园艺学报 ›› 2021, Vol. 48 ›› Issue (12): 2458-2470.doi: 10.16420/j.issn.0513-353x.2021-0382
张婷婷1, 李雨欣1, 张德遥1, 康宇乾1, 王健2, 宋希强2, 周扬1,*()
收稿日期:
2021-07-20
修回日期:
2021-08-26
发布日期:
2022-01-04
通讯作者:
周扬
E-mail:zhouyang@hainanu.edu.cn
基金资助:
ZHANG Tingting1, LI Yuxin1, ZHANG Deyao1, KANG Yuqian1, WANG Jian2, SONG Xiqiang2, ZHOU Yang1,*()
Received:
2021-07-20
Revised:
2021-08-26
Published:
2022-01-04
Contact:
ZHOU Yang
E-mail:zhouyang@hainanu.edu.cn
摘要:
利用生物信息学方法对铁皮石斛(Dendrobium catenatum Lindl.)2C型蛋白磷酸酶(protein phosphatase 2C,PP2C)家族基因进行鉴定分析,采用实时荧光定量PCR技术分析PP2C基因的组织表达特性,以及在不同非生物胁迫和植物生长调节剂处理下的表达模式。共鉴定出67个家族成员,其编码的氨基酸在80 ~ 1 075 aa之间,分子量在8.86 ~ 119.59 kD之间,等电点在4.13 ~ 9.36之间。进化树分析可将其分为13个亚族,且每个亚族蛋白的保守基元相似。基因结构分析表明,PP2C基因的外显子数量为1 ~ 15不等。空间表达模式分析发现有4个PP2C在铁皮石斛合蕊柱中表达量最高,5个基因在蒴果中表达量最高,有1个基因在根中表达量最高,PP2C的表达具有组织特异性。DcPP2C5、DcPP2C20和DcPP2C56表达量受干旱胁迫(20% PEG6000)上调较明显,DcPP2C24受高温(42 ℃)诱导最显著,DcPP2C5受低温(4 ℃)诱导上调表达,DcPP2C20受到盐胁迫(200 mmol · L-1 NaCl)诱导,这些基因的表达均在不同程度上受100 μmol · L-1ABA和20 μmol · L-1 SA的诱导,说明它们在铁皮石斛应对逆境胁迫中发挥作用。
中图分类号:
张婷婷, 李雨欣, 张德遥, 康宇乾, 王健, 宋希强, 周扬. 铁皮石斛蛋白磷酸酶PP2C家族基因鉴定及其表达分析[J]. 园艺学报, 2021, 48(12): 2458-2470.
ZHANG Tingting, LI Yuxin, ZHANG Deyao, KANG Yuqian, WANG Jian, SONG Xiqiang, ZHOU Yang. Genome-Wide Identification and Expression Analyses of PP2C Gene Family in Dendrobium catenatum[J]. Acta Horticulturae Sinica, 2021, 48(12): 2458-2470.
