. 1918742 (SEQ ID
NO:174), Ceres CLONE ID no. 1940023 (SEQ ID NO:176), Ceres CLONE ID
no. 1923495 (SEQ ID NO:178), Ceres CLONE ID no. 1851388 (SEQ ID
NO:182), Ceres ANNOT ID no. 1477681 (SEQ ID NO:186), Ceres ANNOT ID
no. 1508800 (SEQ ID NO:188), Ceres ANNOT ID no. 1455597 (SEQ ID
NO:190), Ceres ANNOT ID no. 1532016 (SEQ ID NO:192), Ceres ANNOT ID
no. 1478526 (SEQ ID NO:194), Ceres ANNOT ID no. 1492567 (SEQ ID
NO:196), Ceres ANNOT ID no. 1515273 (SEQ ID NO:198), Ceres ANNOT ID
no. 1461979 (SEQ ID NO:200), Ceres ANNOT ID no. 1439702 (SEQ ID
NO:202), Ceres ANNOT ID no. 1448044 (SEQ ID NO:204), Ceres ANNOT ID
no. 1483370 (SEQ ID NO:206), Ceres ANNOT ID no. 1445651 (SEQ ID
NO:208), Ceres ANNOT ID no. 1490863 (SEQ ID NO:212), Ceres ANNOT ID
no. 1501043 (SEQ ID NO:214), Ceres ANNOT ID no. 1527507 (SEQ ID
NO:216), Ceres ANNOT ID no. 1498633 (SEQ ID NO:218), Ceres ANNOT ID
no. 1465403 (SEQ ID NO:220), Ceres ANNOT ID no. 1510956 (SEQ ID
NO:222), Ceres ANNOT ID no. 1457735 (SEQ ID NO:224), Public GI ID
no. 15224711 (SEQ ID NO:225), Ceres CLONE ID no. 40062 (SEQ ID
NO:228), Public GI ID no. 15220315 (SEQ ID NO:229), Ceres CLONE ID
no. 36480 (SEQ ID NO:231), Ceres CLONE ID no. 2443 (SEQ ID NO:233),
Public GI ID no. 30685375 (SEQ ID NO:234), Public GI ID no.
126009434 (SEQ ID NO:235), Ceres CLONE ID no. 965753 (SEQ ID
NO:237), Ceres CLONE ID no. 948834 (SEQ ID NO:239), Public GI ID
no. 33347409 (SEQ ID NO:240), Ceres CLONE ID no. 643614 (SEQ ID
NO:244), Ceres CLONE ID no. 1049362 (SEQ ID NO:246), Ceres CLONE ID
no. 546675 (SEQ ID NO:248), Ceres CLONE ID no. 522046 (SEQ ID
NO:250), Ceres CLONE ID no. 651076 (SEQ ID NO:252), Public GI ID
no. 92897590 (SEQ ID NO:254), Ceres CLONE ID no. 634176 (SEQ ID
NO:258), Ceres CLONE ID no. 1041432 (SEQ ID NO:260), Ceres CLONE ID
no. 1065203 (SEQ ID NO:262), Ceres CLONE ID no. 555361 (SEQ ID
NO:264), Ceres CLONE ID no. 704227 (SEQ ID NO:266), Ceres CLONE ID
no. 100819945 (SEQ ID NO:269), Ceres CLONE ID no. 282584 (SEQ ID
NO:271), Ceres CLONE ID no. 1448469 (SEQ ID NO:273), Ceres CLONE ID
no. 241246 (SEQ ID NO:275), Ceres CLONE ID no. 234443 (SEQ ID
NO:277), Ceres CLONE ID no. 293549 (SEQ ID NO:279), Public GI ID
no. 109450926 (SEQ ID NO:280), Ceres CLONE ID no. 1803923 (SEQ ID
NO:284), Ceres CLONE ID no. 2024557 (SEQ ID NO:286), Ceres CLONE ID
no. 1820364 (SEQ ID NO:288), Public GI ID no. 125554274 (SEQ ID
NO:289), Public GI ID no. 125531165 (SEQ ID NO:290), Public GI ID
no. 125541514 (SEQ ID NO:291), Public GI ID no. 125557559 (SEQ ID
NO:292), Public GI ID no. 125557844 (SEQ ID NO:293), Public GI ID
no. 115449295 (SEQ ID NO:295), Public GI ID no. 115481182 (SEQ ID
NO:296), Public GI ID no. 20177639 (SEQ ID NO:297), Public GI ID
no. 115471021 (SEQ ID NO:298), Public GI ID no. 115456011 (SEQ ID
NO:299), Public GI ID no. 125588290 (SEQ ID NO:300), Public GI ID
no. 115477799 (SEQ ID NO:301), Public GI ID no. 115471385 (SEQ ID
NO:302), Public GI ID no. 125599705 (SEQ ID NO:303), Public GI ID
no. 115489810 (SEQ ID NO:304), Public GI ID no. 55276718 (SEQ ID
NO:305), Ceres ANNOT ID no. 6025568 (SEQ ID NO:307), and Ceres
ANNOT ID no. 6091128 (SEQ ID NO:309).
[0110] Polypeptides are shown in the sequence listing that have HMM
bit scores greater than 85 when fitted to an HMM generated from the
amino acid sequences set forth in FIG. 14. Such polypeptides
include Ceres CLONE ID no. 14572 (SEQ ID NO:2), Public GI ID no.
115470807 (SEQ ID NO:5), Ceres CLONE ID no. 1842931 (SEQ ID NO:15),
Ceres CLONE ID no. 321308 (SEQ ID NO:17), Ceres CLONE ID no.
1725811 (SEQ ID NO:19), Ceres CLONE ID no. 1357455 (SEQ ID NO:21),
Ceres CLONE ID no. 943370 (SEQ ID NO:23), Ceres CLONE ID no.
1327712 (SEQ ID NO:25), Ceres CLONE ID no. 1764692 (SEQ ID NO:27),
Ceres ANNOT ID no. 1512656 (SEQ ID NO:41), Public GI ID no.
18403425 (SEQ ID NO:3), Public GI ID no. 11994285 (SEQ ID NO:4),
Public GI ID no. 108706298 (SEQ ID NO:6), Public GI ID no. 15233585
(SEQ ID NO:7), Public GI ID no. 125599279 (SEQ ID NO:8), Public GI
ID no. 115450865 (SEQ ID NO:9), Public GI ID no. 22773244 (SEQ ID
NO:10), Public GI ID no. 92874749 (SEQ ID NO:11), Ceres CLONE ID
no. 1857034 (SEQ ID NO:13), Ceres CLONE ID no. 1059300 (SEQ ID
NO:29), Ceres CLONE ID no. 1804869 (SEQ ID NO:31), Ceres CLONE ID
no. 378863 (SEQ ID NO:33), Ceres CLONE ID no. 1930271 (SEQ ID
NO:35), Ceres CLONE ID no. 2034728 (SEQ ID NO:37), Ceres CLONE ID
no. 1866421 (SEQ ID NO:39), Ceres ANNOT ID no. 1463492 (SEQ ID
NO:43), Ceres ANNOT ID no. 1459391 (SEQ ID NO:45), Ceres ANNOT ID
no. 1442920 (SEQ ID NO:47), Ceres ANNOT ID no. 6011447 (SEQ ID
NO:49), Ceres ANNOT ID no. 6068218 (SEQ ID NO:51), Ceres ANNOT ID
no. 6044121 (SEQ ID NO:53), and Ceres ANNOT ID no. 6100755 (SEQ ID
NO:55).
[0111] Polypeptides are shown in the sequence listing that have HMM
bit scores greater than 70 when fitted to an HMM generated from the
amino acid sequences set forth in FIG. 15. Such polypeptides
include Ceres SEEDLINE ID no. ME02401 (SEQ ID NO:749), Ceres CLONE
ID no. 1847516 (SEQ ID NO:1195), Ceres CLONE ID no. 1961986 (SEQ ID
NO:1201), Ceres ANNOT ID no. 6091930 (SEQ ID NO:1207), Ceres CLONE
ID no. 2025938 (SEQ ID NO:885), Ceres CLONE ID no. 1821214 (SEQ ID
NO:1189), Ceres CLONE ID no. 1040399 (SEQ ID NO:1191), Ceres CLONE
ID no. 1093691 (SEQ ID NO:1193), Ceres CLONE ID no. 974539 (SEQ ID
NO:1197), Ceres CLONE ID no. 1832340 (SEQ ID NO:1199), Ceres CLONE
ID no. 1933211 (SEQ ID NO:1203), Ceres CLONE ID no. 997558 (SEQ ID
NO:1205), Ceres ANNOT ID no. 6041596 (SEQ ID NO:1209), Public GI ID
no. 125564176 (SEQ ID NO:1210), Ceres CLONE ID no. 1836064 (SEQ ID
NO:1212), Ceres CLONE ID no. 1909693 (SEQ ID NO:1214), Public GI ID
no. 169363 (SEQ ID NO:1215), Public GI ID no. 40287480 (SEQ ID
NO:1216), Ceres CLONE ID no. 1765346 (SEQ ID NO:1218), Public GI ID
no. 125546008 (SEQ ID NO:1219), Ceres CLONE ID no. 1950900 (SEQ ID
NO:1221), Public GI ID no. 41350259 (SEQ ID NO:1222), Public GI ID
no. 125588210 (SEQ ID NO:1223), Ceres CLONE ID no. 1954395 (SEQ ID
NO:1225), Public GI ID no. 18403408 (SEQ ID NO:1226), Ceres CLONE
ID no. 2010121 (SEQ ID NO:1228), Ceres ANNOT ID no. 6011486 (SEQ ID
NO:1230), Public GI ID no. 25082726 (SEQ ID NO:1231), Public GI ID
no. 113196593 (SEQ ID NO:1232), Ceres CLONE ID no. 1843021 (SEQ ID
NO:1234), Ceres CLONE ID no. 1931194 (SEQ ID NO:1236), Ceres CLONE
ID no. 1652996 (SEQ ID NO:1238), Ceres CLONE ID no. 1930044 (SEQ ID
NO:1240), Ceres CLONE ID no. 24255 (SEQ ID NO:1242), Ceres ANNOT ID
no. 6034955 (SEQ ID NO:1244), Ceres ANNOT ID no. 6119444 (SEQ ID
NO:1246), Ceres ANNOT ID no. 6063956 (SEQ ID NO:1248), Ceres ANNOT
ID no. 6015461 (SEQ ID NO:1250), Ceres CLONE ID no. 696244 (SEQ ID
NO:1252), Ceres ANNOT ID no. 1468973 (SEQ ID NO:1254), Ceres CLONE
ID no. 2019529 (SEQ ID NO:1256), and Ceres CLONE ID no. 1492169
(SEQ ID NO:1258).
[0112] Polypeptides are shown in the sequence listing that have HMM
bit scores greater than about 300 when fitted to an HMM generated
from the amino acid sequences set forth in FIG. 16. Such
polypeptides include Ceres SEEDLINE ID no. ME004246 (SEQ ID NO:
1311), CeresClone: 971761 (SEQ ID NO: 1313), CeresClone: 1946574
(SEQ ID NO: 1331), CeresClone: 2055551 (SEQ ID NO: 1340),
CeresClone: 100045499 (SEQ ID NO: 1347), CeresClone: 1465853 (SEQ
ID NO: 1353), CeresClone: 753982 (SEQ ID NO: 1360), CeresClone:
1076093 (SEQ ID NO: 1315), CeresClone: 1627875 (SEQ ID NO: 1317),
CeresAnnot: 1508362 (SEQ ID NO: 1319), CeresAnnot: 1526950 (SEQ ID
NO: 1321), CeresClone: 1832333 (SEQ ID NO: 1323), GI: 74272607 (SEQ
ID NO: 1324), GI: 192910782 (SEQ ID NO: 1325), GI: 157342563 (SEQ
ID NO: 1326), GI: 159483497 (SEQ ID NO: 1327), CeresClone: 1839999
(SEQ ID NO: 1329), CeresClone: 467335 (SEQ ID NO: 1333), GI:
168010087 (SEQ ID NO: 1334), CeresClone: 1728202 (SEQ ID NO: 1336),
CeresClone: 1994239 (SEQ ID NO: 1338), CeresAnnot: 8704704 (SEQ ID
NO: 1342), GI: 115453877 (SEQ ID NO: 1343), CeresClone: 1646411
(SEQ ID NO: 1345), CeresClone: 100920260 (SEQ ID NO: 1349), GI:
116786147 (SEQ ID NO: 1350), GI: 116783944 (SEQ ID NO: 1351), GI:
82623397 (SEQ ID NO: 1354), CeresAnnot: 8656662 (SEQ ID NO: 1356),
CeresClone: 1417803 (SEQ ID NO: 1358), and GI: 145345927 (SEQ ID
NO: 1361).
[0113] D. Percent Identity
[0114] In some embodiments, a heat and/or drought-tolerance
polypeptide has an amino acid sequence with at least 20% sequence
identity, e.g., 21%, 22%, 24%, 27%, 30%, 33%, 38%, 45%, 46%, 49%,
50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 97%,
98%, or 99% sequence identity, to one of the amino acid sequences
set forth in SEQ ID NO:442, SEQ ID NO:489, SEQ ID NO:568, SEQ ID
NO:590, SEQ ID NO:705, SEQ ID NO:711, SEQ ID NO:742, SEQ ID
NO:1012, SEQ ID NO:1129, SEQ ID NO:60, SEQ ID NO:364, SEQ ID
NO:311, SEQ ID NO:160, SEQ ID NO:2, SEQ ID NO:749, SEQ ID NO: 1131,
and SEQ ID NO:57. Polypeptides having such a percent sequence
identity often have a domain indicative of a heat and/or
drought-tolerance polypeptide and/or have an HMM bit score that is
greater than 20, as discussed above. Amino acid sequences of heat
and/or drought-tolerance polypeptides having at least 20% sequence
identity to one of the amino acid sequences set forth in SEQ ID
NO:442, SEQ ID NO:489, SEQ ID NO:568, SEQ ID NO:590, SEQ ID NO:705,
SEQ ID NO:711, SEQ ID NO:742, SEQ ID NO:1012, SEQ ID NO:1129, SEQ
ID NO:60, SEQ ID NO:364, SEQ ID NO:311, SEQ ID NO:160, SEQ ID NO:2,
SEQ ID NO: 1131, and SEQ ID NO:749 are provided in FIGS. 1-16.
[0115] "Percent sequence identity" refers to the degree of sequence
identity between any given reference sequence, e.g., SEQ ID NO:2,
and a candidate heat and/or drought-tolerance sequence. A candidate
sequence typically has a length that is from 80 percent to 200
percent of the length of the reference sequence, e.g., 82, 85, 87,
89, 90, 93, 95, 97, 99, 100, 105, 110, 115, 120, 130, 140, 150,
160, 170, 180, 190, or 200 percent of the length of the reference
sequence. A percent identity for any candidate nucleic acid or
polypeptide relative to a reference nucleic acid or polypeptide can
be determined as follows. A reference sequence (e.g., a nucleic
acid sequence or an amino acid sequence) is aligned to one or more
candidate sequences using the computer program ClustalW (version
1.83, default parameters), which allows alignments of nucleic acid
or polypeptide sequences to be carried out across their entire
length (global alignment). Chenna et al., Nucleic Acids Res.,
31(13):3497-500 (2003).
[0116] ClustalW calculates the best match between a reference and
one or more candidate sequences, and aligns them so that
identities, similarities and differences can be determined. Gaps of
one or more residues can be inserted into a reference sequence, a
candidate sequence, or both, to maximize sequence alignments. For
fast pairwise alignment of nucleic acid sequences, the following
default parameters are used: word size: 2; window size: 4; scoring
method: percentage; number of top diagonals: 4; and gap penalty: 5.
For multiple alignment of nucleic acid sequences, the following
parameters are used: gap opening penalty: 10.; gap extension
penalty: 5.0; and weight transitions: yes. For fast pairwise
alignment of protein sequences, the following parameters are used:
word size: 1; window size: 5; scoring method: percentage; number of
top diagonals: 5; gap penalty: 3. For multiple alignment of protein
sequences, the following parameters are used: weight matrix:
blosum; gap opening penalty: 10.; gap extension penalty: 0.5;
hydrophilic gaps: on; hydrophilic residues: Gly, Pro, Ser, Asn,
Asp, Gln, Glu, Arg, and Lys; residue-specific gap penalties: on.
The ClustalW output is a sequence alignment that reflects the
relationship between sequences. ClustalW can be run, for example,
at the Baylor College of Medicine Search Launcher site
(searchlauncher.bcm.tmc.edu/multi-align/multi-align.html) and at
the European Bioinformatics Institute site on the World Wide Web
(ebi.ac.uk/clustalw).
[0117] To determine percent identity of a candidate nucleic acid or
amino acid sequence to a reference sequence, the sequences are
aligned using ClustalW, the number of identical matches in the
alignment is divided by the length of the reference sequence, and
the result is multiplied by 100. It is noted that the percent
identity value can be rounded to the nearest tenth. For example,
78.11, 78.12, 78.13, and 78.14 are rounded down to 78.1, while
78.15, 78.16, 78.17, 78.18, and 78.19 are rounded up to 78.2.
[0118] In some cases, a drought-tolerance polypeptide has an amino
acid sequence with at least 40% sequence identity, e.g., 50%, 52%,
56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 97%, 98%, or 99%
sequence identity, to the amino acid sequence set forth in SEQ ID
NO:442. Amino acid sequences of polypeptides having greater than
40% sequence identity to the polypeptide set forth in SEQ ID NO:442
are provided in FIG. 1. Such polypeptides include Public GI ID no.
72384401 (SEQ ID NO:443), Ceres CLONE ID no. 1079382 (SEQ ID
NO:445), Ceres CLONE ID no. 1853461 (SEQ ID NO:447), Ceres CLONE ID
no. 1626485 (SEQ ID NO:451), Ceres CLONE ID no. 1713920 (SEQ ID
NO:459), Ceres CLONE ID no. 1772747 (SEQ ID NO:463), Ceres CLONE ID
no. 225960 (SEQ ID NO:465), Public GI ID no. 115443807 (SEQ ID
NO:470), and Ceres CLONE ID no. 569388 (SEQ ID NO:483).
[0119] In some cases, a drought-tolerance polypeptide has an amino
acid sequence with at least 30% sequence identity, e.g., 30%, 35%,
40%, 48%, 50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%,
95%, 97%, 98%, or 99% sequence identity, to the amino acid sequence
set forth in SEQ ID NO:489. Amino acid sequences of polypeptides
having greater than 30% sequence identity to the polypeptide set
forth in SEQ ID NO:489 are provided in FIG. 2. Such polypeptides
include Ceres CLONE ID no. 571200 (SEQ ID NO:494), Ceres CLONE ID
no. 1928532 (SEQ ID NO:496), Ceres ANNOT ID no. 1490637 (SEQ ID
NO:498), Ceres CLONE ID no. 295496 (SEQ ID NO:514), Public GI ID
no. 115463637 (SEQ ID NO:515), and Ceres CLONE ID no. 1999444 (SEQ
ID NO:518).
[0120] In some cases, a drought-tolerance polypeptide has an amino
acid sequence with at least 35% sequence identity, e.g., 37%, 40%,
45%, 50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%, 95%,
97%, 98%, or 99% sequence identity, to the amino acid sequence set
forth in SEQ ID NO:568. Amino acid sequences of polypeptides having
greater than 35% sequence identity to the polypeptide set forth in
SEQ ID NO:568 are provided in FIG. 3. Such polypeptides include
Ceres CLONE ID no. 695006 (SEQ ID NO:578) and Ceres ANNOT ID no.
1527488 (SEQ ID NO:584).