基因 Gene | 上游引物(5′-3′) Forward primer | 下游引物(5′-3′) Reverse primer |
---|---|---|
ACTIN | TTGTGTTGGATTCTGGTGATGGTGT | TTTCCCGTTCTGCTGTTGTTGTGAA |
DcPP2C5 | CTTTGAAGATGTTGAGTTCCCAC | TGCTTTCCGCACTGAATCCT |
DcPP2C6 | ATCACTGTTGTGGACCTTTGCG | CGCACACGGCAGAGAGGCTA |
DcPP2C16 | AGGGCAAACTTCTGGGACTACTG | AGGGCAAACTTCTGGGACTACTG |
DcPP2C20 | GTGGGAAAAAATGTGAGAAAGAGT | TTGGGTCCATCTCCAGTATTCTT |
DcPP2C24 | CCTGAGGACTCTTTAGCCATTGC | GTGATGTAAGGTGAGGTTCCAATGT |
DcPP2C29 | TTCAAGGCAGGTATGGGCGTA | TCGCATCATTCATCTGCTCCA |
DcPP2C35 | CTTCAGGAACGACAGTTACATTTG | TCCAACCTCACCCCCACTT |
DcPP2C38 | CACCTGGGAGTGACGACGAT | CCAGACATACCAAGTTCAAATCCC |
DcPP2C56 | CTTGGCAACCGTTTATTTGTAGC | CACATTACTACTCCTCCCGCATC |
DcPP2C62 | CATTGTTGAGGGGCAAGAGAG | GCACCACTCATCATCAACGC |
表1 荧光定量PCR引物
Table 1 Primer sequences of quantitative real time PCR
基因 Gene | 上游引物(5′-3′) Forward primer | 下游引物(5′-3′) Reverse primer |
---|---|---|
ACTIN | TTGTGTTGGATTCTGGTGATGGTGT | TTTCCCGTTCTGCTGTTGTTGTGAA |
DcPP2C5 | CTTTGAAGATGTTGAGTTCCCAC | TGCTTTCCGCACTGAATCCT |
DcPP2C6 | ATCACTGTTGTGGACCTTTGCG | CGCACACGGCAGAGAGGCTA |
DcPP2C16 | AGGGCAAACTTCTGGGACTACTG | AGGGCAAACTTCTGGGACTACTG |
DcPP2C20 | GTGGGAAAAAATGTGAGAAAGAGT | TTGGGTCCATCTCCAGTATTCTT |
DcPP2C24 | CCTGAGGACTCTTTAGCCATTGC | GTGATGTAAGGTGAGGTTCCAATGT |
DcPP2C29 | TTCAAGGCAGGTATGGGCGTA | TCGCATCATTCATCTGCTCCA |
DcPP2C35 | CTTCAGGAACGACAGTTACATTTG | TCCAACCTCACCCCCACTT |
DcPP2C38 | CACCTGGGAGTGACGACGAT | CCAGACATACCAAGTTCAAATCCC |
DcPP2C56 | CTTGGCAACCGTTTATTTGTAGC | CACATTACTACTCCTCCCGCATC |
DcPP2C62 | CATTGTTGAGGGGCAAGAGAG | GCACCACTCATCATCAACGC |
基因名称 Gene name | 基因 ID Gene ID | 基因位置 Genome location | 氨基酸/aa Amino acid | 分子量/kD Molecular weight | 等电点 Isoelectric point |
---|---|---|---|---|---|
DcPP2C1 | LOC110092834 | NW_021319682.1 12353861…12363554 | 429 | 46.53 | 4.79 |
DcPP2C2 | LOC110093201 | NW_021319838.1 79636…86716 | 357 | 38.54 | 5.92 |
DcPP2C3 | LOC110097041 | NW_021445045.1 22930…89839 | 376 | 41.44 | 5.73 |
DcPP2C4 | LOC110092981 | NW_021479630.1 566050…569127 | 437 | 48.44 | 5.73 |
DcPP2C5 | LOC110106882 | NW_021319908.1 1599017…1626996 | 656 | 73.43 | 6.05 |
DcPP2C6 | LOC110093401 | NW_021319500.1 2605723…2631661 | 355 | 39.03 | 5.44 |
DcPP2C7 | LOC110096102 | NW_021318918.1 327845…329678 | 349 | 38.17 | 5.57 |
DcPP2C8 | LOC110092255 | NW_021319550.1 589338…598830 | 289 | 31.62 | 6.90 |
DcPP2C9 | LOC110102146 | NW_021318673.1 106681…130525 | 283 | 31.37 | 6.13 |
DcPP2C10 | LOC110101792 | NW_021437127.1 239…864 | 169 | 18.38 | 5.19 |