[0121] In some cases, a drought-tolerance polypeptide has an amino
acid sequence with at least 25% sequence identity, e.g., 28%, 32%,
40%, 44%, 48%, 50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%,
90%, 95%, 97%, 98%, or 99% sequence identity, to the amino acid
sequence set forth in SEQ ID NO:590. Amino acid sequences of
polypeptides having greater than 25% sequence identity to the
polypeptide set forth in SEQ ID NO:590 are provided in FIG. 4. Such
polypeptides include Ceres ANNOT ID no. 1501305 (SEQ ID NO:623),
Ceres CLONE ID no. 607280 (SEQ ID NO:627), Public GI ID no.
92887174 (SEQ ID NO:632), Ceres CLONE ID no. 1857162 (SEQ ID
NO:634), Public GI ID no. 115477272 (SEQ ID NO:674), and Ceres
CLONE ID no. 264002 (SEQ ID NO:682).
[0122] In some cases, a drought-tolerance polypeptide has an amino
acid sequence with at least 40% sequence identity, e.g., 50%, 52%,
56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 97%, 98%, or 99%
sequence identity, to the amino acid sequence set forth in SEQ ID
NO:705. Amino acid sequences of polypeptides having greater than
40% sequence identity to the polypeptide set forth in SEQ ID NO:705
are provided in FIG. 5. Such polypeptides include Public GI ID no.
79320952 (SEQ ID NO:706) and Public GI ID no. 79320957 (SEQ ID
NO:707).
[0123] In some cases, a drought-tolerance polypeptide has an amino
acid sequence with at least 30% sequence identity, e.g., 33%, 41%,
45%, 50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%, 95%,
97%, 98%, or 99% sequence identity, to the amino acid sequence set
forth in SEQ ID NO:711. Amino acid sequences of polypeptides having
greater than 30% sequence identity to the polypeptide set forth in
SEQ ID NO:711 are provided in FIG. 6. Such polypeptides include
Ceres CLONE ID no. 964616 (SEQ ID NO:714), Ceres CLONE ID no.
100009667 (SEQ ID NO:715), Ceres ANNOT ID no. 1444568 (SEQ ID
NO:719), Ceres CLONE ID no. 719489 (SEQ ID NO:725), Ceres CLONE ID
no. 587748 (SEQ ID NO:733), Public GI ID no. 125528114 (SEQ ID
NO:734), and Ceres CLONE ID no. 274172 (SEQ ID NO:737).
[0124] In some cases, a drought-tolerance polypeptide has an amino
acid sequence with at least 30% sequence identity, e.g., 35%, 38%,
43%, 47%, 50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%,
95%, 97%, 98%, or 99% sequence identity, to the amino acid sequence
set forth in SEQ ID NO:742. Amino acid sequences of polypeptides
having greater than 30% sequence identity to the polypeptide set
forth in SEQ ID NO:742 are provided in FIG. 7. Such polypeptides
include Public GI ID no. 115477170 (SEQ ID NO:757), Ceres CLONE ID
no. 1620215 (SEQ ID NO:759), Ceres CLONE ID no. 1931889 (SEQ ID
NO:761), Ceres ANNOT ID no. 1460527 (SEQ ID NO:801), Ceres CLONE ID
no. 975913 (SEQ ID NO:846), Ceres CLONE ID no. 708446 (SEQ ID
NO:859), Ceres CLONE ID no. 1748922 (SEQ ID NO:944), Ceres CLONE ID
no. 1775820 (SEQ ID NO:955), Public GI ID no. 115468934 (SEQ ID
NO:991), and Public GI ID no. 118424243 (SEQ ID NO:1000).
[0125] In some cases, a drought-tolerance polypeptide has an amino
acid sequence with at least 35% sequence identity, e.g., 36%, 40%,
47%, 50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%, 95%,
97%, 98%, or 99% sequence identity, to the amino acid sequence set
forth in SEQ ID NO:1012. Amino acid sequences of polypeptides
having greater than 35% sequence identity to the polypeptide set
forth in SEQ ID NO:1012 are provided in FIG. 8. Such polypeptides
include Ceres CLONE ID no. 287430 (SEQ ID NO:1018), Ceres ANNOT ID
no. 451889 (SEQ ID NO:1020), Ceres CLONE ID no. 936084 (SEQ ID
NO:1055), Ceres CLONE ID no. 1792501 (SEQ ID NO:1066), Ceres ANNOT
ID no. 1437875 (SEQ ID NO:1068), Public GI ID no. 1853968 (SEQ ID
NO:1093), Public GI ID no. 27530032 (SEQ ID NO:1094), Ceres CLONE
ID no. 1834483 (SEQ ID NO:1110), Public GI ID no. 84579418 (SEQ ID
NO:1113), Public GI ID no. 15077030 (SEQ ID NO:1117), and Public GI
ID no. 13752458 (SEQ ID NO:1118).
[0126] In some cases, a drought-tolerance polypeptide has an amino
acid sequence with at least 30% sequence identity, e.g., 31%, 35%,
40%, 41%, 49%, 50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%,
90%, 95%, 97%, 98%, or 99% sequence identity, to the amino acid
sequence set forth in SEQ ID NO:1129. Amino acid sequences of
polypeptides having greater than 30% sequence identity to the
polypeptide set forth in SEQ ID NO:1129 are provided in FIG. 9.
Such polypeptides include Public GI ID no. 72255610 (SEQ ID
NO:933), Ceres SEEDLINE ID no. ME20095 (SEQ ID NO:1129), Ceres
CLONE ID no. 1562633 (SEQ ID NO:1140), Ceres ANNOT ID no. 1692728
(SEQ ID NO:1142), Ceres CLONE ID no. 921110 (SEQ ID NO:1144), Ceres
CLONE ID no. 1791180 (SEQ ID NO:1146), Ceres CLONE ID no. 527891
(SEQ ID NO:1148), Ceres ANNOT ID no. 1522414 (SEQ ID NO:1150), and
Public GI ID no. 51458330 (SEQ ID NO:1182).
[0127] In some cases, a heat-tolerance polypeptide has an amino
acid sequence with at least 25% sequence identity, e.g., 26%, 27%,
30%, 32%, 40%, 42%, 47%, 50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%,
80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity, to the
amino acid sequence set forth in SEQ ID NO:60. Amino acid sequences
of polypeptides having greater than 25% sequence identity to the
polypeptide set forth in SEQ ID NO:60 are provided in FIG. 10. Such
polypeptides include Ceres CLONE ID no. 872030 (SEQ ID NO:62),
Ceres CLONE ID no. 100029223 (SEQ ID NO:74), Ceres CLONE ID no.
1939845 (SEQ ID NO:98), Ceres CLONE ID no. 2015383 (SEQ ID NO:102),
Ceres CLONE ID no. 1607893 (SEQ ID NO:104), Ceres CLONE ID no.
1075133 (SEQ ID NO:152), and Ceres CLONE ID no. 1218065 (SEQ ID
NO:915).
[0128] In some cases, a heat-tolerance polypeptide has an amino
acid sequence with at least 25% sequence identity, e.g., 27%, 28%,
31%, 36%, 44%, 50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%,
90%, 95%, 97%, 98%, or 99% sequence identity, to the amino acid
sequence set forth in SEQ ID NO:364. Amino acid sequences of
polypeptides having greater than 25% sequence identity to the
polypeptide set forth in SEQ ID NO:364 are provided in FIG. 11.
Such polypeptides include Ceres ANNOT ID no. 1455221 (SEQ ID
NO:407), Public GI ID no. 729774 (SEQ ID NO:408), Ceres CLONE ID
no. 1414288 (SEQ ID NO:410), Public GI ID no. 115482048 (SEQ ID
NO:413), and Public GI ID no. 56117815 (SEQ ID NO:421).
[0129] In some cases, a heat-tolerance polypeptide has an amino
acid sequence with at least 25% sequence identity, e.g., 29%, 35%,
40%, 47%, 50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%,
95%, 97%, 98%, or 99% sequence identity, to the amino acid sequence
set forth in SEQ ID NO:311. Amino acid sequences of polypeptides
having greater than 25% sequence identity to the polypeptide set
forth in SEQ ID NO:311 are provided in FIG. 12. Such polypeptides
include Ceres ANNOT ID no. 1538958 (SEQ ID NO:319), Public GI ID
no. 115521213 (SEQ ID NO:320), Public GI ID no. 42415865 (SEQ ID
NO:321), Public GI ID no. 729775 (SEQ ID NO:322), Public GI ID no.
11386827 (SEQ ID NO:323), Public GI ID no. 115456675 (SEQ ID
NO:326), and Public GI ID no. 89274218 (SEQ ID NO:335).
[0130] In some cases, a heat-tolerance polypeptide has an amino
acid sequence with at least 25% sequence identity, e.g., 30%, 33%,
40%, 45%, 50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%, 90%,
95%, 97%, 98%, or 99% sequence identity, to the amino acid sequence
set forth in SEQ ID NO:160. Amino acid sequences of polypeptides
having greater than 25% sequence identity to the polypeptide set
forth in SEQ ID NO:160 are provided in FIG. 13. Such polypeptides
include Ceres CLONE ID no. 1837065 (SEQ ID NO:162), Ceres ANNOT ID
no. 1531178 (SEQ ID NO:184), Ceres CLONE ID no. 470694 (SEQ ID
NO:242), Public GI ID no. 92867368 (SEQ ID NO:253), Ceres CLONE ID
no. 859707 (SEQ ID NO:256), Ceres CLONE ID no. 392275 (SEQ ID
NO:268), Ceres CLONE ID no. 1828394 (SEQ ID NO:282), and Public GI
ID no. 115466694 (SEQ ID NO:294).
[0131] In some cases, a heat-tolerance polypeptide has an amino
acid sequence with at least 20% sequence identity, e.g., 21%, 22%,
30%, 35%, 40%, 45%, 50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%,
85%, 90%, 95%, 97%, 98%, or 99% sequence identity, to the amino
acid sequence set forth in SEQ ID NO:2. Amino acid sequences of
polypeptides having greater than 20% sequence identity to the
polypeptide set forth in SEQ ID NO:2 are provided in FIG. 14. Such
polypeptides include Public GI ID no. 115470807 (SEQ ID NO:5),
Ceres CLONE ID no. 1842931 (SEQ ID NO:15), Ceres CLONE ID no.
321308 (SEQ ID NO:17), Ceres CLONE ID no. 1725811 (SEQ ID NO:19),
Ceres CLONE ID no. 1357455 (SEQ ID NO:21), Ceres CLONE ID no.
943370 (SEQ ID NO:23), Ceres CLONE ID no. 1327712 (SEQ ID NO:25),
Ceres CLONE ID no. 1764692 (SEQ ID NO:27), and Ceres ANNOT ID no.
1512656 (SEQ ID NO:41).
[0132] In some cases, a drought-tolerance polypeptide has an amino
acid sequence with at least 20% sequence identity, e.g., 22%, 23%,
30%, 35%, 40%, 50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%,
90%, 95%, 97%, 98%, or 99% sequence identity, to the amino acid
sequence set forth in SEQ ID NO:749. Amino acid sequences of
polypeptides having greater than 20% sequence identity to the
polypeptide set forth in SEQ ID NO:749 are provided in FIG. 15.
Such polypeptides include Ceres CLONE ID no. 1847516 (SEQ ID
NO:1195), Ceres CLONE ID no. 1961986 (SEQ ID NO:1201), and Ceres
ANNOT ID no. 6091930 (SEQ ID NO:1207).
[0133] In some cases, a drought-tolerance polypeptide has an amino
acid sequence with at least 30% sequence identity, e.g., 31%, 35%,
40%, 41%, 49%, 50%, 52%, 56%, 59%, 61%, 65%, 70%, 75%, 80%, 85%,
90%, 95%, 97%, 98%, or 99% sequence identity, to the amino acid
sequence set forth in SEQ ID NO:1311. Amino acid sequences of
polypeptides having greater than 30% sequence identity to the
polypeptide set forth in SEQ ID NO:1311 are provided in FIG. 16.
Such polypeptides include CeresClone: 971761 (SEQ ID NO: 1313),
CeresClone: 1946574 (SEQ ID NO: 1331), CeresClone: 2055551 (SEQ ID
NO: 1340), CeresClone: 100045499 (SEQ ID NO: 1347), CeresClone:
1465853 (SEQ ID NO: 1353), CeresClone: 753982 (SEQ ID NO: 1360),
CeresClone: 1076093 (SEQ ID NO: 1315), CeresClone: 1627875 (SEQ ID
NO: 1317), CeresAnnot: 1508362 (SEQ ID NO: 1319), CeresAnnot:
1526950 (SEQ ID NO: 1321), CeresClone: 1832333 (SEQ ID NO: 1323),
GI: 74272607 (SEQ ID NO: 1324), GI: 192910782 (SEQ ID NO: 1325),
GI: 157342563 (SEQ ID NO: 1326), GI: 159483497 (SEQ ID NO: 1327),
CeresClone: 1839999 (SEQ ID NO: 1329), CeresClone: 467335 (SEQ ID
NO: 1333), GI: 168010087 (SEQ ID NO: 1334), CeresClone: 1728202
(SEQ ID NO: 1336), CeresClone: 1994239 (SEQ ID NO: 1338),
CeresAnnot: 8704704 (SEQ ID NO: 1342), GI: 115453877 (SEQ ID NO:
1343), CeresClone: 1646411 (SEQ ID NO: 1345), CeresClone: 100920260
(SEQ ID NO: 1349), GI: 116786147 (SEQ ID NO: 1350), GI: 116783944
(SEQ ID NO: 1351), GI: 82623397 (SEQ ID NO: 1354), CeresAnnot:
8656662 (SEQ ID NO: 1356), CeresClone: 1417803 (SEQ ID NO: 1358),
and GI: 145345927 (SEQ ID NO: 1361).
[0134] E. Other Sequences
[0135] It should be appreciated that a heat and/or
drought-tolerance polypeptide can include additional amino acids
that are not involved in heat and/or drought-tolerance, and thus
such a polypeptide can be longer than would otherwise be the case.
For example, a heat and/or drought-tolerance polypeptide can
include a purification tag, a chloroplast transit peptide, a
mitochondrial transit peptide, an amyloplast peptide, or a leader
sequence added to the amino or carboxy terminus. In some
embodiments, a heat and/or drought-tolerance polypeptide includes
an amino acid sequence that functions as a reporter, e.g., a green
fluorescent protein or yellow fluorescent protein.
III. NUCLEIC ACIDS
[0136] Nucleic acids described herein include nucleic acids that
are effective to increase heat and/or drought tolerance when
transcribed in a plant or plant cell. Such nucleic acids include
those that encode a heat and/or drought-tolerance polypeptide.
[0137] A. Nucleic Acids Encoding Heat and/or Drought-Tolerance
Polypeptides
[0138] Nucleic acids encoding heat and/or drought-tolerance
polypeptides are described herein. Such nucleic acids include SEQ
ID NO:441, SEQ ID NO:488, SEQ ID NO:567, SEQ ID NO:589, SEQ ID
NO:703, SEQ ID NO:704, SEQ ID NO:710, SEQ ID NO:1241, SEQ ID
NO:1260, SEQ ID NO:1126, SEQ ID NO:1127, SEQ ID NO:1128, SEQ ID
NO:58, SEQ ID NO:59, SEQ ID NO:363, SEQ ID NO:310, SEQ ID NO:159,
SEQ ID NO:1, SEQ ID NO:1188, SEQ ID NO:1311, and SEQ ID NO:56, as
described in more detail below. A nucleic acid also can be a
fragment that is at least 40% (e.g., at least 45, 50, 55, 60, 65,
70, 75, 80, 85, 90, 95, or 99%) of the length of the full-length
nucleic acid set forth in SEQ ID NO:441, SEQ ID NO:488, SEQ ID
NO:567, SEQ ID NO:589, SEQ ID NO:703, SEQ ID NO:704, SEQ ID NO:710,
SEQ ID NO:1241, SEQ ID NO:1260, SEQ ID NO:1126, SEQ ID NO:1127, SEQ
ID NO:1128, SEQ ID NO:58, SEQ ID NO:59, SEQ ID NO:363, SEQ ID
NO:310, SEQ ID NO:159, SEQ ID NO:1, SEQ ID NO:1008, and SEQ ID
NO:56.
[0139] A drought-tolerance nucleic acid can comprise the nucleotide
sequence set forth in SEQ ID NO:441. Alternatively, a
drought-tolerance nucleic acid can be a variant of the nucleic acid
having the nucleotide sequence set forth in SEQ ID NO:441. For
example, a drought-tolerance nucleic acid can have a nucleotide
sequence with at least 80% sequence identity, e.g., 81%, 85%, 90%,
95%, 97%, 98%, or 99% sequence identity, to the nucleotide sequence
set forth in SEQ ID NO:441 or a functional homolog thereof such as
those identified in the sequence listing.
[0140] A drought-tolerance nucleic acid can comprise the nucleotide
sequence set forth in SEQ ID NO:488. Alternatively, a
drought-tolerance nucleic acid can be a variant of the nucleic acid
having the nucleotide sequence set forth in SEQ ID NO:488. For
example, a drought-tolerance nucleic acid can have a nucleotide
sequence with at least 80% sequence identity, e.g., 81%, 85%, 90%,
95%, 97%, 98%, or 99% sequence identity, to the nucleotide sequence
set forth in SEQ ID NO:488 or a functional homolog thereof such as
those identified in the sequence listing.
[0141] A drought-tolerance nucleic acid can comprise the nucleotide
sequence set forth in SEQ ID NO:567. Alternatively, a
drought-tolerance nucleic acid can be a variant of the nucleic acid
having the nucleotide sequence set forth in SEQ ID NO:567. For
example, a drought-tolerance nucleic acid can have a nucleotide
sequence with at least 80% sequence identity, e.g., 81%, 85%, 90%,
95%, 97%, 98%, or 99% sequence identity, to the nucleotide sequence
set forth in SEQ ID NO:567 or a functional homolog thereof such as
those identified in the sequence listing.
[0142] A drought-tolerance nucleic acid can comprise the nucleotide
sequence set forth in SEQ ID NO:589. Alternatively, a
drought-tolerance nucleic acid can be a variant of the nucleic acid
having the nucleotide sequence set forth in SEQ ID NO:589. For
example, a drought-tolerance nucleic acid can have a nucleotide
sequence with at least 80% sequence identity, e.g., 81%, 85%, 90%,
95%, 97%, 98%, or 99% sequence identity, to the nucleotide sequence
set forth in SEQ ID NO:589 or a functional homolog thereof such as
those identified in the sequence listing.
[0143] A drought-tolerance nucleic acid can comprise the nucleotide
sequence set forth in SEQ ID NO:703. Alternatively, a
drought-tolerance nucleic acid can be a variant of the nucleic acid
having the nucleotide sequence set forth in SEQ ID NO:703. For
example, a drought-tolerance nucleic acid can have a nucleotide
sequence with at least 80% sequence identity, e.g., 81%, 85%, 90%,
95%, 97%, 98%, or 99% sequence identity, to the nucleotide sequence
set forth in SEQ ID NO:703 or a functional homolog thereof such as
those identified in the sequence listing.
[0144] A drought-tolerance nucleic acid can comprise the nucleotide
sequence set forth in SEQ ID NO:710. Alternatively, a
drought-tolerance nucleic acid can be a variant of the nucleic acid
having the nucleotide sequence set forth in SEQ ID NO:710. For
example, a drought-tolerance nucleic acid can have a nucleotide
sequence with at least 80% sequence identity, e.g., 81%, 85%, 90%,
95%, 97%, 98%, or 99% sequence identity, to the nucleotide sequence
set forth in SEQ ID NO:710 or a functional homolog thereof such as
those identified in the sequence listing.
[0145] A drought-tolerance nucleic acid can comprise the nucleotide
sequence set forth in SEQ ID NO:1241. Alternatively, a
drought-tolerance nucleic acid can be a variant of the nucleic acid
having the nucleotide sequence set forth in SEQ ID NO:1241. For
example, a drought-tolerance nucleic acid can have a nucleotide
sequence with at least 80% sequence identity, e.g., 81%, 85%, 90%,
95%, 97%, 98%, or 99% sequence identity, to the nucleotide sequence
set forth in SEQ ID NO:1241 or a functional homolog thereof such as
those identified in the sequence listing.
[0146] A drought-tolerance nucleic acid can comprise the nucleotide
sequence set forth in SEQ ID NO:1260. Alternatively, a
drought-tolerance nucleic acid can be a variant of the nucleic acid
having the nucleotide sequence set forth in SEQ ID NO:1260. For
example, a drought-tolerance nucleic acid can have a nucleotide
sequence with at least 80% sequence identity, e.g., 81%, 85%, 90%,
95%, 97%, 98%, or 99% sequence identity, to the nucleotide sequence
set forth in SEQ ID NO:1260 or a functional homolog thereof such as
those identified in the sequence listing.
[0147] A drought-tolerance nucleic acid can comprise the nucleotide
sequence set forth in SEQ ID NO:1126. Alternatively, a
drought-tolerance nucleic acid can be a variant of the nucleic acid
having the nucleotide sequence set forth in SEQ ID NO:1126. For
example, a drought-tolerance nucleic acid can have a nucleotide
sequence with at least 80% sequence identity, e.g., 81%, 85%, 90%,
95%, 97%, 98%, or 99% sequence identity, to the nucleotide sequence
set forth in SEQ ID NO:1126 or a functional homolog thereof such as
those identified in the sequence listing.
[0148] A heat-tolerance nucleic acid can comprise the nucleotide
sequence set forth in SEQ ID NO:58. Alternatively, a heat-tolerance
nucleic acid can be a variant of the nucleic acid having the
nucleotide sequence set forth in SEQ ID NO:58. For example, a
heat-tolerance nucleic acid can have a nucleotide sequence with at
least 80% sequence identity, e.g., 81%, 85%, 90%, 95%, 97%, 98%, or
99% sequence identity, to the nucleotide sequence set forth in SEQ
ID NO:58 or a functional homolog thereof such as those identified
in the sequence listing.
[0149] A heat-tolerance nucleic acid can comprise the nucleotide
sequence set forth in SEQ ID NO:363. Alternatively, a
heat-tolerance nucleic acid can be a variant of the nucleic acid
having the nucleotide sequence set forth in SEQ ID NO:363. For
example, a heat-tolerance nucleic acid can have a nucleotide
sequence with at least 80% sequence identity, e.g., 81%, 85%, 90%,
95%, 97%, 98%, or 99% sequence identity, to the nucleotide sequence
set forth in SEQ ID NO:363 or a functional homolog thereof such as
those identified in the sequence listing.
[0150] A heat-tolerance nucleic acid can comprise the nucleotide
sequence set forth in SEQ ID NO:310. Alternatively, a
heat-tolerance nucleic acid can be a variant of the nucleic acid
having the nucleotide sequence set forth in SEQ ID NO:310. For
example, a heat-tolerance nucleic acid can have a nucleotide
sequence with at least 80% sequence identity, e.g., 81%, 85%, 90%,
95%, 97%, 98%, or 99% sequence identity, to the nucleotide sequence
set forth in SEQ ID NO:310 or a functional homolog thereof such as
those identified in the sequence listing.
[0151] A heat-tolerance nucleic acid can comprise the nucleotide
sequence set forth in SEQ ID NO:159. Alternatively, a
heat-tolerance nucleic acid can be a variant of the nucleic acid
having the nucleotide sequence set forth in SEQ ID NO:159. For
example, a heat-tolerance nucleic acid can have a nucleotide
sequence with at least 80% sequence identity, e.g., 81%, 85%, 90%,
95%, 97%, 98%, or 99% sequence identity, to the nucleotide sequence
set forth in SEQ ID NO:159 or a functional homolog thereof such as
those identified in the sequence listing.
[0152] A heat-tolerance nucleic acid can comprise the nucleotide
sequence set forth in SEQ ID NO:1. Alternatively, a heat-tolerance
nucleic acid can be a variant of the nucleic acid having the
nucleotide sequence set forth in SEQ ID NO:1. For example, a
heat-tolerance nucleic acid can have a nucleotide sequence with at
least 80% sequence identity, e.g., 81%, 85%, 90%, 95%, 97%, 98%, or
99% sequence identity, to the nucleotide sequence set forth in SEQ
ID NO:1 or a functional homolog thereof such as those identified in
the sequence listing.
[0153] A drought-tolerance nucleic acid can comprise the nucleotide
sequence set forth in SEQ ID NO:1008. Alternatively, a
drought-tolerance nucleic acid can be a variant of the nucleic acid
having the nucleotide sequence set forth in SEQ ID NO:1008. For
example, a drought-tolerance nucleic acid can have a nucleotide
sequence with at least 80% sequence identity, e.g., 81%, 85%, 90%,
95%, 97%, 98%, or 99% sequence identity, to the nucleotide sequence
set forth in SEQ ID NO:1008 or a functional homolog thereof such as
those identified in the sequence listing.
[0154] A heat-tolerance nucleic acid can comprise the nucleotide
sequence set forth in SEQ ID NO:56. Alternatively, a heat-tolerance
nucleic acid can be a variant of the nucleic acid having the
nucleotide sequence set forth in SEQ ID NO:56. For example, a
heat-tolerance nucleic acid can have a nucleotide sequence with at
least 80% sequence identity, e.g., 81%, 85%, 90%, 95%, 97%, 98%, or
99% sequence identity, to the nucleotide sequence set forth in SEQ
ID NO:56.
[0155] A drought-tolerance nucleic acid can comprise the nucleotide
sequence set forth in SEQ ID NO:1362. Alternatively, a
drought-tolerance nucleic acid can be a variant of the nucleic acid
having the nucleotide sequence set forth in SEQ ID NO:1362. For
example, a drought-tolerance nucleic acid can have a nucleotide
sequence with at least 30% sequence identity, e.g., 35%, 40%, 45%,
50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 97%, 98%, or 99%
sequence identity, to the nucleotide sequence set forth in SEQ ID
NO:1362 or a functional homolog thereof such as those identified in
the sequence listing.
[0156] Isolated nucleic acid molecules can be produced by standard
techniques. For example, polymerase chain reaction (PCR) techniques
can be used to obtain an isolated nucleic acid containing a
nucleotide sequence described herein. PCR can be used to amplify
specific sequences from DNA as well as RNA, including sequences
from total genomic DNA or total cellular RNA. Various PCR methods
are described, for example, in PCR Primer: A Laboratory Manual,
Dieffenbach and Dveksler, eds., Cold Spring Harbor Laboratory
Press, 1995. Generally, sequence information from the ends of the
region of interest or beyond is employed to design oligonucleotide
primers that are identical or similar in sequence to opposite
strands of the template to be amplified. Various PCR strategies
also are available by which site-specific nucleotide sequence
modifications can be introduced into a template nucleic acid.
Isolated nucleic acids also can be chemically synthesized, either
as a single nucleic acid molecule (e.g., using automated DNA
synthesis in the 3' to 5' direction using phosphoramidite
technology) or as a series of oligonucleotides. For example, one or
more pairs of long oligonucleotides (e.g., >100 nucleotides) can
be synthesized that contain the desired sequence, with each pair
containing a short segment of complementarity (e.g., about 15
nucleotides) such that a duplex is formed when the oligonucleotide
pair is annealed. DNA polymerase is used to extend the
oligonucleotides, resulting in a single, double-stranded nucleic
acid molecule per oligonucleotide pair, which then can be ligated
into a vector. Isolated nucleic acids of the invention also can be
obtained by mutagenesis of, e.g., a naturally occurring DNA.
[0157] B. Use of Nucleic Acids to Modulate Expression of
Polypeptides
Expression of a Heat and/or Drought-Tolerance Polypeptide
[0158] A nucleic acid encoding one of the heat and/or
drought-tolerance polypeptides described herein can be used to
express the polypeptide in a plant species of interest, typically
by transforming a plant cell with a nucleic acid having the coding
sequence for the polypeptide operably linked in sense orientation
to one or more regulatory regions. Suitable polynucleotides include
full-length nucleic acids encoding heat and/or drought-tolerance
polypeptides or fragments of such full-length nucleic acids. In
some embodiments, a complement of the full-length nucleic acid or a
fragment thereof can be used. Typically, a fragment is at least 50
nucleotides, e.g., at least 50, 55, 60, 75, 100, 200, 300, 500,
600, 700, 1000, 2000 nucleotides or more.
[0159] It will be appreciated that because of the degeneracy of the
genetic code, a number of nucleic acids can encode a particular
heat and/or drought-tolerance polypeptide; i.e., for many amino
acids, there is more than one nucleotide triplet that serves as the
codon for the amino acid. Thus, codons in the coding sequence for a
given heat and/or drought-tolerance polypeptide can be modified
such that optimal expression in a particular plant species is
obtained, using appropriate codon bias tables for that species.
[0160] In some cases, expression of a heat and/or drought-tolerance
polypeptide inhibits one or more functions of an endogenous
polypeptide. For example, a nucleic acid that encodes a dominant
negative polypeptide can be used to inhibit protein function. A
dominant negative polypeptide typically is mutated or truncated
relative to an endogenous wild type polypeptide, and its presence
in a cell inhibits one or more functions of the wild type
polypeptide in that cell, i.e., the dominant negative polypeptide
is genetically dominant and confers a loss of function. The
mechanism by which a dominant negative polypeptide confers such a
phenotype can vary but often involves a protein-protein interaction
or a protein-DNA interaction. For example, a dominant negative
polypeptide can be an enzyme that is truncated relative to a native
wild type enzyme, such that the truncated polypeptide retains
domains involved in binding a first protein but lacks domains
involved in binding a second protein. The truncated polypeptide is
thus unable to properly modulate the activity of the second
protein. See, e.g., US 2007/0056058. As another example, a point
mutation that results in a non-conservative amino acid substitution
in a catalytic domain can result in a dominant negative
polypeptide. See, e.g., US 2005/032221. As another example, a
dominant negative polypeptide can be a transcription factor that is
truncated relative to a native wild type transcription factor, such
that the truncated polypeptide retains the DNA binding domain(s)
but lacks the activation domain(s). Such a truncated polypeptide
can inhibit the wild type transcription factor from binding DNA,
thereby inhibiting transcription activation.
[0161] C. Constructs/Vectors
[0162] Recombinant constructs provided herein can be used to
transform plants or plant cells in order to increase drought and/or
heat tolerance. A recombinant nucleic acid construct can comprise a
nucleic acid encoding a heat and/or drought-tolerance polypeptide
as described herein, operably linked to a regulatory region
suitable for expressing the heat and/or drought-tolerance
polypeptide in the plant or cell. Thus, a nucleic acid can comprise
a coding sequence that encodes any of the heat and/or
drought-tolerance polypeptides as set forth in SEQ ID NO: 2, 3, 4,
5, 6, 7, 8, 9, 10, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33,
35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 60, 62, 63, 64, 65,
66, 67, 68, 69, 70, 71, 73, 74, 76, 78, 80, 82, 84, 86, 88, 90, 92,
94, 96, 98, 100, 102, 104, 106, 108, 110, 112, 114, 116, 118, 120,
122, 124, 126, 127, 130, 132, 134, 136, 138, 139, 142, 144, 146,
148, 150, 152, 154, 156, 158, 160, 162, 164, 166, 168, 170, 172,
174, 176, 178, 179, 180, 182, 184, 186, 188, 190, 192, 194, 196,
198, 200, 202, 204, 206, 208, 209, 212, 214, 216, 218, 220, 222,
224, 225, 228, 229, 231, 233, 234, 235, 237, 239, 240, 242, 244,
246, 248, 250, 252, 253, 254, 256, 258, 260, 262, 264, 266, 268,
269, 271, 273, 275, 277, 279, 280, 282, 284, 286, 288, 289, 290,
291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303,
304, 305, 307, 309, 311, 312, 314, 315, 316, 317, 319, 320, 321,
322, 323, 325, 326, 327, 329, 330, 332, 334, 335, 336, 337, 338,
339, 340, 341, 343, 344, 346, 348, 349, 350, 351, 353, 355, 356,
358, 360, 362, 364, 366, 368, 369, 370, 371, 373, 375, 377, 379,
381, 383, 385, 387, 389, 390, 392, 394, 396, 398, 400, 401, 402,
404, 405, 407, 408, 410, 412, 413, 414, 416, 418, 419, 420, 421,
422, 423, 424, 426, 428, 430, 431, 432, 433, 434, 435, 436, 438,
440, 442, 443, 445, 447, 449, 451, 453, 455, 457, 459, 461, 463,
465, 467, 469, 470, 472, 474, 476, 478, 480, 481, 483, 485, 487,
489, 490, 491, 492, 494, 496, 498, 500, 502, 504, 506, 508, 510,
512, 514, 515, 516, 518, 519, 520, 522, 524, 526, 528, 530, 531,
533, 535, 536, 538, 540, 541, 543, 545, 547, 548, 550, 552, 553,
555, 557, 558, 559, 561, 562, 564, 566, 568, 569, 570, 572, 574,
576, 578, 580, 582, 584, 586, 588, 590, 591, 592, 593, 594, 595,
596, 597, 598, 599, 600, 602, 603, 604, 605, 607, 608, 610, 612,
614, 615, 616, 617, 618, 619, 620, 621, 623, 625, 627, 629, 631,
632, 634, 635, 636, 638, 640, 642, 644, 646, 647, 648, 650, 651,
653, 654, 655, 656, 658, 660, 661, 662, 663, 664, 665, 666, 667,
668, 669, 671, 673, 674, 675, 676, 678, 680, 682, 684, 685, 687,
688, 690, 691, 692, 694, 696, 698, 700, 702, 705, 706, 707, 708,
709, 711, 712, 714, 715, 717, 719, 721, 723, 725, 726, 727, 729,
731, 733, 734, 735, 737, 738, 740, 741, 742, 743, 744, 745, 746,
747, 748, 749, 751, 752, 753, 754, 755, 756, 757, 759, 760, 761,
763, 765, 767, 768, 769, 771, 773, 774, 775, 777, 779, 781, 783,
785, 787, 789, 791, 793, 795, 796, 797, 798, 799, 801, 803, 805,
807, 808, 809, 810, 811, 812, 813, 815, 816, 817, 819, 821, 823,
825, 827, 829, 832, 834, 836, 839, 840, 841, 842, 843, 844, 845,
846, 848, 849, 850, 851, 852, 853, 854, 856, 857, 859, 861, 863,
865, 867, 869, 870, 871, 872, 873, 874, 876, 878, 880, 882, 885,
886, 887, 888, 889, 890, 891, 892, 893, 894, 896, 897, 898, 900,
901, 902, 904, 906, 908, 910, 912, 913, 914, 915, 916, 918, 920,
922, 924, 926, 929, 931, 932, 933, 935, 937, 939, 941, 943, 944,
945, 947, 949, 951, 952, 953, 955, 957, 958, 959, 960, 961, 962,
963, 964, 966, 968, 969, 970, 971, 973, 974, 975, 977, 978, 979,
980, 981, 983, 985, 987, 988, 989, 990, 991, 992, 993, 995, 997,
998, 999, 1000, 1001, 1003, 1005, 1007, 1010, 1012, 1013, 1014,
1015, 1016, 1018, 1020, 1022, 1023, 1025, 1027, 1028, 1030, 1031,
1033, 1035, 1037, 1039, 1041, 1043, 1045, 1046, 1048, 1050, 1051,
1053, 1055, 1056, 1058, 1059, 1061, 1062, 1064, 1066, 1068, 1069,
1071, 1073, 1075, 1077, 1079, 1080, 1082, 1084, 1086, 1088, 1090,
1092, 1093, 1094, 1096, 1098, 1100, 1102, 1104, 1106, 1108, 1110,
1112, 1113, 1114, 1116, 1117, 1118, 1119, 1121, 1123, 1125, 1129,
1130, 1131, 1132, 1133, 1134, 1135, 1136, 1137, 1138, 1140, 1142,
1144, 1146, 1148, 1150, 1151, 1152, 1153, 1154, 1156, 1157, 1158,
1159, 1160, 1161, 1162, 1164, 1166, 1168, 1170, 1172, 1174, 1175,
1177, 1179, 1181, 1182, 1184, 1186, 1188, 1189, 1191, 1193, 1195,
1197, 1199, 1201, 1203, 1205, 1207, 1209, 1210, 1212, 1214, 1215,
1216, 1218, 1219, 1221, 1222, 1223, 1225, 1226, 1228, 1230, 1231,
1232, 1234, 1236, 1238, 1240, 1242, 1244, 1246, 1248, 1250, 1252,
1254, 1256, 1258, 1262, 1264, 1265, 1267, 1269, 1271, 1273, 1274,
1276, 1277, 1279, 1281, 1283, 1285, 1286, 1287, 1288, 1290, 1292,
1293, 1294, 1296, 1298, 1300, 1302, 1304, 1305, 1311, 1313, 1315,
1317, 1319, 1321, 1323, 1324, 1325, 1326, 1327, 1329, 1331, 1333,
1334, 1336, 1338, 1340, 1342, 1343, 1345, 1347, 1349, 1350, 1351,
1353, 1354, 1356, 1358, 1360, or 1361. The heat and/or
drought-tolerance polypeptide encoded by a recombinant nucleic acid
can be a native heat and/or drought-tolerance polypeptide, or can
be heterologous to the cell. Examples of suitable regulatory
regions are described in the section entitled "Regulatory
Regions."
[0163] Vectors containing recombinant nucleic acid constructs such
as those described herein also are provided. Suitable vector
backbones include, for example, those routinely used in the art
such as plasmids, viruses, artificial chromosomes, BACs, YACs, or
PACs. Suitable expression vectors include, without limitation,
plasmids and viral vectors derived from, for example,
bacteriophage, baculoviruses, and retroviruses. Numerous vectors
and expression systems are commercially available from such
corporations as Novagen (Madison, Wis.), Clontech (Palo Alto,
Calif.), Stratagene (La Jolla, Calif.), and Invitrogen/Life
Technologies (Carlsbad, Calif.).
[0164] The vectors provided herein also can include, for example,
origins of replication, scaffold attachment regions (SARs), and/or
markers. A marker gene can confer a selectable phenotype on a plant
cell. For example, a marker can confer biocide resistance, such as
resistance to an antibiotic (e.g., kanamycin, G418, bleomycin, or
hygromycin), or an herbicide (e.g., glyphosate, chlorsulfuron or
phosphinothricin). In addition, an expression vector can include a
tag sequence designed to facilitate manipulation or detection
(e.g., purification or localization) of the expressed polypeptide.
Tag sequences, such as luciferase, .beta.-glucuronidase (GUS),
green fluorescent protein (GFP), glutathione S-transferase (GST),
polyhistidine, c-myc, hemagglutinin, or Flag.TM. tag (Kodak, New
Haven, Conn.) sequences typically are expressed as a fusion with
the encoded polypeptide. Such tags can be inserted anywhere within
the polypeptide, including at either the carboxyl or amino
terminus.
[0165] D. Regulatory Regions
[0166] The choice of regulatory regions to be included in a
recombinant construct depends upon several factors, including, but
not limited to, efficiency, selectability, inducibility, desired
expression level, and cell- or tissue-preferential expression. It
is a routine matter for one of skill in the art to modulate the
expression of a coding sequence by appropriately selecting and
positioning regulatory regions relative to the coding sequence.
Transcription of a nucleic acid can be modulated in a similar
manner.
[0167] Some suitable regulatory regions initiate transcription
only, or predominantly, in certain cell types. Methods for
identifying and characterizing regulatory regions in plant genomic
DNA are known, including, for example, those described in the
following references: Jordano et al., Plant Cell, 1:855-866 (1989);
Bustos et al., Plant Cell, 1:839-854 (1989); Green et al., EMBO J.,
7:4035-4044 (1988); Meier et al., Plant Cell, 3:309-316 (1991); and
Zhang et al., Plant Physiology, 110:1069-1079 (1996).
[0168] Examples of various classes of regulatory regions are
described below. Some of the regulatory regions indicated below as
well as additional regulatory regions are described in more detail
in U.S. Patent Application Ser. Nos. 60/505,689; 60/518,075;
60/544,771; 60/558,869; 60/583,691; 60/619,181; 60/637,140;
60/757,544; 60/776,307; 10/957,569; 11/058,689; 11/172,703;
11/208,308; 11/274,890; 60/583,609; 60/612,891; 11/097,589;
11/233,726; 11/408,791; 11/414,142; 10/950,321; 11/360,017;
PCT/US05/011105; PCT/US05/23639; PCT/US05/034308; PCT/US05/034343;
and PCT/US06/038236; PCT/US06/040572; and PCT/US07/62762.
[0169] For example, the sequences of regulatory regions p326,
YP0144, YP0190, p13879, YP0050, p32449, 21876, YP0158, YP0214,
YP0380, PT0848, PT0633, YP0128, YP0275, PT0660, PT0683, PT0758,
PT0613, PT0672, PT0688, PT0837, YP0092, PT0676, PT0708, YP0396,
YP0007, YP0111, YP0103, YP0028, YP0121, YP0008, YP0039, YP0115,
YP0119, YP0120, YP0374, YP0101, YP0102, YP0110, YP0117, YP0137,
YP0285, YP0212, YP0097, YP0107, YP0088, YP0143, YP0156, PT0650,
PT0695, PT0723, PT0838, PT0879, PT0740, PT0535, PT0668, PT0886,
PT0585, YP0381, YP0337, PT0710, YP0356, YP0385, YP0384, YP0286,
YP0377, PD1367, PT0863, PT0829, PT0665, PT0678, YP0086, YP0188,
YP0263, PT0743 and YP0096 are set forth in the sequence listing of
PCT/US06/040572; the sequence of regulatory region PT0625 is set
forth in the sequence listing of PCT/US05/034343; the sequences of
regulatory regions PT0623, YP0388, YP0087, YP0093, YP0108, YP0022
and YP0080 are set forth in the sequence listing of U.S. patent
application Ser. No. 11/172,703; the sequence of regulatory region
PR0924 is set forth in the sequence listing of PCT/US07/62762; and
the sequences of regulatory regions p530c10, pOsFIE2-2, pOsMEA,
pOsYp102, and pOsYp285 are set forth in the sequence listing of
PCT/US06/038236.
[0170] It will be appreciated that a regulatory region may meet
criteria for one classification based on its activity in one plant
species, and yet meet criteria for a different classification based
on its activity in another plant species.
[0171] i. Broadly Expressing Promoters
[0172] A promoter can be said to be "broadly expressing" when it
promotes transcription in many, but not necessarily all, plant
tissues. For example, a broadly expressing promoter can promote
transcription of an operably linked sequence in one or more of the
shoot, shoot tip (apex), and leaves, but weakly or not at all in
tissues such as roots or stems. As another example, a broadly
expressing promoter can promote transcription of an operably linked
sequence in one or more of the stem, shoot, shoot tip (apex), and
leaves, but can promote transcription weakly or not at all in
tissues such as reproductive tissues of flowers and developing
seeds. Non-limiting examples of broadly expressing promoters that
can be included in the nucleic acid constructs provided herein
include the p326, YP0144, YP0190, p13879, YP0050, p32449, 21876,
YP0158, YP0214, YP0380, PT0848, and PT0633 promoters. Additional
examples include the cauliflower mosaic virus (CaMV) 35S promoter,
the mannopine synthase (MAS) promoter, the 1' or 2' promoters
derived from T-DNA of Agrobacterium tumefaciens, the figwort mosaic
virus 34S promoter, actin promoters such as the rice actin
promoter, and ubiquitin promoters such as the maize ubiquitin-1
promoter. In some cases, the CaMV 35S promoter is excluded from the
category of broadly expressing promoters.
[0173] ii. Root Promoters
[0174] Root-active promoters confer transcription in root tissue,
e.g., root endodermis, root epidermis, or root vascular tissues. In
some embodiments, root-active promoters are root-preferential
promoters, i.e., confer transcription only or predominantly in root
tissue. Root-preferential promoters include the YP0128, YP0275,
PT0625, PT0660, PT0683, and PT0758 promoters. Other
root-preferential promoters include the PT0613, PT0672, PT0688, and
PT0837 promoters, which drive transcription primarily in root
tissue and to a lesser extent in ovules and/or seeds. Other
examples of root-preferential promoters include the root-specific
subdomains of the CaMV 35S promoter (Lam et al., Proc. Natl. Acad.
Sci. USA, 86:7890-7894 (1989)), root cell specific promoters
reported by Conkling et al., Plant Physiol., 93:1203-1211 (1990),
and the tobacco RD2 promoter.
[0175] iii. Maturing Endosperm Promoters
[0176] In some embodiments, promoters that drive transcription in
maturing endosperm can be useful. Transcription from a maturing
endosperm promoter typically begins after fertilization and occurs
primarily in endosperm tissue during seed development and is
typically highest during the cellularization phase. Most suitable
are promoters that are active predominantly in maturing endosperm,
although promoters that are also active in other tissues can
sometimes be used. Non-limiting examples of maturing endosperm
promoters that can be included in the nucleic acid constructs
provided herein include the napin promoter, the Arcelin-5 promoter,
the phaseolin promoter (Bustos et al., Plant Cell, 1(9):839-853
(1989)), the soybean trypsin inhibitor promoter (Riggs et al.,
Plant Cell, 1(6):609-621 (1989)), the ACP promoter (Baerson et al.,
Plant Mol. Biol., 22(2):255-267 (1993)), the stearoyl-ACP
desaturase promoter (Slocombe et al., Plant Physiol.,
104(4):167-176 (1994)), the soybean .alpha.' subunit of
.beta.-conglycinin promoter (Chen et al., Proc. Natl. Acad. Sci.
USA, 83:8560-8564 (1986)), the oleosin promoter (Hong et al., Plant
Mol. Biol., 34(3):549-555 (1997)), and zein promoters, such as the
15 kD zein promoter, the 16 kD zein promoter, 19 kD zein promoter,
22 kD zein promoter and 27 kD zein promoter. Also suitable are the
Osgt-1 promoter from the rice glutelin-1 gene (Zheng et al., Mol.
Cell Biol., 13:5829-5842 (1993)), the beta-amylase promoter, and
the barley hordein promoter. Other maturing endosperm promoters
include the YP0092, PT0676, and PT0708 promoters.
[0177] iv. Ovary Tissue Promoters
[0178] Promoters that are active in ovary tissues such as the ovule
wall and mesocarp can also be useful, e.g., a polygalacturonidase
promoter, the banana TRX promoter, the melon actin promoter,
YP0396, and PT0623. Examples of promoters that are active primarily
in ovules include YP0007, YP0111, YP0092, YP0103, YP0028, YP0121,
YP0008, YP0039, YP0115, YP0119, YP0120, and YP0374.
[0179] v. Embryo Sac/Early Endosperm Promoters
[0180] To achieve expression in embryo sac/early endosperm,
regulatory regions can be used that are active in polar nuclei
and/or the central cell, or in precursors to polar nuclei, but not
in egg cells or precursors to egg cells. Most suitable are
promoters that drive expression only or predominantly in polar
nuclei or precursors thereto and/or the central cell. A pattern of
transcription that extends from polar nuclei into early endosperm
development can also be found with embryo sac/early
endosperm-preferential promoters, although transcription typically
decreases significantly in later endosperm development during and
after the cellularization phase. Expression in the zygote or
developing embryo typically is not present with embryo sac/early
endosperm promoters.
[0181] Promoters that may be suitable include those derived from
the following genes: Arabidopsis viviparous-1 (see, GenBank No.
U93215); Arabidopsis atmycl (see, Urao (1996) Plant Mol. Biol.,
32:571-57; Conceicao (1994) Plant, 5:493-505); Arabidopsis FIE
(GenBank No. AF129516); Arabidopsis MEA; Arabidopsis FIS2 (GenBank
No. AF096096); and FIE 1.1 (U.S. Pat. No. 6,906,244). Other
promoters that may be suitable include those derived from the
following genes: maize MAC1 (see, Sheridan (1996) Genetics,
142:1009-1020); maize Cat3 (see, GenBank No. L05934; Abler (1993)
Plant Mol. Biol., 22:10131-1038). Other promoters include the
following Arabidopsis promoters: YP0039, YP0101, YP0102, YP0110,
YP0117, YP0119, YP0137, DME, YP0285, and YP0212. Other promoters
that may be useful include the following rice promoters: p530c10,
pOsFIE2-2, pOsMEA, pOsYp102, and pOsYp285.
[0182] vi. Embryo Promoters
[0183] Regulatory regions that preferentially drive transcription
in zygotic cells following fertilization can provide
embryo-preferential expression. Most suitable are promoters that
preferentially drive transcription in early stage embryos prior to
the heart stage, but expression in late stage and maturing embryos
is also suitable. Embryo-preferential promoters include the barley
lipid transfer protein (Ltpl) promoter (Plant Cell Rep (2001)
20:647-654), YP0097, YP0107, YP0088, YP0143, YP0156, PT0650,
PT0695, PT0723, PT0838, PT0879, and PT0740.
[0184] vii. Photosynthetic Tissue Promoters
[0185] Promoters active in photosynthetic tissue confer
transcription in green tissues such as leaves and stems. Most
suitable are promoters that drive expression only or predominantly
in such tissues. Examples of such promoters include the
ribulose-1,5-bisphosphate carboxylase (RbcS) promoters such as the
RbcS promoter from eastern larch (Larix laricina), the pine cab6
promoter (Yamamoto et al., Plant Cell Physiol., 35:773-778 (1994)),
the Cab-1 promoter from wheat (Fejes et al., Plant Mol. Biol.,
15:921-932 (1990)), the CAB-1 promoter from spinach (Lubberstedt et
al., Plant Physiol., 104:997-1006 (1994)), the cablR promoter from
rice (Luan et al., Plant Cell, 4:971-981 (1992)), the pyruvate
orthophosphate dikinase (PPDK) promoter from corn (Matsuoka et al.,
Proc. Natl. Acad. Sci. USA, 90:9586-9590 (1993)), the tobacco
Lhcb1*2 promoter (Cerdan et al., Plant Mol. Biol., 33:245-255
(1997)), the Arabidopsis thaliana SUC2 sucrose-H+ symporter
promoter (Truernit et al., Planta, 196:564-570 (1995)), and
thylakoid membrane protein promoters from spinach (psaD, psaF,
psaE, PC, FNR, atpC, atpD, cab, rbcS). Other photosynthetic tissue
promoters include PT0535, PT0668, PT0886, YP0144, YP0380 and
PT0585.
[0186] viii. Vascular Tissue Promoters
[0187] Examples of promoters that have high or preferential
activity in vascular bundles include YP0087, YP0093, YP0108,
YP0022, and YP0080. Other vascular tissue-preferential promoters
include the glycine-rich cell wall protein GRP 1.8 promoter (Keller
and Baumgartner, Plant Cell, 3(10):1051-1061 (1991)), the Commelina
yellow mottle virus (CoYMV) promoter (Medberry et al., Plant Cell,
4(2):185-192 (1992)), and the rice tungro bacilliform virus (RTBV)
promoter (Dai et al., Proc. Natl. Acad. Sci. USA, 101(2):687-692
(2004)).
[0188] ix. Inducible Promoters
[0189] Inducible promoters confer transcription in response to
external stimuli such as chemical agents or environmental stimuli.
For example, inducible promoters can confer transcription in
response to hormones such as giberellic acid or ethylene, or in
response to light or drought. Examples of drought-inducible
promoters include YP0380, PT0848, YP0381, YP0337, PT0633, YP0374,
PT0710, YP0356, YP0385, YP0396, YP0388, YP0384, PT0688, YP0286,
YP0377, PD1367, and PD0901. Examples of nitrogen-inducible
promoters include PT0863, PT0829, PT0665, and PT0886. Examples of
shade-inducible promoters include PR0924 and PT0678. An example of
a promoter induced by salt is rd29A (Kasuga et al., Nature Biotech
17: 287-291 (1999)).
[0190] x. Basal Promoters
[0191] A basal promoter is the minimal sequence necessary for
assembly of a transcription complex required for transcription
initiation. Basal promoters frequently include a "TATA box" element
that may be located between about 15 and about 35 nucleotides
upstream from the site of transcription initiation. Basal promoters
also may include a "CCAAT box" element (typically the sequence
CCAAT) and/or a GGGCG sequence, which can be located between about
40 and about 200 nucleotides, typically about 60 to about 120
nucleotides, upstream from the transcription start site.
[0192] xi. Stem Promoters
[0193] A stem promoter may be specific to one or more stem tissues
or specific to stem and other plant parts. Stem promoters may have
high or preferential activity in, for example, epidermis and
cortex, vascular cambium, procambium, or xylem. Examples of stem
promoters include YP0018 which is disclosed in US20060015970 and
CryIA(b) and CryIA(c) (Braga et al., Journal of New Seeds 5:209-221
(2003)).
[0194] xii. Other Promoters
[0195] Other classes of promoters include, but are not limited to,
shoot-preferential, callus-preferential, trichome
cell-preferential, guard cell-preferential such as PT0678,
tuber-preferential, parenchyma cell-preferential, and
senescence-preferential promoters. Promoters designated YP0086,
YP0188, YP0263, PT0758, PT0743, PT0829, YP0119, and YP0096, as
described in the above-referenced patent applications, may also be
useful.
[0196] xiii. Other Regulatory Regions
[0197] A 5' untranslated region (UTR) can be included in nucleic
acid constructs described herein. A 5' UTR is transcribed, but is
not translated, and lies between the start site of the transcript
and the translation initiation codon and may include the +1
nucleotide. A 3' UTR can be positioned between the translation
termination codon and the end of the transcript. UTRs can have
particular functions such as increasing mRNA stability or
attenuating translation. Examples of 3' UTRs include, but are not
limited to, polyadenylation signals and transcription termination
sequences, e.g., a nopaline synthase termination sequence.
[0198] It will be understood that more than one regulatory region
may be present in a recombinant polynucleotide, e.g., introns,
enhancers, upstream activation regions, transcription terminators,
and inducible elements. Thus, for example, more than one regulatory
region can be operably linked to the sequence of a polynucleotide
encoding a heat and/or drought-tolerance polypeptide.
[0199] Regulatory regions, such as promoters for endogenous genes,
can be obtained by chemical synthesis or by subcloning from a
genomic DNA that includes such a regulatory region. A nucleic acid
comprising such a regulatory region can also include flanking
sequences that contain restriction enzyme sites that facilitate
subsequent manipulation.
IV. TRANSGENIC PLANTS AND PLANT CELLS
A. Transformation
[0200] The invention also features transgenic plant cells and
plants comprising at least one recombinant nucleic acid construct
described herein. A plant or plant cell can be transformed by
having a construct integrated into its genome, i.e., can be stably
transformed. Stably transformed cells typically retain the
introduced nucleic acid with each cell division. A plant or plant
cell can also be transiently transformed such that the construct is
not integrated into its genome. Transiently transformed cells
typically lose all or some portion of the introduced nucleic acid
construct with each cell division such that the introduced nucleic
acid cannot be detected in daughter cells after a sufficient number
of cell divisions. Both transiently transformed and stably
transformed transgenic plants and plant cells can be useful in the
methods described herein.
[0201] Transgenic plant cells used in methods described herein can
constitute part or all of a whole plant. Such plants can be grown
in a manner suitable for the species under consideration, either in
a growth chamber, a greenhouse, or in a field. Transgenic plants
can be bred as desired for a particular purpose, e.g., to introduce
a recombinant nucleic acid into other lines, to transfer a
recombinant nucleic acid to other species, or for further selection
of other desirable traits. Alternatively, transgenic plants can be
propagated vegetatively for those species amenable to such
techniques. As used herein, a transgenic plant also refers to
progeny of an initial transgenic plant provided the progeny
inherits the transgene. Seeds produced by a transgenic plant can be
grown and then selfed (or outcrossed and selfed) to obtain seeds
homozygous for the nucleic acid construct.
[0202] Transgenic plants can be grown in suspension culture, or
tissue or organ culture. For the purposes of this invention, solid
and/or liquid tissue culture techniques can be used. When using
solid medium, transgenic plant cells can be placed directly onto
the medium or can be placed onto a filter that is then placed in
contact with the medium. When using liquid medium, transgenic plant
cells can be placed onto a flotation device, e.g., a porous
membrane that contacts the liquid medium. A solid medium can be,
for example, Murashige and Skoog (MS) medium containing agar and a
suitable concentration of an auxin, e.g., 2,4-dichlorophenoxyacetic
acid (2,4-D), and a suitable concentration of a cytokinin, e.g.,
kinetin.
[0203] When transiently transformed plant cells are used, a
reporter sequence encoding a reporter polypeptide having a reporter
activity can be included in the transformation procedure and an
assay for reporter activity or expression can be performed at a
suitable time after transformation. A suitable time for conducting
the assay typically is about 1-21 days after transformation, e.g.,
about 1-14 days, about 1-7 days, or about 1-3 days. The use of
transient assays is particularly convenient for rapid analysis in
different species, or to confirm expression of a heterologous heat
and/or drought-tolerance polypeptide whose expression has not
previously been confirmed in particular recipient cells.
[0204] Techniques for introducing nucleic acids into
monocotyledonous and dicotyledonous plants are known in the art,
and include, without limitation, Agrobacterium-mediated
transformation, viral vector-mediated transformation,
electroporation and particle gun transformation, e.g., U.S. Pat.
Nos. 5,538,880; 5,204,253; 6,329,571 and 6,013,863. If a cell or
cultured tissue is used as the recipient tissue for transformation,
plants can be regenerated from transformed cultures if desired, by
techniques known to those skilled in the art.
B. Screening/Selection
[0205] A population of transgenic plants can be screened and/or
selected for those members of the population that have a trait or
phenotype conferred by expression of the transgene. For example, a
population of progeny of a single transformation event can be
screened for those plants having a desired level of expression of a
heat and/or drought-tolerance polypeptide or nucleic acid. Physical
and biochemical methods can be used to identify expression levels.
These include Southern analysis or PCR amplification for detection
of a polynucleotide; Northern blots, S1 RNase protection,
primer-extension, or RT-PCR amplification for detecting RNA
transcripts; enzymatic assays for detecting enzyme or ribozyme
activity of polypeptides and polynucleotides; and protein gel
electrophoresis, Western blots, immunoprecipitation, and
enzyme-linked immunoassays to detect polypeptides. Other techniques
such as in situ hybridization, enzyme staining, and immunostaining
also can be used to detect the presence or expression of
polypeptides and/or polynucleotides. Methods for performing all of
the referenced techniques are known. As an alternative, a
population of plants comprising independent transformation events
can be screened for those plants having a desired trait, such as
increased drought tolerance or increased heat tolerance. Selection
and/or screening can be carried out over one or more generations,
and/or in more than one geographic location. In some cases,
transgenic plants can be grown and selected under conditions which
induce a desired phenotype or are otherwise necessary to produce a
desired phenotype in a transgenic plant. In addition, selection
and/or screening can be applied during a particular developmental
stage in which the phenotype is expected to be exhibited by the
plant. Selection and/or screening can be carried out to choose
those transgenic plants having a statistically significant
difference in heat and/or drought tolerance relative to a control
plant that lacks the transgene. Selected or screened transgenic
plants have an altered phenotype as compared to a corresponding
control plant, as described in the "Transgenic Plant Phenotypes"
section herein.
C. Plant Species
[0206] The polynucleotides and vectors described herein can be used
to transform a number of monocotyledonous and dicotyledonous plants
and plant cell systems, including species from one of the following
families: Acanthaceae, Alliaceae, Alstroemeriaceae, Amaryllidaceae,
Apocynaceae, Arecaceae, Asteraceae, Berberidaceae, Bixaceae,
Brassicaceae, Bromeliaceae, Cannabaceae, Caryophyllaceae,
Cephalotaxaceae, Chenopodiaceae, Colchicaceae, Cucurbitaceae,
Dioscoreaceae, Ephedraceae, Erythroxylaceae, Euphorbiaceae,
Fabaceae, Lamiaceae, Linaceae, Lycopodiaceae, Malvaceae,
Melanthiaceae, Musaceae, Myrtaceae, Nyssaceae, Papaveraceae,
Pinaceae, Plantaginaceae, Poaceae, Rosaceae, Rubiaceae, Salicaceae,
Sapindaceae, Solanaceae, Taxaceae, Theaceae, or Vitaceae.
[0207] Suitable species may include members of the genus
Abelmoschus, Abies, Acer, Agrostis, Allium, Alstroemeria, Ananas,
Andrographis, Andropogon, Artemisia, Arundo, Atropa, Berberis,
Beta, Bixa, Brassica, Calendula, Camellia, Camptotheca, Cannabis,
Capsicum, Carthamus, Catharanthus, Cephalotaxus, Chrysanthemum,
Cinchona, Citrullus, Coffea, Colchicum, Coleus, Cucumis, Cucurbita,
Cynodon, Datura, Dianthus, Digitalis, Dioscorea, Elaeis, Ephedra,
Erianthus, Erythroxylum, Eucalyptus, Festuca, Fragaria, Galanthus,
Glycine, Gossypium, Helianthus, Hevea, Hordeum, Hyoscyamus,
Jatropha, Lactuca, Linum, Lolium, Lupinus, Lycopersicon,
Lycopodium, Manihot, Medicago, Mentha, Miscanthus, Musa, Nicotiana,
Oryza, Panicum, Papaver, Parthenium, Pennisetum, Petunia, Phalaris,
Phleum, Pinus, Poa, Poinsettia, Populus, Rauwolfia, Ricinus, Rosa,
Saccharum, Salix, Sanguinaria, Scopolia, Secale, Solanum, Sorghum,
Spartina, Spinacea, Tanacetum, Taxus, Theobroma, Triticosecale,
Triticum, Uniola, Veratrum, Vinca, Vitis, and Zea.
[0208] Suitable species include Panicum spp. or hybrids thereof,
Sorghum spp. or hybrids thereof, sudangrass, Miscanthus spp. or
hybrids thereof, Saccharum spp. or hybrids thereof, Erianthus spp.,
Populus spp., Andropogon gerardii (big bluestem), Pennisetum
purpureum (elephant grass) or hybrids thereof (e.g., Pennisetum
purpureum.times.Pennisetum typhoidum), Phalaris arundinacea (reed
canarygrass), Cynodon dactylon (bermudagrass), Festuca arundinacea
(tall fescue), Spartina pectinata (prairie cord-grass), Medicago
sativa (alfalfa), Arundo donax (giant reed) or hybrids thereof,
Secale cereale (rye), Salix spp. (willow), Eucalyptus spp.
(eucalyptus), Triticosecale (Triticum--wheat X rye), Tripsicum
dactyloides (Eastern gammagrass), Leymus cinereus (basin wildrye),
Leymus condensatus (giant wildrye), and bamboo.
[0209] In some embodiments, a suitable species can be a wild,
weedy, or cultivated sorghum species such as, but not limited to,
Sorghum almum, Sorghum amplum, Sorghum angustum, Sorghum
arundinaceum, Sorghum bicolor (such as bicolor, guinea, caudatum,
kafir, and durra), Sorghum brachypodum, Sorghum bulbosum, Sorghum
burmahicum, Sorghum controversum, Sorghum drummondii, Sorghum
ecarinatum, Sorghum exstans, Sorghum grande, Sorghum halepense,
Sorghum interjectum, Sorghum intrans, Sorghum laxiflorum, Sorghum
leiocladum, Sorghum macrospermum, Sorghum matarankense, Sorghum
miliaceum, Sorghum nigrum, Sorghum nitidum, Sorghum plumosum,
Sorghum propinquum, Sorghum purpureosericeum, Sorghum stipoideum,
Sorghum sudanensese, Sorghum timorense, Sorghum trichocladum,
Sorghum versicolor, Sorghum virgatum, Sorghum vulgare, or hybrids
such as Sorghum.times.almum, Sorghum.times.sudangrass or
Sorghum.times.drummondii.
[0210] Suitable species also include Helianthus annuus (sunflower),
Carthamus tinctorius (safflower), Jatropha curcas (jatropha),
Ricinus communis (castor), Elaeis guineensis (palm), Linum
usitatissimum (flax), and Brassica juncea.
[0211] Suitable species also include Beta vulgaris (sugarbeet), and
Manihot esculenta (cassava).
[0212] Suitable species also include Lycopersicon esculentum
(tomato), Lactuca sativa (lettuce), Musa paradisiaca (banana),
Solanum tuberosum (potato), Brassica oleracea (broccoli,
cauliflower, brusselsprouts), Camellia sinensis (tea), Fragaria
ananassa (strawberry), Theobroma cacao (cocoa), Coffea arabica
(coffee), Vitis vinifera (grape), Ananas comosus (pineapple),
Capsicum annum (hot & sweet pepper), Allium cepa (onion),
Cucumis melo (melon), Cucumis sativus (cucumber), Cucurbita maxima
(squash), Cucurbita moschata (squash), Spinacea oleracea (spinach),
Citrullus lanatus (watermelon), Abelmoschus esculentus (okra), and
Solanum melongena (eggplant).
[0213] Suitable species also include Papaver somniferum (opium
poppy), Papaver orientale, Taxus baccata, Taxus brevifolia,
Artemisia annua, Cannabis sativa, Camptotheca acuminate,
Catharanthus roseus, Vinca rosea, Cinchona officinalis, Colchicum
autumnale, Veratrum californica, Digitalis lanata, Digitalis
purpurea, Dioscorea spp., Andrographis paniculata, Atropa
belladonna, Datura stomonium, Berberis spp., Cephalotaxus spp.,
Ephedra sinica, Ephedra spp., Erythroxylum coca, Galanthus
wornorii, Scopolia spp., Lycopodium serratum (=Huperzia serrata),
Lycopodium spp., Rauwolfia serpentina, Rauwolfia spp., Sanguinaria
canadensis, Hyoscyamus spp., Calendula officinalis, Chrysanthemum
parthenium, Coleus forskohlii, and Tanacetum parthenium.
[0214] Suitable species also include Parthenium argentatum
(guayule), Hevea spp. (rubber), Mentha spicata (mint), Mentha
piperita (mint), Bixa orellana, and Alstroemeria spp.
[0215] Suitable species also include Rosa spp. (rose), Dianthus
caryophyllus (carnation), Petunia spp. (petunia) and Poinsettia
pulcherrima (poinsettia).
[0216] Suitable species also include Nicotiana tabacum (tobacco),
Lupinus albus (lupin), Uniola paniculata (oats), bentgrass
(Agrostis spp.), Populus tremuloides (aspen), Pinus spp. (pine),
Abies spp. (fir), Acer spp. (maple, Hordeum vulgare (barley), Poa
pratensis (bluegrass), Lolium spp. (ryegrass) and Phleum pratense
(timothy).
[0217] Thus, the methods and compositions can be used over a broad
range of plant species, including species from the dicot genera
Brassica, Carthamus, Glycine, Gossypium, Helianthus, Jatropha,
Parthenium, Populus, and Ricinus; and the monocot genera Elaeis,
Festuca, Hordeum, Lolium, Oryza, Panicum, Pennisetum, Phleum, Poa,
Saccharum, Secale, Sorghum, Triticosecale, Triticum, and Zea. In
some embodiments, a plant is a member of the species Panicum
virgatum (switchgrass), Sorghum bicolor (sorghum, sudangrass),
Miscanthus giganteus (miscanthus), Saccharum sp. (energycane),
Populus balsamifera (poplar), Zea mays (corn), Glycine max
(soybean), Brassica napus (canola), Triticum aestivum (wheat),
Gossypium hirsutum (cotton), Oryza sativa (rice), Helianthus annuus
(sunflower), Medicago sativa (alfalfa), Beta vulgaris (sugarbeet),
or Pennisetum glaucum (pearl millet).
[0218] In certain embodiments, the polynucleotides and vectors
described herein can be used to transform a number of
monocotyledonous and dicotyledonous plants and plant cell systems,
wherein such plants are hybrids of different species or varieties
of a specific species (e.g., Saccharum sp. X Miscanthus sp.,
Saccharum sp. X Sorghum sp., Panicum virgatum.times.Panicum amarum,
Panicum virgatum.times.Panicum amarulum, and Pennisetum
purpureum.times.Pennisetum typhoidum).
D. Transgenic Plant Phenotypes
[0219] In some embodiments, a plant in which expression of a heat
and/or drought-tolerance polypeptide is modulated can have
increased heat tolerance or drought tolerance. In some cases, a
plant in which expression of a heat and/or drought-tolerance
polypeptide is modulated can exhibit both heat tolerance and
drought tolerance. The phenotype of a transgenic plant in which
expression of a heat and/or drought-tolerance polypeptide is
modulated and a corresponding control plant that either lacks the
transgene or does not express the transgene can be evaluated under
particular environmental conditions that are useful for simulating
drought conditions. The phenotype of a transgenic plant in which
expression of a heat and/or drought-tolerance polypeptide is
modulated and a corresponding control plant can also be evaluated
under heat shock conditions.
[0220] For example, a heat and/or drought-tolerance polypeptide
described herein can be expressed in a transgenic plant, resulting
in increased growth as compared to a plant that does not express
the polypeptide when grown under drought conditions or heat shock
conditions, or following such conditions. Growth can be assessed
with physiological parameters such as, for example, plant height,
number of new shoots, number of new leaves, leaf length, seedling
area, or seed number.
[0221] In some cases, a transgenic plant expressing a heat and/or
drought tolerance polypeptide described herein can exhibit a height
that is from about 5% to about 100% greater (e.g., about 5% to
about 12%; about 5% to about 40%; about 5% to about 80%; about 7%
to about 20%; about 10% to about 15%; about 10% to about 50%; about
10% to about 90%; about 20% to about 25%; about 20% to about 45%;
about 20% to about 75%; about 25% to about 60%; about 25% to about
100%; about 30% to about 50%; about 30% to about 70%; about 40% to
about 50%; about 45% to about 60%; about 50% to about 80%; about
55% to about 75%; about 60% to about 80%; about 60% to about 95%;
about 75% to about 100%; about 80% to about 100%; about 90% to
about 95%; or about 95% to about 100% greater) than a plant not
expressing the polypeptide when grown under drought conditions or
heat shock conditions, or following such conditions.
[0222] In some instances, a transgenic plant expressing a heat
and/or drought-tolerance polypeptide can exhibit greater leaf area
or greater leaf length than a corresponding control plant that does
not express the polypeptide when grown under heat shock conditions
or drought conditions, or following such conditions. For example, a
transgenic plant expressing a heat and/or drought-tolerance
polypeptide can have a leaf area that is 5% to about 100% greater
(e.g., about 5% to about 7%; about 5% to about 20%; about 8% to
about 80%; about 10% to about 20%; about 10% to about 25%; about
10% to about 50%; about 10% to about 90%; about 15% to about 25%;
about 20% to about 45%; about 20% to about 70%; about 25% to about
40%; about 25% to about 100%; about 30% to about 50%; about 30% to
about 70%; about 40% to about 50%; about 45% to about 60%; about
50% to about 80%; about 55% to about 75%; about 60% to about 80%;
about 60% to about 95%; about 75% to about 100%; about 80% to about
100%; about 90% to about 95%; or about 95% to about 100% greater)
than a corresponding control plant that does not express the
polypeptide when grown under heat shock conditions or drought
conditions, or following such conditions. In another example, a
transgenic plant expressing a heat and/or drought-tolerance
polypeptide can have a leaf length that is from about 5% to about
100% greater (e.g., about 5% to about 12%; about 5% to about 40%;
about 5% to about 80%; about 7% to about 20%; about 10% to about
15%; about 10% to about 50%; about 10% to about 90%; about 20% to
about 25%; about 20% to about 45%; about 20% to about 75%; about
25% to about 60%; about 25% to about 100%; about 30% to about 50%;
about 30% to about 70%; about 40% to about 50%; about 45% to about
60%; about 50% to about 80%; about 55% to about 75%; about 60% to
about 80%; about 60% to about 95%; about 75% to about 100%; about
80% to about 100%; about 90% to about 95%; or about 95% to about
100% greater) than a corresponding control plant that does not
express the polypeptide when grown under heat shock conditions or
drought conditions, or following such conditions.
[0223] In other cases, when a polypeptide described herein is
expressed in a transgenic plant, the transgenic plant can exhibit a
seed number (number of seeds per plant) from about 10% to about 95%
greater (e.g., from about 10% to about 20%; from about 10% to about
50%; from about 10% to about 70%; from about 20% to about 60%; from
about 20% to about 75%; from about 25% to about 85%; from about 30%
to about 70%; from about 35% to about 90%; from about 40% to about
60%; from about 40% to about 85%; from about 50% to about 80%; from
about 50% to about 90%; or from about 70% to about 90% greater)
than a control plant not expressing the polypeptide when grown
under heat shock conditions or drought conditions. In certain
cases, when a polypeptide described herein is expressed in a
transgenic plant, the transgenic plant can exhibit an increase in
seed weight per plant from about 5% to about 100% greater (e.g.,
about 5% to about 12%; about 5% to about 40%; about 5% to about
80%; about 7% to about 20%; about 10% to about 15%; about 10% to
about 50%; about 10% to about 90%; about 20% to about 25%; about
20% to about 45%; about 20% to about 75%; about 25% to about 60%;
about 25% to about 100%; about 30% to about 50%; about 30% to about
70%; about 40% to about 50%; about 45% to about 60%; about 50% to
about 80%; about 55% to about 75%; about 60% to about 80%; about
60% to about 95%; about 75% to about 100%; about 80% to about 100%;
about 90% to about 95%; or about 95% to about 100% greater) than
the seed weight in a plant not expressing the polypeptide when
grown under heat shock conditions or drought conditions.
[0224] Transgenic plants expressing a heat and/or drought-tolerance
polypeptide also can exhibit a lower transpiration rate compared to
control plants of the same genetic background. Transpiration rate
is another physiological parameter that is indicative of how well a
plant can tolerate drought conditions. For example, plants with a
low transpiration rate are expected to lose water more slowly than
plants with higher transpiration rates and therefore would be
expected to better withstand drought conditions (i.e., have better
drought tolerance). When a polypeptide described herein is
expressed in a transgenic plant, the transgenic plant can exhibit a
transpiration rate that is reduced by about 0.25% to 100% (e.g.,
0.27%, 0.3%, 0.43%, 0.55%, 0.7%, 0.99%, 1%, 2%, 4%, 6%, 8%, 10%,
12%, 15%, 18%, 22%, 28%, 35%, 37%, 42%, 45%, 47%, 50%, 55%, 64%,
68%, 71%, 75%, 77%, 80%, 83%, 86%, 89%, 90%, 92%, 95%, 98%, or 99%)
as compared to the transpiration rate in a corresponding control
plant when grown under drought conditions.
[0225] In some cases, a transgenic plant expressing a heat and/or
drought-tolerance polypeptide described herein can exhibit a
decreased change in photosynthetic activity (.DELTA.Fv/Fm) after
exposure to heat shock conditions as compared to a corresponding
control plant that does not express the polypeptide when grown
under the same conditions. In some cases, a transgenic plant
expressing a heat and/or drought-tolerance polypeptide described
herein can exhibit an increased change in photosynthetic activity
(.DELTA.Fv/Fm-D.sub.2) two days post heat-shock treatment as
compared to a corresponding control plant that does not express the
polypeptide when grown under the same conditions. For example, a
transgenic plant expressing a heat and/or drought-tolerance
polypeptide can exhibit a .DELTA.Fv/Fm of from about 0.1 to about
0.8 (e.g., about 0.2 to about 0.28; about 0.2 to about 0.32; about
0.22 to about 0.35; about 0.29 to about 0.4; about 0.3 to about
0.45; about 0.33 to about 0.41; about 0.35 to about 0.5; about 0.4
to about 0.8; about 0.46 to about 0.52; about 0.5 to about 0.65;
about 0.5 to about 0.8; about 0.6 to about 0.7; about 0.6 to about
0.9; about 0.65 to about 0.75; about 0.7 to about 0.9; or about
0.75 to about 0.8) or a .DELTA.Fv/Fm-D.sub.2 range of from about
0.03 to about 0.8 (e.g., about 0.03 to about 0.08; about 0.03 to
about 0.032; about 0.04 to about 0.05; about 0.09 to about 0.4;
about 0.05 to about 0.5; about 0.075 to about 0.1; about 0.08 to
about 0.2; about 0.3 to about 0.45; about 0.33 to about 0.41; about
0.35 to about 0.5; about 0.4 to about 0.8; about 0.46 to about
0.52; about 0.5 to about 0.65; about 0.5 to about 0.8; about 0.6 to
about 0.7; about 0.6 to about 0.9; about 0.65 to about 0.75; about
0.7 to about 0.9; about 0.75 to about 0.85; or about 0.8 to about
0.9). In some embodiments, photosynthetic activity can be reduced
by about 0.25% to about 100% (e.g., about 0.25% to about 0.4%,
about 0.25% to about 1%, about 0.25% to about 5%, about 0.5% to
about 10%, about 1% to about 5%, about 1% to about 10%, about 2% to
about 8%, about 3% to about 20%, about 5% to about 7%; about 5% to
about 20%; about 5% to about 45%, about 8% to about 80%; about 10%
to about 20%; about 10% to about 25%; about 10% to about 50%; about
10% to about 90%; about 15% to about 25%; about 20% to about 45%;
about 20% to about 70%; about 25% to about 40%; about 25% to about
99%; about 30% to about 50%; about 30% to about 70%; about 40% to
about 50%; about 45% to about 60%; about 50% to about 80%; about
55% to about 75%; about 60% to about 80%; about 60% to about 95%;
about 75% to about 99%; about 80% to about 99%; about 90% to about
95%; or about 95% to about 100%) as compared to the photosynthetic
activity in a corresponding control plant following heat shock
conditions.
[0226] Typically, a difference in the heat and/or drought tolerance
in a transgenic plant relative to a control plant is considered
statistically significant at p.ltoreq.0.05 with an appropriate
parametric or non-parametric statistic, e.g., Chi-square test,
Student's t-test, Mann-Whitney test, or F-test. In some
embodiments, a difference in the heat and/or drought tolerance is
statistically significant at p<0.1, p<0.05, or p<0.01.
[0227] The phenotype of a transgenic plant is evaluated relative to
a control plant. A plant is said "not to express" a polypeptide
when the plant exhibits less than 10%, e.g., less than 9%, 8%, 7%,
6%, 5%, 4%, 3%, 2%, 1%, 0.5%, 0.1%, 0.1%, or 0.01%, of the amount
of polypeptide or mRNA encoding the polypeptide exhibited by the
plant of interest. Expression can be evaluated using methods
including, for example, RT-PCR, Northern blots, S1 RNase
protection, primer extensions, Western blots, protein gel
electrophoresis, immunoprecipitation, enzyme-linked immunoassays,
chip assays, and mass spectrometry. It should be noted that if a
polypeptide is expressed under the control of a tissue-preferential
or broadly expressing promoter, expression can be evaluated in the
entire plant or in a selected tissue. Similarly, if a polypeptide
is expressed at a particular time, e.g., at a particular time in
development or upon induction, expression can be evaluated
selectively at a desired time period.
V. PLANT BREEDING
[0228] Genetic polymorphisms are discrete allelic sequence
differences in a population. Typically, an allele that is present
at 1% or greater is considered to be a genetic polymorphism. The
discovery that polypeptides disclosed herein can increase heat
and/or drought tolerance is useful in plant breeding, because
genetic polymorphisms exhibiting a degree of linkage with loci for
such polypeptides are more likely to be correlated with variation
in heat and/or drought tolerance. For example, genetic
polymorphisms linked to the loci for such polypeptides are more
likely to be useful in marker-assisted breeding programs to create
lines having increased heat and/or drought tolerance.
[0229] Thus, one aspect of the invention includes methods of
identifying whether one or more genetic polymorphisms are
associated with heat and/or drought tolerance. Such methods involve
determining whether genetic polymorphisms in a given population
exhibit linkage with the locus for one of the polypeptides depicted
in FIGS. 1-16 and/or a functional homolog thereof. The correlation
is measured between variation in heat and/or drought tolerance in
plants of the population and the presence of the genetic
polymorphism(s) in plants of the population, thereby identifying
whether or not the genetic polymorphism(s) are associated with
variation for the trait. If the presence of a particular allele is
statistically significantly correlated with a modulation in heat
and/or drought tolerance, the allele is associated with variation
for the trait and is useful as a marker for the trait. If, on the
other hand, the presence of a particular allele is not
significantly correlated with the desired modulation, the allele is
not associated with variation for the trait and is not useful as a
marker.
[0230] Such methods are applicable to populations containing the
naturally occurring endogenous polypeptide rather than an exogenous
nucleic acid encoding the polypeptide, i.e., populations that are
not transgenic for the exogenous nucleic acid. It will be
appreciated, however, that populations suitable for use in the
methods may contain a transgene for another, different trait, e.g.,
herbicide resistance.
[0231] Genetic polymorphisms that are useful in such methods
include simple sequence repeats (SSRs, or microsatellites), rapid
amplification of polymorphic DNA (RAPDs), single nucleotide
polymorphisms (SNPs), amplified fragment length polymorphisms
(AFLPs) and restriction fragment length polymorphisms (RFLPs). SSR
polymorphisms can be identified, for example, by making sequence
specific probes and amplifying template DNA from individuals in the
population of interest by PCR. For example, PCR techniques can be
used to enzymatically amplify a genetic marker associated with a
nucleotide sequence conferring a specific trait (e.g., nucleotide
sequences described herein). PCR can be used to amplify specific
sequences from DNA as well as RNA, including sequences from total
genomic DNA or total cellular RNA. When using RNA as a source of
template, reverse transcriptase can be used to synthesize
complementary DNA (cDNA) strands. Various PCR methods are
described, for example, in PCR Primer: A Laboratory Manual,
Dieffenbach and Dveksler, eds., Cold Spring Harbor Laboratory
Press, 1995.
[0232] Generally, sequence information from polynucleotides
flanking the region of interest or beyond is employed to design
oligonucleotide primers that are identical or similar in sequence
to opposite strands of the template to be amplified. Primers are
typically 14 to 40 nucleotides in length, but can range from 10
nucleotides to hundreds of nucleotides in length. Template and
amplified DNA is repeatedly denatured at a high temperature to
separate the double strand, then cooled to allow annealing of
primers and the extension of nucleotide sequences through the
microsatellite, resulting in sufficient DNA for detection of PCR
products. If the probes flank an SSR in the population, PCR
products of different sizes will be produced. See, e.g., U.S. Pat.
No. 5,766,847.
[0233] PCR products can be qualitative or quantitatively analyzed
using several techniques. For example, PCR products can be stained
with a fluorescent molecule (e.g., PicoGreen.RTM. or OliGreen.RTM.)
and detected in solution using spectrophotometry or capillary
electrophoresis. In some cases, PCR products can be separated in a
gel matrix (e.g., agarose or polyacrylamide) by electrophoresis,
and size-fractionated bands comprising PCR products can be
visualized using nucleic acid stains. Suitable stains can fluoresce
under UV light (e.g., Ethidium bromide, GR Safe, SYBR.RTM. Green,
or SYBR.RTM. Gold). The results can be visualized via
transillumination or epi-illumination, and an image of the
fluorescent pattern can be acquired using a camera or scanner, for
example. The image can be processed and analyzed using specialized
software (e.g., ImageJ) to measure and compare the intensity of a
band of interest against a standard loaded on the same gel.
[0234] Alternatively, SSR polymorphisms can be identified by using
PCR product(s) as a probe against Southern blots from different
individuals in the population. See, U. H. Refseth et al., (1997)
Electrophoresis 18: 1519. Briefly, PCR products are separated by
length through gel electrophoresis and transferred to a membrane.
SSR-specific DNA probes, such as oligonucleotides labeled with
radioactive, fluorescent, or chromogenic molecules, are applied to
the membrane and hybridize to bound PCR products with a
complementary nucleotide sequence. The pattern of hybridization can
be visualized by autoradiography or by development of color on the
membrane, for example.
[0235] In some cases, PCR products can be quantified using a
real-time thermocycler detection system. For example, Quantitative
real-time PCR can use a fluorescent dye that forms a
DNA-dye-complex (e.g., SYBR.RTM. Green), or a
fluorophore-containing DNA probe, such as single-stranded
oligonucleotides covalently bound to a fluorescent reporter or
fluorophore (e.g. 6-carboxyfluorescein or tetrachlorofluorescin)
and quencher (e.g., tetramethylrhodamine or
dihydrocyclopyrroloindole tripeptide minor groove binder). The
fluorescent signal allows detection of the amplified product in
real time, thereby indicating the presence of a sequence of
interest, and allowing quantification of the copy number of a
sequence of interest in cellular DNA or expression level of a
sequence of interest from cellular mRNA.
[0236] The identification of RFLPs is discussed, for example, in
Alonso-Blanco et al. (Methods in Molecular Biology, vol. 82,
"Arabidopsis Protocols", pp. 137-146, J. M. Martinez-Zapater and J.
Salinas, eds., c. 1998 by Humana Press, Totowa, N.J.); Burr
("Mapping Genes with Recombinant Inbreds", pp. 249-254, in
Freeling, M. and V. Walbot (Ed.), The Maize Handbook, c. 1994 by
Springer-Verlag New York, Inc.: New York, N.Y., USA; Berlin
Germany; Burr et al. Genetics (1998) 118: 519; and Gardiner, J. et
al., (1993) Genetics 134: 917). For example, to produce a RFLP
library enriched with single- or low-copy expressed sequences,
total DNA can be digested with a methylation-sensitive enzyme
(e.g., PstI). The digested DNA can be separated by size on a
preparative gel. Polynucleotide fragments (500 to 2000 bp) can be
excised, eluted and cloned into a plasmid vector (e.g., pUC18).
Southern blots of plasmid digests can be probed with total sheared
DNA to select clones that hybridize to single- and low-copy
sequences. Additional restriction endonucleases can be tested to
increase the number of polymorphisms detected.
[0237] The identification of AFLPs is discussed, for example, in EP
0 534 858 and U.S. Pat. No. 5,878,215. In general, total cellular
DNA is digested with one or more restriction enzymes. Restriction
halfsite-specific adapters are ligated to all restriction fragments
and the fragments are selectively amplified with two PCR primers
that have corresponding adaptor and restriction site specific
sequences. The PCR products can be visualized after
size-fractionation, as described above.
[0238] In some embodiments, the methods are directed to breeding a
plant line. Such methods use genetic polymorphisms identified as
described above in a marker assisted breeding program to facilitate
the development of lines that have increased heat and/or drought
tolerance. Once a suitable genetic polymorphism is identified as
being associated with variation for the trait, one or more
individual plants are identified that possess the polymorphic
allele correlated with the desired variation. Those plants are then
used in a breeding program to combine the polymorphic allele with a
plurality of other alleles at other loci that are correlated with
the desired variation. Techniques suitable for use in a plant
breeding program are known in the art and include, without
limitation, backcrossing, mass selection, pedigree breeding, bulk
selection, crossing to another population and recurrent selection.
These techniques can be used alone or in combination with one or
more other techniques in a breeding program. Thus, each identified
plants is selfed or crossed a different plant to produce seed which
is then germinated to form progeny plants. At least one such
progeny plant is then selfed or crossed with a different plant to
form a subsequent progeny generation. The breeding program can
repeat the steps of selfing or outcrossing for an additional 0 to 5
generations as appropriate in order to achieve the desired
uniformity and stability in the resulting plant line, which retains
the polymorphic allele. In most breeding programs, analysis for the
particular polymorphic allele will be carried out in each
generation, although analysis can be carried out in alternate
generations if desired.
[0239] In some cases, selection for other useful traits is also
carried out, e.g., selection for fungal resistance or bacterial
resistance. Selection for such other traits can be carried out
before, during or after identification of individual plants that
possess the desired polymorphic allele.
VI. ARTICLES OF MANUFACTURE
[0240] Transgenic plants provided herein have various uses in the
agricultural and energy production industries. For example,
transgenic plants described herein can be used to make animal feed
and food products. Such plants, however, are often particularly
useful as a feedstock for energy production.
[0241] Transgenic plants described herein often produce higher
yields of grain and/or biomass per hectare, relative to control
plants that lack the exogenous nucleic acid. In some embodiments,
such transgenic plants provide equivalent or even increased yields
of grain and/or biomass per hectare relative to control plants when
grown under conditions of reduced inputs such as fertilizer and/or
water. Thus, such transgenic plants can be used to provide yield
stability at a lower input cost and/or under environmentally
stressful conditions such as drought. In some embodiments, plants
described herein have a composition that permits more efficient
processing into free sugars, and subsequently ethanol, for energy
production. In some embodiments, such plants provide higher yields
of ethanol, other biofuel molecules, and/or sugar-derived
co-products per kilogram of plant material, relative to control
plants. By providing higher yields higher yields under
environmentally stressful conditions such as drought conditions
and/or heat shock conditions, the transgenic plants described
herein improve profitability for farmers and processors as well as
decrease costs to consumers.
[0242] Seeds from transgenic plants described herein can be
conditioned and bagged in packaging material by means known in the
art to form an article of manufacture. Packaging material such as
paper and cloth are well known in the art. A package of seed can
have a label, e.g., a tag or label secured to the packaging
material, a label printed on the packaging material, or a label
inserted within the package, that describes the nature of the seeds
therein.
[0243] The invention will be further described in the following
examples, which do not limit the scope of the invention described
in the claims.
VII. EXAMPLES
Example 1
Transgenic Arabidopsis Plants
[0244] The following symbols are used in the Examples with respect
to Arabidopsis transformation: T.sub.1: first generation
transformant; T.sub.2: second generation, progeny of
self-pollinated T.sub.1 plants; T.sub.3: third generation, progeny
of self-pollinated T.sub.2 plants; T.sub.4: fourth generation,
progeny of self-pollinated T.sub.3 plants. Independent
transformations are referred to as events.
[0245] Nucleic acids that were isolated from Arabidopsis thaliana
plants, and cloned into a Ti plasmid vector, CRS338, under the
control of a 35S promoter, p32449 promoter, or p326 promoter, as
indicated. Each construct contained a phosphinothricin
acetyltransferase gene which confers Finale.TM. resistance to
transformed plants. Wild-type Arabidopsis thaliana ecotype
Wassilewskija (Ws) plants were transformed separately with each
construct. The transformations were performed essentially as
described in Bechtold et al., C.R. Acad. Sci. Paris, 316:1194-1199
(1993). introduced into Arabidopsis plants.
[0246] Transgenic Arabidopsis lines containing SEQ ID NO:441, SEQ
ID NO:488, SEQ ID NO:567, SEQ ID NO:589, SEQ ID NO:704, SEQ ID
NO:710, SEQ ID NO:1241, SEQ ID NO:1260, SEQ ID NO:1128, SEQ ID
NO:59, SEQ ID NO:363, SEQ ID NO:310, SEQ ID NO:159, SEQ ID NO:1,
SEQ ID NO:1008, or SEQ ID NO:56 were designated ME00029, ME00045,
ME02190, ME02549, ME02865, ME03227, ME04477, ME18396, ME20095,
ME02932, ME18240, ME20867, ME03268, ME06551, ME02401, or ME06919,
respectively. The presence of each vector containing a nucleic acid
described above in the respective transgenic Arabidopsis line
transformed with the vector was confirmed by Finale.TM. resistance,
PCR amplification from green leaf tissue extract, and/or sequencing
of PCR products.
Example 2
Screening for Drought Tolerance in Transgenic Arabidopsis
Seedlings
[0247] Soil was made from 60% Sunshine Mix #5 (Sun Gro; Bellevue,
Wash., USA) and 40% vermiculite. For each superpool, 3 flats were
seeded with about 3000 seeds. Prior to seeding, each flat was
watered with 2 L of filtered water and the surface was misted. The
flats were covered with a humidity dome and kept in the dark at
4.degree. C. for 3 days.
[0248] After 3 days of cold treatment, the flats covered with
humidity domes were transferred to a greenhouse and covered with a
shade cloth (day 1). The humidity domes were removed on day 4, or
when cotyledons are fully expanded. Each flat was watered with 2 L
water within 1 day of removing the humidity domes by adding
filtered water to the bottom of the flat and allowing 30-45 minutes
for absorption of the water. On days 6 and 8, the flats were
assessed for Finale.TM. resistance. On day 10, the no-hole flats
were removed from the bottom of each flat to speed soil drying. The
flats were watered in no-hole flats as above after 2.5 weeks and 4
weeks. Candidates with increased height, increased branching,
normal siliques, larger rosettes, and persistent flowering, as
compared to the wild-type Ws seedlings that were grown and treated
in the same conditions were selected for analysis in a chronic soil
drought assay.
[0249] Chronic Soil Drought Assay
[0250] Soil was made from 60% Sunshine Mix #5 (Sun Gro; Bellevue,
Wash., USA) and 40% vermiculite. For each candidate event, 24 pots
were prepared in a 24-pot no-hole flat with 28 grams of dry soil in
each pot, and 4 L of filtered water was added to the tray. The
water was allowed to soak into the soil and the soil surface was
misted before seeding. For each candidate event, 18 pots were
seeded with 3-5 seeds each of candidate seeds and 6 pots were
seeded with 3-5 seeds each of wild-type control seeds. The seeded
pots were covered with a humidity dome and kept in the dark at
4.degree. C. for 3 days.
[0251] After 3 days of cold treatment, the pots covered with
humidity domes were transferred to a Conviron.RTM. growth chamber
set at 22.degree. C., 16:8 hour light:dark cycle, 70% humidity, and
120 .mu.E light intensity. The humidity domes were removed on day
5, or when cotyledons are fully expanded. After removal of the
domes, each pot was irrigated to saturation with 0.5.times.
Hoagland's solution, allowing the excess solution to drain.
Seedlings were then thinned to 1 per pot. On day 8 or 9, the pots
were irrigated to saturation with filtered water, allowing the
excess water to drain after about 30 minutes of soaking, and the
weight of each 24-pot flat was recorded. Each pot contained about
107 g water at saturation. Each pot was allowed to dry to about 15%
of saturation between waterings. 30 g of water was added at each
watering. Rosette area was measured at day 19. On day 24, plants
were staked and measurements were taken of the longest leaf and the
plant height. Terminal plant height was measured at the cease of
flowering. The plants were allowed to dry and seed weight was
measured. Finale resistance of a cauline leaf was determined at day
24. Significance of drought tolerance measurements was assessed
using a one-tailed Student's t-test, assuming unequal variance, at
p.ltoreq.0.5.
Example 3
Screening for Heat Tolerance in Transgenic Arabidopsis
Seedlings
[0252] Agar plates were made from the media composed of 2.15 g/L MS
salt (PhytoTechnology Laboratories.RTM.), 5 g/L sucrose (Sigma),
0.7% agar (PhytoTechnology Laboratories.RTM.), pH 5.7. For each
superpool, 3000 seeds were sterilized using 30% Clorox containing
0.1% Triton X-100 and plated onto two agar plates at a density of
1500 seeds per plate. The plates were wrapped with vent tape and
kept in the dark at 4.degree. C. for 3 days.
[0253] After 3 days of stratification, the plates were transferred
to a Conviron.RTM. growth chamber set at 22.degree. C., 16:8 hour
light:dark cycle, 70% humidity, and 60 .mu.E light intensity. The
seedlings were grown in the chamber for 3.5-4 days. For heat shock,
the plates were placed in an incubator set at 47.degree. C. for 2
hours. After heat shock, the plates were transferred back to the
Conviron.RTM. growth chamber and the seedlings were allowed to
recover and grow for several days (about 7 days). Candidates with
enhanced root growth and greenness of leaves, faster emergence of
photosynthetically active true leaves and increased size of
rosettes, as compared to the wild-type Ws seedlings that were grown
and treated in the same conditions were selected for analysis in a
heat shock assay.
[0254] Heat Shock I Assay
[0255] Seeds were plated on heat germination agar plates containing
45 mL of half-strength MS medium (2.15 g MS salt (PhytoTechnology
Laboratories.RTM.), 5 g sucrose (Sigma), and 0.7% agar
(PhytoTechnology Laboratories.RTM.) per liter, pH 5.7) per Petri
dish (square, 100 mm.times.15 mm). One transgenic event (40 seeds)
and 9 wild-type Ws control seeds were plated on each plate. Plates
were wrapped with vent tape and placed in the dark at 4.degree. C.
for 3 days. Plates were then transferred into a Conviron.RTM.
growth chamber set at 22.degree. C., 16:8 hour light:dark cycle,
and 70% humidity with fluorescent lamps emitting a light intensity
of about 60 .mu.E.
[0256] After the plants were grown in the Conviron.RTM. growth
chamber for 3-4 days, the plates were treated at 47.degree. C. in a
Yamato IC800 incubator for 1 hour 50 minutes. Following the heat
treatment, the plates were placed immediately back into the
Conviron.RTM. growth chamber. On the 7th, 9th, and 11th day at
22.degree. C. after the heat treatment, the plates were scanned
using Epson Perfection 4870 scanner. The area of each seedling
(green area) was quantified using the WinRhizo software (Regent
Instruments). The transgenic status of each plant was assessed by
Finale.TM. resistance.
[0257] The seedling area (a measure of growth after heat treatment)
of transgenic plants was compared to that of the pooled controls
comprising non-transgenic segregants and wild-type Ws controls
grown on the same plate. Significance of heat tolerance was
assessed using a one-tailed Student's t-test, assuming unequal
variance, at p.ltoreq.0.5.
[0258] Heat Shock II Assay
[0259] Seeds were plated on heat germination agar plates containing
45 mL of half-strength MS medium (2.15 g MS salt (PhytoTechnology
Laboratories.RTM.), 5 g sucrose (Sigma-Aldrich.RTM.), and 0.7% agar
(PhytoTechnology Laboratories.RTM.) per liter, pH 5.7) per Petri
dish (square, 100 mm.times.15 mm). One transgenic event (30 seeds),
4 wild-type Ws control seeds, and 2 seeds of a transgenic positive
control were plated on each plate. Plates were wrapped with vent
tape and placed in the dark at 4.degree. C. for 3 days. Plates were
then transferred into a Conviron.RTM. growth chamber set at
22.degree. C., 16:8 hour light:dark cycle, and 70% humidity with
fluorescent lamps emitting a light intensity of about 60 .mu.E.
[0260] After the plants were grown in the Conviron.RTM. growth
chamber for 11 days, the plates were scanned using a CF imager
(Technologica) to record the PSII operating efficiency
(.DELTA..PHI..sub.PSII) and seedling area. The plates were then
transferred to a Conviron.RTM. reach-in heat chamber set at
42.degree. C. for 5 hours. Following the heat treatment, the plants
were immediately scanned using the CF imager, and then transferred
back into the Conviron.RTM. growth chamber. After 2 and 4 days of
recovery in the growth chamber, the plates were scanned using the
CF imager again. The transgenic status of each plant was assessed
by Finale.TM. resistance.
[0261] The differences in the PSII operating efficiency and
seedling size prior to heat shock, immediately after heat shock,
and after recovery following heat shock were calculated. The
differences in the PSII operating efficiency and seedling size of
transgenic plants were compared to the pooled controls comprising
non-transgenic segregants and wild-type Ws controls grown on the
same plate. Significance of heat tolerance was assessed using a
one-tailed Student's t-test, assuming unequal variance, at
p.ltoreq.0.5.
Example 4
Results for ME00029 Events
[0262] T.sub.2 and T.sub.3 seed from two events of ME00029
containing SEQ ID NO:441 under control of a p32449 promoter was
analyzed for rosette area, terminal plant height, and seed yield as
described in Example 2. The results are provided in Table 1.
TABLE-US-00001 TABLE 1 Chronic drought response of seedlings from
ME00029 Control Population Plants Measurement Internal Pooled
T.sub.2 seedlings from event -01 of Rosette Area N.S. Sig. ME00029
T.sub.2 seedlings from event -04 of Rosette Area N.A. Sig. ME00029
T.sub.2 seedlings from event -01 of Longest Leaf N.S. Sig. ME00029
T.sub.2 seedlings from event -01 of Terminal Plant Sig. Sig.
ME00029 Height T.sub.3 seedlings from event -01 of Terminal Plant
Sig. Sig. ME00029 Height T.sub.2 seedlings from event -04 of
Terminal Plant N.A. Sig. ME00029 Height T.sub.3 seedlings from
event -04 of Terminal Plant N.A. Sig. ME00029 Height T.sub.3
seedlings from event -01 of Seed Yield N.S. Sig. ME00029
N.A.--results not available due to insufficient sample size (n <
5); N.S.--not significant; Sig.--significant (p .ltoreq. 0.5).
Example 5
Results for ME00045 Events
[0263] T.sub.2 and T.sub.3 seed from three events of ME00045
containing SEQ ID NO:488 under control of a p32449 promoter was
analyzed for rosette area, terminal plant height, longest leaf
length, and seed yield as described in Example 2. The results are
provided in Table 2.
TABLE-US-00002 TABLE 2 Chronic drought response of seedlings from
ME00045 Control Population Plants Measurement Internal Pooled Ws
T.sub.3 seedlings from event -02 of Rosette Area N.S. N.S. Sig.
ME00045 T.sub.3 seedlings from event -02 of Rosette Area N.S. N.S.
N.S. ME00045 T.sub.2 seedlings from event -07 of Rosette Area N.A.
N.S. Sig. ME00045 T.sub.2 seedlings from event -07 of Rosette Area
N.S. N.S. Sig. ME00045 T.sub.2 seedlings from event -08 of Rosette
Area N.A. N.S. Sig. ME00045 T.sub.2 seedlings from event -08 of
Rosette Area N.S. N.S. N.S. ME00045 T.sub.3 seedlings from event
-02 of Longest Leaf Sig. Sig. Sig. ME00045 T.sub.3 seedlings from
event -02 of Longest Leaf N.S. N.S. N.S. ME00045 T.sub.2 seedlings
from event -07 of Longest Leaf N.A. N.S. Sig. ME00045 T.sub.2
seedlings from event -07 of Longest Leaf N.S. N.S. N.S. ME00045
T.sub.2 seedlings from event -08 of Longest Leaf N.A. N.S. Sig.
ME00045 T.sub.2 seedlings from event -08 of Longest Leaf N.S. N.S.
N.S. ME00045 T.sub.3 seedlings from event -02 of Terminal Plant
N.S. N.S. N.S. ME00045 Height T.sub.3 seedlings from event -02 of
Terminal Plant N.S. N.S. N.S.* ME00045 Height T.sub.2 seedlings
from event -07 of Terminal Plant N.A. Sig. Sig. ME00045 Height
T.sub.2 seedlings from event -07 of Terminal Plant N.S. Sig. Sig.
ME00045 Height T.sub.2 seedlings from event -08 of Terminal Plant
N.A. N.S.* N.S. ME00045 Height T.sub.2 seedlings from event -08 of
Terminal Plant N.S. N.S. Sig. ME00045 Height T.sub.3 seedlings from
event -02 of Seed Yield Sig. N.S.* Sig. ME00045 T.sub.3 seedlings
from event -02 of Seed Yield N.S. N.S. N.S. ME00045 T.sub.2
seedlings from event -07 of Seed Yield N.A. N.S. N.S. ME00045
T.sub.2 seedlings from event -07 of Seed Yield N.S. N.S. N.S.
ME00045 T.sub.2 seedlings from event -08 of Seed Yield N.A. N.S.*
Sig. ME00045 T.sub.2 seedlings from event -08 of Seed Yield N.S.
N.S. Sig. ME00045 N.A.--results not available due to insufficient
sample size; N.S.--not significant (p > 0.10); N.S.*--not
significant (0.5 < p < 0.10); Sig.--significant (p < 0.5).
Data in bold represents a second independent replicate.
Example 6
Results for ME02190 Events
[0264] T.sub.2 and T.sub.4 seed from two events of ME02190
containing SEQ ID NO:567 under control of a 35S promoter was
analyzed for rosette area, terminal plant height, and longest leaf
length as described in Example 2. The results are provided in Table
3.
TABLE-US-00003 TABLE 3 Chronic drought response of seedlings from
ME02190 Control Population Plants Measurement Internal Pooled
T.sub.2 seedlings from event -10 of Rosette Area N.S. Sig. ME02190
T.sub.4 seedlings from event -02 of Longest Leaf N.A. Sig. ME02190
T.sub.2 seedlings from event -10 of Longest Leaf N.S. Sig. ME02190
T.sub.4 seedlings from event -02 of Terminal Plant N.A. Sig.
ME02190 Height T.sub.2 seedlings from event -10 of Terminal Plant
N.S. Sig. ME02190 Height N.A.--results not available due to
insufficient sample size (n < 5); N.S.--not significant;
Sig.--significant (p .ltoreq. 0.5).
Example 7
Results for ME02549 Events
[0265] T.sub.2 and T.sub.3 seed from three events of ME02549
containing SEQ ID NO:589 under control of a 35S promoter was
analyzed for rosette area, terminal plant height, and longest leaf
length as described in Example 2. The results are provided in Table
4.
TABLE-US-00004 TABLE 4 Chronic drought response of seedlings from
ME02549 Control Population Plants Measurement Internal Pooled Ws
T.sub.2 seedlings from event -01 of Rosette Area Sig. N.S. Sig.
ME02549 T.sub.3 seedlings from event -01 of Rosette Area Sig. N.S.
N.S. ME02549 T.sub.2 seedlings from event -02 of Rosette Area N.S.*
N.S.* Sig. ME02549 T.sub.3 seedlings from event -02 of Rosette Area
N.S. N.S. Sig. ME02549 T.sub.2 seedlings from event -05 of Rosette
Area N.A. N.S. Sig. ME02549 T.sub.3 seedlings from event -05 of
Rosette Area N.S. N.S.* Sig. ME02549 T.sub.3 seedlings from event
-01 of Longest Leaf N.S. N.S. Sig. ME02549 T.sub.3 seedlings from
event -02 of Longest Leaf N.S. N.S. Sig. ME02549 T.sub.3 seedlings
from event -05 of Longest Leaf N.S. N.S. Sig. ME02549 T.sub.2
seedlings from event -01 of Terminal Plant N.A. N.S. N.S. ME02549
Height T.sub.3 seedlings from event -01 of Terminal Plant N.S. N.S.
Sig. ME02549 Height T.sub.3 seedlings from event -02 of Terminal
Plant N.S. Sig. Sig. ME02549 Height T.sub.2 seedlings from event
-05 of Terminal Plant N.A. N.S. N.S. ME02549 Height T.sub.3
seedlings from event -05 of Terminal Plant Sig. N.S. Sig. ME02549
Height N.A.--results not available due to insufficient sample size
(n < 5); N.S.--not significant; N.S.*--not significant (0.5 <
p < 0.10); Sig.--significant (p < 0.5).
Example 8
Results for ME02865 Events
[0266] T.sub.2 and T.sub.3 seed from two events of ME02865
containing SEQ ID NO:704 under control of a 35S promoter was
analyzed for rosette area, terminal plant height, longest leaf
length, and seed weight as described in Example 2. The results are
provided in Table 5.
TABLE-US-00005 TABLE 5 Chronic drought response of seedlings from
ME02865 Control Population Plants Measurement Internal Pooled
T.sub.2 seedlings from event -02 of Rosette Area N.S. N.S. ME02865
T.sub.3 seedlings from event -02-01 of Rosette Area -- N.S. ME02865
T.sub.3 seedlings from event -02-02 of Rosette Area N.S. N.S.
ME02865 T.sub.2 seedlings from event -05 of Rosette Area -- Sig.
ME02865 T.sub.3 seedlings from event -05-01 of Rosette Area -- Sig.
ME02865 T.sub.3 seedlings from event -05-02 of Rosette Area N.S.
N.S. ME02865 T.sub.2 seedlings from event -02 of Longest Leaf N.S.
N.S. ME02865 T.sub.3 seedlings from event -02-01 of Longest Leaf --
N.S. ME02865 T.sub.3 seedlings from event -02-02 of Longest Leaf
N.S.* Sig. ME02865 T.sub.2 seedlings from event -05 of Longest Leaf
-- Sig. ME02865 T.sub.3 seedlings from event -05-01 of Longest Leaf
-- N.S. ME02865 T.sub.3 seedlings from event -05-02 of Longest Leaf
N.S. Sig. ME02865 T.sub.2 seedlings from event -02 of Terminal
Plant Sig. Sig. ME02865 Height T.sub.3 seedlings from event -02-01
of Terminal Plant -- N.S.* ME02865 Height T.sub.3 seedlings from
event -02-02 of Terminal Plant N.S. Sig. ME02865 Height T.sub.2
seedlings from event -05 of Terminal Plant -- Sig. ME02865 Height
T.sub.3 seedlings from event -05-01 of Terminal Plant -- N.S.*
ME02865 Height T.sub.3 seedlings from event -05-02 of Terminal
Plant N.S.* Sig. ME02865 Height T.sub.2 seedlings from event -02 of
Seed Weight N.S. N.S. ME02865 T.sub.3 seedlings from event -02-01
of Seed Weight -- N.S. ME02865 T.sub.3 seedlings from event -02-02
of Seed Weight Sig. N.S. ME02865 T.sub.2 seedlings from event -05
of Seed Weight -- Sig. ME02865 T.sub.3 seedlings from event -05-01
of Seed Weight -- N.S. ME02865 T.sub.3 seedlings from event -05-02
of Seed Weight N.S. N.S.* ME02865 N.A.--results not available due
to insufficient sample size (n < 5); N.S.--not significant;
N.S.*--not significant (0.5 < p < 0.10); Sig.--significant (p
< 0.5).
[0267] The physical appearances of T.sub.1 ME02865 plants were
similar to those of corresponding control plants grown under
standard conditions. There were no observable or statistically
significant differences between T.sub.2 plants from events -02 and
-05 of ME02865 and control plants in germination, onset of
flowering, rosette area, fertility, and general
morphology/architecture when grown under standard conditions.
Example 9
Results for ME03227 Events
[0268] T.sub.2 and T.sub.3 seed from two events of ME03227
containing SEQ ID NO:710 under control of a 35S promoter was
analyzed for rosette area, terminal plant height, and longest leaf
length as described in Example 2. The results are provided in Table
6.
TABLE-US-00006 TABLE 6 Chronic drought response of seedlings from
ME03227 Control Population Plants Measurement Internal Pooled
T.sub.2 seedlings from event -01 of Rosette Area N.A. Sig. ME03227
T.sub.3 seedlings from event -01 of Rosette Area N.A. Sig. ME03227
T.sub.2 seedlings from event -05 of Rosette Area N.A. Sig. ME03227
T.sub.3 seedlings from event -05 of Rosette Area N.A. Sig. ME03227
T.sub.3 seedlings from event -05 of Longest Leaf N.A. Sig. ME03227
T.sub.3 seedlings from event -01 of Terminal Plant N.A. N.S.*
ME03227 Height T.sub.3 seedlings from event -05 of Terminal Plant
N.A. N.S.* ME03227 Height N.A.--results not available due to
insufficient sample size (n < 5); N.S.--not significant;
N.S.*--not significant (0.5 < p < 0.10); Sig.--significant (p
.ltoreq. 0.5).
Example 10
Results for ME04477 Events
[0269] T.sub.2 seed from five events of ME04477 containing SEQ ID
NO:1241 under control of a 35S promoter was analyzed for rosette
area, terminal plant height, and longest leaf length as described
in Example 2. The results are provided in Table 7.
TABLE-US-00007 TABLE 7 Chronic drought response of seedlings from
ME04477 Control Population Plants Measurement Internal Pooled
T.sub.2 seedlings from event -03 of Rosette Area N.A. Sig. ME04477
T.sub.2 seedlings from event -01 of Longest Leaf N.A. Sig. ME04477
T.sub.2 seedlings from event -02 of Longest Leaf N.A. Sig. ME04477
T.sub.2 seedlings from event -03 of Longest Leaf N.A. Sig. ME04477
T.sub.2 seedlings from event -04 of Longest Leaf Sig. Sig. ME04477
T.sub.2 seedlings from event -05 of Longest Leaf N.A. Sig. ME04477
T.sub.2 seedlings from event -01 of Terminal Plant N.A. Sig.
ME04477 Height T.sub.2 seedlings from event -02 of Terminal Plant
N.A. Sig. ME04477 Height T.sub.2 seedlings from event -03 of
Terminal Plant N.A. Sig. ME04477 Height T.sub.2 seedlings from
event -04 of Terminal Plant Sig. Sig. ME04477 Height T.sub.2
seedlings from event -05 of Terminal Plant N.A. Sig. ME04477 Height
N.A.--results not available due to insufficient sample size (n <
5); N.S.--not significant; Sig.--significant (p < 0.5).
Example 11
Results for ME18396 Events
[0270] T.sub.2 seed from one event of ME18396 containing SEQ ID
NO:1260 under control of a p326 promoter was analyzed for rosette
area and terminal plant height as described in Example 2. The
results are provided in Table 8.
TABLE-US-00008 TABLE 8 Chronic drought response of seedlings from
ME18396 Control Population Plants Measurement Internal Pooled
T.sub.2 seedlings from event -02 of Rosette Area N.S. Sig. ME18396
T.sub.2 seedlings from event -02 of Terminal Plant Sig. Sig.
ME18396 Height N.S.--not significant; Sig.--significant (p <
0.5).
Example 12
Results for ME20095 Events
[0271] T.sub.2 and T.sub.3 seed from two events of ME20095
containing SEQ ID NO:1128 under control of a 35S promoter was
analyzed for rosette area, longest leaf length, terminal plant
height, and seed yield as described in Example 2. The results are
provided in Table 9.
TABLE-US-00009 TABLE 9 Chronic drought response of seedlings from
ME20095 Control Population Plants Measurement Internal Pooled
T.sub.2 seedlings from event -01 of Rosette Area Sig. Sig. ME20095
T.sub.3 seedlings from event -01-01 of Rosette Area N.S. N.S.
ME20095 T.sub.3 seedlings from event -01-02 of Rosette Area N.S.
N.S. ME20095 T.sub.2 seedlings from event -04 of Rosette Area Sig.
Sig. ME20095 T.sub.3 seedlings from event -04-01 of Rosette Area
N.S. N.S. ME20095 T.sub.3 seedlings from event -04-02 of Rosette
Area N.S. N.S.* ME20095 T.sub.2 seedlings from event -01 of Longest
Leaf N.S. Sig. ME20095 T.sub.3 seedlings from event -01-01 of
Longest Leaf Sig. Sig. ME20095 T.sub.3 seedlings from event -01-02
of Longest Leaf N.S. N.S. ME20095 T.sub.2 seedlings from event -04
of Longest Leaf Sig. Sig. ME20095 T.sub.3 seedlings from event
-04-01 of Longest Leaf Sig. Sig. ME20095 T.sub.3 seedlings from
event -04-02 of Longest Leaf N.S.* Sig. ME20095 T.sub.2 seedlings
from event -01 of Terminal Plant N.S. Sig. ME20095 Height T.sub.3
seedlings from event -01-01 of Terminal Plant N.S.* Sig. ME20095
Height T.sub.3 seedlings from event -01-02 of Terminal Plant N.S.
N.S.* ME20095 Height T.sub.2 seedlings from event -04 of Terminal
Plant Sig. Sig. ME20095 Height T.sub.3 seedlings from event -04-01
of Terminal Plant Sig. Sig. ME20095 Height T.sub.3 seedlings from
event -04-02 of Terminal Plant N.S. Sig. ME20095 Height T.sub.2
seedlings from event -01 of Seed Weight Sig. Sig. ME20095 T.sub.3
seedlings from event -01-01 of Seed Weight Sig. N.S.* ME20095
T.sub.3 seedlings from event -01-02 of Seed Weight Sig. Sig.
ME20095 T.sub.2 seedlings from event -04 of Seed Weight N.S. N.S.
ME20095 T.sub.3 seedlings from event -04-01 of Seed Weight N.S.
N.S. ME20095 T.sub.3 seedlings from event -04-02 of Seed Weight
N.S. N.S. ME20095 N.S.--not significant; N.S.*--not significant
(0.5 < p < 0.10); Sig.--significant (p < 0.5).
[0272] T.sub.1 ME20095 plants showed delayed flowering and
prolonged vegetative growth compared to corresponding control
plants grown under standard conditions. There were no observable or
statistically significant differences between T.sub.2 plants from
events -02 and -04 of ME20095 and control plants in germination,
onset of flowering, fertility, and general morphology/architecture
when grown under standard conditions. Rosette area in T2 plants
from events -02 and -04 was significantly larger and seed yield was
increased as compared to corresponding control plants grown under
standard conditions.
Example 13
Results for ME02401 Events
[0273] T.sub.2 and T.sub.3 seed from two events of ME02401
containing SEQ ID NO:1188 under control of a 35S promoter was
analyzed for rosette area, terminal plant height, and longest leaf
length as described in Example 2. The results are provided in Table
10.
TABLE-US-00010 TABLE 10 Chronic drought response of seedlings from
ME02401 Control Population Plants Measurement Internal Pooled
T.sub.2 seedlings from event -05 of Rosette Area N.A. N.S.* ME02401
T.sub.3 seedlings from event -01 of Longest Leaf N.S.* Sig. ME02401
T.sub.2 seedlings from event -05 of Longest Leaf N.A. Sig. ME02401
T.sub.3 seedlings from event -01 of Terminal Plant Sig. Sig.
ME02401 Height T.sub.2 seedlings from event -05 of Terminal Plant
N.A. Sig. ME02401 Height N.A.--results not available due to
insufficient sample size (n < 5); N.S.--not significant;
N.S.*--not significant (0.5 < p < 0.10); Sig.--significant (p
< 0.5).
Example 14
Results for ME02932 Events
[0274] T.sub.2 and T.sub.3 seed from two events of ME02932
containing SEQ ID NO:59 under control of a 35S promoter was
analyzed for seedling area following a Heat Shock I assay as
described in Example 3. The results are provided in Table 11.
TABLE-US-00011 TABLE 11 ME02932 seedling area following Heat Shock
I assay Average seedling Plants area SE N p-value T.sub.2 seedlings
from event -01 of ME02932 0.07 0.008 33 8.77E-04 T.sub.2
non-transgenic segregants of event -01 of 0.04 0.006 15 ME02932
T.sub.3 seedlings from event -01 of ME02932 0.10 0.003 40 2.21E-04
T.sub.3 non-transgenic segregants of event -01 of 0.07 0.007 9
ME02932 T.sub.2 seedlings from event -03 of ME02932 0.08 0.012 21
1.29E-02 T.sub.2 non-transgenic segregants of event -03 of 0.04
0.009 23 ME02932 T.sub.3 seedlings from event -03 of ME02932 0.09
0.004 36 1.54E-03 T.sub.3 non-transgenic segregants of event -03 of
0.07 0.005 13 ME02932
[0275] The physical appearances of T.sub.1 ME02932 plants were
similar to those of corresponding control plants grown under
standard conditions. There were no observable or statistically
significant differences between T.sub.2 plants from events -01 and
-03 of ME02932 and control plants in germination, onset of
flowering, rosette area, fertility, and general
morphology/architecture when grown under standard conditions.
Example 15
Results for ME18240 Events
[0276] T.sub.2 and T.sub.3 seed from three events of ME18240
containing SEQ ID NO:363 under control of a 35S promoter was
analyzed for seedling area following a Heat Shock I assay and
change in photosynthetic activity during a Heat Shock II assay as
described in Example 3. The results are provided in Tables 12 and
13.
TABLE-US-00012 TABLE 12 ME18240 seedling area following Heat Shock
I assay Average seedling Plants area SE N p-value T.sub.2 seedlings
from event -02 of ME18240 0.08 0.007 27 4.22E-08 T.sub.2
non-transgenic segregants of event -02 of 0.03 0.004 16 ME18240
T.sub.3 seedlings from event -02 of ME18240 0.14 0.007 31 5.83E-11
T.sub.3 non-transgenic segregants of event -02 of 0.06 0.007 14
ME18240 T.sub.2 seedlings from event -03 of ME18240 0.13 0.007 35
3.31E-05 T.sub.2 non-transgenic segregants of event -03 of 0.08
0.01 14 ME18240 T.sub.3 seedlings from event -03 of ME18240 0.14
0.008 28 5.13E-06 T.sub.3 non-transgenic segregants of event -03 of
0.09 0.008 19 ME18240 T.sub.2 seedlings from event -04 of ME18240
0.13 0.008 33 2.39E-07 T.sub.2 non-transgenic segregants of event
-04 of 0.06 0.009 16 ME18240 T.sub.3 seedlings from event -04 of
ME18240 0.13 0.008 29 4.68E-05 T.sub.3 non-transgenic segregants of
event -04 of 0.08 0.008 18 ME18240
TABLE-US-00013 TABLE 13 ME18240 .DELTA.Fv/Fm in response to Heat
Shock II assay Average Plants .DELTA.Fv/Fm SE N p-value T.sub.2
seedlings from event -02 of ME18240 0.24 0.013 47 3.64E-05 T.sub.2
non-transgenic segregants of event -02 of 0.32 0.016 25 ME18240
T.sub.3 seedlings from event -02 of ME18240 0.36 0.012 43 1.92E-04
T.sub.3 non-transgenic segregants of event -02 of 0.43 0.016 22
ME18240 T.sub.2 seedlings from event -03 of ME18240 0.24 0.010 41
9.27E-11 T.sub.2 non-transgenic segregants of event -03 of 0.37
0.011 28 ME18240 T.sub.3 seedlings from event -03 of ME18240 0.33
0.011 24 1.91E-02 T.sub.3 non-transgenic segregants of event -03 of
0.39 0.022 10 ME18240 T.sub.2 seedlings from event -04 of ME18240
0.21 0.009 43 5.55E-17 T.sub.2 non-transgenic segregants of event
-04 of 0.40 0.007 29 ME18240 T.sub.3 seedlings from event -04 of
ME18240 0.28 0.011 46 9.38E-11 T.sub.3 non-transgenic segregants of
event -04 of 0.41 0.008 21 ME18240 Where .DELTA.Fv/Fm = Fv/Fm prior
to heat shock - Fv/Fm after heat shock.
[0277] The physical appearances of T.sub.1 ME18240 plants were
similar to those of corresponding control plants grown under
standard conditions. There were no observable or statistically
significant differences between T.sub.2 plants from events -02,
-03, and -04 of ME18240 and control plants in germination, onset of
flowering, rosette area, fertility, and general
morphology/architecture when grown under standard conditions.
Example 16
Results for ME20867 Events
[0278] T.sub.2 and T.sub.3 seed from four events of ME20867
containing SEQ ID NO:310 under control of a 35S promoter was
analyzed for seedling area following a Heat Shock I assay and
change in photosynthetic activity during a Heat Shock II assay as
described in Example 3. The results are provided in Tables 14 and
15.
TABLE-US-00014 TABLE 14 ME20867 seedling area in response to Heat
Shock I assay Average seedling Plants area SE N p-value T.sub.2
seedlings from event -01 of ME20867 0.14 0.008 28 4.86E-07 T.sub.2
non-transgenic segregants of event -01 of 0.07 0.011 12 ME20867
T.sub.3 seedlings from event -01 of ME20867 0.07 0.007 31 9.84E-10
T.sub.3 non-transgenic segregants of event -01 of 0.01 0.010 16
ME20867 T.sub.2 seedlings from event -02 of ME20867 0.13 0.006 37
1.03E-02 T.sub.2 non-transgenic segregants of event -02 of 0.08
0.024 3 ME20867 T.sub.3 seedlings from event -02 of ME20867 0.09
0.006 32 6.01E-07 T.sub.3 non-transgenic segregants of event -02 of
0.02 0.011 9 ME20867 T.sub.2 seedlings from event -04 of ME20867
0.12 0.006 31 1.47E-02 T.sub.2 non-transgenic segregants of event
-04 of 0.10 0.010 9 ME20867 T.sub.3 seedlings from event -04 of
ME20867 0.03 0.005 20 2.14E-04 T.sub.3 non-transgenic segregants of
event -04 of 0.01 0.007 12 ME20867 T.sub.2 seedlings from event -05
of ME20867 0.11 0.009 30 1.51E-02 T.sub.2 non-transgenic segregants
of event -05 of 0.08 0.012 10 ME20867 T.sub.3 seedlings from event
-05 of ME20867 0.03 0.004 23 1.98E-04 T.sub.3 non-transgenic
segregants of event -05 of 0.01 0.005 20 ME20867
TABLE-US-00015 TABLE 15 ME20867 .DELTA.Fv/Fm-D.sub.2 in response to
Heat Shock II assay Average Plants .DELTA.Fv/Fm-D.sub.2 SE N
p-value T.sub.2 seedlings from event -01 of ME20867 0.03 0.012 33
2.17E-08 T.sub.2 non-transgenic segregants of event -01 of -0.07
0.008 12 ME20867 T.sub.3 seedlings from event -01 of ME20867 0.21
0.010 44 3.71E-12 T.sub.3 non-transgenic segregants of event -01 of
0.06 0.015 22 ME20867 T.sub.2 seedlings from event -02 of ME20867
0.06 0.011 25 9.39E-11 T.sub.2 non-transgenic segregants of event
-02 of -0.10 0.006 12 ME20867 T.sub.3 seedlings from event -02 of
ME20867 0.25 0.012 50 4.79E-08 T.sub.3 non-transgenic segregants of
event -02 of 0.06 0.029 16 ME20867 T.sub.2 seedlings from event -04
of ME20867 0.17 0.010 43 8.71E-11 T.sub.2 non-transgenic segregants
of event -04 of 0.03 0.014 22 ME20867 T.sub.3 seedlings from event
-04 of ME20867 0.16 0.011 24 1.39E-03 T.sub.3 non-transgenic
segregants of event -04 of 0.06 0.027 9 ME20867 T.sub.2 seedlings
from event -05 of ME20867 0.21 0.016 28 1.35E-09 T.sub.2
non-transgenic segregants of event -05 of 0.07 0.012 31 ME20867
T.sub.3 seedlings from event -05 of ME20867 0.24 0.015 19 3.72E-03
T.sub.3 non-transgenic segregants of event -05 of 0.18 0.015 14
ME20867 .DELTA.Fv/Fm-D.sub.2 = Fv/Fm immediately after heat shock -
Fv/Fm after 2 days recovery.
[0279] The physical appearances of T.sub.1 ME20867 plants were
similar to those of corresponding control plants grown under
standard conditions. There were no observable or statistically
significant differences between T.sub.2 plants from events -01,
-02, -04 and -05 of ME20867 and control plants in germination,
onset of flowering, rosette area, fertility, and general
morphology/architecture when grown under standard conditions.
Example 17
Results for ME03268 Events
[0280] T.sub.2 and T.sub.3 seed from one event of ME03268, and
T.sub.3 and T.sub.4 seed from one event of ME03268, each containing
SEQ ID NO:159 under control of a 35S promoter, was analyzed for
seedling area following a Heat Shock I assay as described in
Example 3. The results are provided in Table 16.
TABLE-US-00016 TABLE 16 ME03268 seedling area in response to Heat
Shock I assay Average seedling Plants area SE N p-value T.sub.2
seedlings from event -02 of ME03268 0.165 0.015 26 1.09E-03 T.sub.2
non-transgenic segregants of event -02 of 0.070 0.024 3 ME03268
T.sub.3 seedlings from event -02 of ME03268 0.137 0.009 40 1.97E-07
T.sub.3 non-transgenic segregants of event -02 of 0.048 0.012 9
ME03268 T.sub.3 seedlings from event -03 of ME03268 0.062 0.007 26
3.65E-02 T.sub.3 non-transgenic segregants of event -03 of 0.037
0.011 4 ME03268 T.sub.4 seedlings from event -03 of ME03268 0.094
0.008 30 3.96E-03 T.sub.4 non-transgenic segregants of event -03 of
0.054 0.012 10 ME03268
[0281] The physical appearances of T.sub.1 ME03268 plants were
similar to those of corresponding control plants grown under
standard conditions. There were no observable or statistically
significant differences between T.sub.2 plants from events -02 and
-03 of ME03268 and control plants in germination, onset of
flowering, rosette area, fertility, and general
morphology/architecture when grown under standard conditions.
Example 18
Results for ME06551 Events
[0282] T.sub.3 and T.sub.4 seed from two events of ME06551
containing SEQ ID NO:1 under control of a 35S promoter was analyzed
for change in photosynthetic activity during a Heat Shock II assay
as described in Example 3. The results are provided in Table17.
TABLE-US-00017 TABLE 17 ME06551 .DELTA.Fv/Fm in response to Heat
Shock II assay Average Plants .DELTA.Fv/Fm SE N p-value T.sub.3
seedlings from event -04 of ME06551 0.42 0.016 11 1.32E-02 T.sub.3
non-transgenic segregants of event -04 of 0.47 0.012 15 ME06551
T.sub.4 seedlings from event -04 of ME06551 0.39 0.006 49 4.73E-03
T.sub.4 non-transgenic segregants of event -04 of 0.43 0.014 11
ME06551 T.sub.3 seedlings from event -05 of ME06551 0.39 0.013 11
4.21E-04 T.sub.3 non-transgenic segregants of event -05 of 0.46
0.010 13 ME06551 T.sub.4 seedlings from event -05 of ME06551 0.39
0.007 50 4.79E-02 T.sub.4 non-transgenic segregants of event -05 of
0.41 0.010 17 ME06551 Where .DELTA.Fv/Fm = Fv/Fm prior to heat
shock - Fv/Fm after heat shock.
[0283] The physical appearances of T.sub.1 ME06551 plants were
similar to those of corresponding control plants grown under
standard conditions. There were no observable or statistically
significant differences between T.sub.2 plants from events -04 and
-05 of ME06551 and control plants in germination, onset of
flowering, rosette area, fertility, and general
morphology/architecture when grown under standard conditions.
Example 19
Results for ME06919 Events
[0284] T.sub.2 and T.sub.3 seed from two events of ME06919
containing SEQ ID NO:56 under control of a 35S promoter was
analyzed for change in photosynthetic activity during a Heat Shock
II assay as described in Example 3. The results are provided in
Table 18.
TABLE-US-00018 TABLE 18 ME06919 .DELTA.Fv/Fm in response to Heat
Shock II assay Average Plants .DELTA.Fv/Fm SE N p-value T.sub.2
seedlings from event -03 of ME06919 0.433 0.007 65 2.55E-09 T.sub.2
non-transgenic segregants of event -03 of 0.497 0.008 37 ME06919
T.sub.3 seedlings from event -03 of ME06919 0.281 0.013 46 1.87E-03
T.sub.3 non-transgenic segregants of event -03 of 0.341 0.015 18
ME06919 T.sub.2 seedlings from event -04 of ME06919 0.490 0.007 21
3.06E-04 T.sub.2 non-transgenic segregants of event -04 of 0.531
0.009 26 ME06919 T.sub.3 seedlings from event -04 of ME06919 0.296
0.006 44 6.15E-08 T.sub.3 non-transgenic segregants of event -04 of
0.412 0.018 18 ME06919 Where .DELTA.Fv/Fm = Fv/Fm prior to heat
shock - Fv/Fm after heat shock.
[0285] The physical appearances of T.sub.1 ME06919 plants were
similar to those of corresponding control plants grown under
standard conditions. There were no observable or statistically
significant differences between T.sub.2 plants from events -03 and
-04 of ME06919 and control plants in germination, onset of
flowering, rosette area, fertility, and general
morphology/architecture when grown under standard conditions.
Example 20
Results for ME04246 Events
[0286] T.sub.2 and T.sub.3 seed from two events of ME04246
containing SEQ ID NO:1362 under control of a 35S promoter was
analyzed for rosette area, terminal plant height, and longest leaf
length as described in Example 2. The results are provided in Table
19.
TABLE-US-00019 TABLE 19 Chronic drought response of seedlings from
ME04246 Control Population Plants Measurement Internal Pooled Ws
T.sub.2 seedlings from event -01 of ME04246 Longest Leaf Sig. Sig.
N.S. T.sub.2 seedlings from event -03 of ME04246 Longest Leaf N.S.
N.S. N.S. T.sub.2 seedlings from event -04 of ME04246 Longest Leaf
N.S. N.S. N.S. T.sub.2 seedlings from event -01 of ME04246 Dry
Biomass N.A. N.S.* Sig. T.sub.2 seedlings from event -02 of ME04246
Dry Biomass N.A. N.A. N.S. T.sub.2 seedlings from event -04 of
ME04246 Dry Biomass N.A. N.S.* Sig. N.A.--results not available due
to insufficient sample size (n < 5); N.S.--not significant;
N.S.*--not significant (0.5 < p < 0.10); Sig.--significant (p
< 0.5).
[0287] T.sub.2 ME04246 plants showed increased vegetative biomass
compared to corresponding control plants under water-limiting
conditions as measured directly and to a lesser degree by longest
leaf length. There were no observable or statistically significant
differences between T.sub.1 plants from events -01, -03 and -04 of
ME04246 and control plants in germination, onset of flowering,
fertility, and general morphology/architecture when grown under
standard conditions.
Example 21
Determination of Functional Homologs by Reciprocal BLAST
[0288] A candidate sequence was considered a functional homolog of
a reference sequence if the candidate and reference sequences
encoded proteins having a similar function and/or activity. A
process known as Reciprocal BLAST (Rivera et al., Proc. Natl. Acad.
Sci. USA, 95:6239-6244 (1998)) was used to identify potential
functional homolog sequences from databases consisting of all
available public and proprietary peptide sequences, including NR
from NCBI and peptide translations from Ceres clones.
[0289] Before starting a Reciprocal BLAST process, a specific
reference polypeptide was searched against all peptides from its
source species using BLAST in order to identify polypeptides having
BLAST sequence identity of 80% or greater to the reference
polypeptide and an alignment length of 85% or greater along the
shorter sequence in the alignment. The reference polypeptide and
any of the aforementioned identified polypeptides were designated
as a cluster.
[0290] The BLASTP version 2.0 program from Washington University at
Saint Louis, Mo., USA was used to determine BLAST sequence identity
and E-value. The BLASTP version 2.0 program includes the following
parameters: 1) an E-value cutoff of 1.0e-5; 2) a word size of 5;
and 3) the -postsw option. The BLAST sequence identity was
calculated based on the alignment of the first BLAST HSP
(High-scoring Segment Pairs) of the identified potential functional
homolog sequence with a specific reference polypeptide. The number
of identically matched residues in the BLAST HSP alignment was
divided by the HSP length, and then multiplied by 100 to get the
BLAST sequence identity. The HSP length typically included gaps in
the alignment, but in some cases gaps were excluded.
[0291] The main Reciprocal BLAST process consists of two rounds of
BLAST searches; forward search and reverse search. In the forward
search step, a reference polypeptide sequence, "polypeptide A,"
from source species SA was BLASTed against all protein sequences
from a species of interest. Top hits were determined using an
E-value cutoff of 10.sup.-5 and a sequence identity cutoff of 35%.
Among the top hits, the sequence having the lowest E-value was
designated as the best hit, and considered a potential functional
homolog or ortholog. Any other top hit that had a sequence identity
of 80% or greater to the best hit or to the original reference
polypeptide was considered a potential functional homolog or
ortholog as well. This process was repeated for all species of
interest.
[0292] In the reverse search round, the top hits identified in the
forward search from all species were BLASTed against all protein
sequences from the source species SA. A top hit from the forward
search that returned a polypeptide from the aforementioned cluster
as its best hit was also considered as a potential functional
homolog.
[0293] Functional homologs were identified by manual inspection of
potential functional homolog sequences. Representative functional
homologs for SEQ ID NO:442, SEQ ID NO:489, SEQ ID NO:568, SEQ ID
NO:590, SEQ ID NO:705, SEQ ID NO:711, SEQ ID NO:742, SEQ ID
NO:1012, SEQ ID NO:1129, SEQ ID NO:60, SEQ ID NO:364, SEQ ID
NO:311, SEQ ID NO:160, SEQ ID NO:2, and SEQ ID NO:749 are shown in
FIGS. 1-15, respectively.
Example 22
Determination of Functional Homologs by Hidden Markov Models
[0294] Hidden Markov Models (HMMs) were generated by the program
HMMER 2.3.2. To generate each HMM, the default HMMER 2.3.2 program
parameters, configured for global alignments, were used.
[0295] An HMM was generated using the sequences shown in FIG. 1 as
input. These sequences were fitted to the model and a
representative HMM bit score for each sequence is shown in the
Sequence Listing. Additional sequences were fitted to the model,
and representative HMM bit scores for any such additional sequences
are shown in the Sequence Listing. The results indicate that these
additional sequences are functional homologs of SEQ ID NO:442.
[0296] The procedure above was repeated and an HMM was generated
for each group of sequences shown in FIGS. 2-15 using the sequences
shown in each Figure as input for that HMM. A representative bit
score for each sequence is shown in the Sequence Listing.
Additional sequences were fitted to certain HMMs, and
representative HMM bit scores for such additional sequences are
shown in the Sequence Listing. The results indicate that these
additional sequences are functional homologs of the sequences used
to generate that HMM.
Other Embodiments
[0297] It is to be understood that while the invention has been
described in conjunction with the detailed description thereof, the
foregoing description is intended to illustrate and not limit the
scope of the invention, which is defined by the scope of the
appended claims. Other aspects, advantages, and modifications are
within the scope of the following claims.
Sequence CWU 0 SQTB SEQUENCE LISTING The patent application
contains a lengthy "Sequence Listing" section. A copy of the
"Sequence Listing" is available in electronic form from the USPTO
web site
(http://seqdata.uspto.gov/?pageRequest=docDetail&DocID=US20110023193A1).
An electronic copy of the "Sequence Listing" will also be available
from the USPTO upon request and payment of the fee set forth in 37
CFR 1.19(b)(3).
0 SQTB SEQUENCE LISTING The patent application contains a lengthy
"Sequence Listing" section. A copy of the "Sequence Listing" is
available in electronic form from the USPTO web site
(http://seqdata.uspto.gov/?pageRequest=docDetail&DocID=US20110023193A1).
An electronic copy of the "Sequence Listing" will also be available
from the USPTO upon request and payment of the fee set forth in 37
CFR 1.19(b)(3).
* * * * *
References