DcPP2C11 | LOC110116114 | NW_021319083.1 7218367…7229170 | 363 | 39.77 | 5.22 |
DcPP2C12 | LOC110103211 | NW_021319536.1 1067495…1070936 | 342 | 37.17 | 5.22 |
DcPP2C13 | LOC110108169 | NW_021320036.1 164362…168605 | 403 | 44.84 | 6.30 |
DcPP2C14 | LOC110101987 | NW_021319486.1 1718938…1721588 | 394 | 43.43 | 4.77 |
DcPP2C15 | LOC110097073 | NW_021319809.1 112607…115681 | 404 | 44.61 | 5.01 |
DcPP2C16 | LOC110094890 | NW_021319875.1 884026…895795 | 433 | 46.69 | 8.12 |
DcPP2C17 | LOC110101160 | NW_021320015.1 317054…320264 | 463 | 51.24 | 5.47 |
DcPP2C18 | LOC110103650 | NW_021319857.1 597472…599426 | 521 | 58.42 | 5.78 |
DcPP2C19 | LOC110107612 | NW_021319184.1 52038…60613 | 536 | 57.64 | 4.97 |
DcPP2C20 | LOC110092282 | NW_021319843.1 1140274…1141088 | 377 | 40.61 | 5.67 |
DcPP2C21 | LOC110092686 | NW_021318775.1 1151130…1158463 | 391 | 43.31 | 6.47 |
DcPP2C22 | LOC110100546 | NW_021542761.1 641641…655285 | 306 | 33.64 | 6.32 |
DcPP2C23 | LOC110116041 | NW_021369505.1 23106…26698 | 217 | 23.13 | 5.31 |
DcPP2C24 | LOC110105480 | NW_021319083.1 17250298…17253552 | 645 | 71.49 | 5.58 |
DcPP2C25 | LOC110093956 | NW_021319004.1 5426598…5430388 | 597 | 65.52 | 5.66 |
DcPP2C26 | LOC110111364 | NW_021319303.1 345392…366805 | 432 | 46.00 | 5.27 |
DcPP2C27 | LOC110095027 | NW_021320057.1 2721345…2737862 | 399 | 43.97 | 8.90 |
DcPP2C28 | LOC110116281 | NW_021319063.1 1121923…1141845 | 376 | 42.47 | 6.60 |
DcPP2C29 | LOC110111869 | NW_021319891.1 154900…158247 | 387 | 42.31 | 5.57 |
DcPP2C30 | LOC110111777 | NW_021319083.1 28116466…28125329 | 285 | 31.08 | 6.18 |
DcPP2C31 | LOC110102478 | NW_021319127.1 3797515…3799946 | 408 | 44.11 | 5.81 |
DcPP2C32 | LOC110110966 | NW_021318758.1 3751…4653 | 270 | 29.36 | 5.99 |
DcPP2C33 | LOC110092340 | NW_021319405.1 21754…29592 | 613 | 67.67 | 5.92 |
DcPP2C34 | LOC110098931 | NW_021319048.1 1082370…1091816 | 440 | 47.25 | 5.98 |
DcPP2C35 | LOC110114396 | NW_021319770.1 96534…111295 | 445 | 48.21 | 6.28 |
DcPP2C36 | LOC110115466 | NW_021319945.1 1826754…1859103 | 438 | 47.35 | 8.10 |
DcPP2C37 | LOC110101385 | NW_021319178.1 7021686…7023466 | 408 | 44.08 | 6.47 |
DcPP2C38 | LOC110109810 | NW_021319281.1 570006…587525 | 564 | 60.50 | 4.75 |
DcPP2C39 | LOC110102541 | NW_021319394.1 554440…558932 | 366 | 40.43 | 8.14 |
DcPP2C40 | LOC110113698 | NW_021319086.1 1277580…1305129 | 843 | 93.73 | 5.15 |
DcPP2C41 | LOC110110144 | NW_021319483.1 1927209…1943550 | 294 | 31.93 | 4.82 |
DcPP2C42 | LOC110113512 | NW_021320183.1 343309…347253 | 405 | 44.74 | 9.13 |
DcPP2C43 | LOC110109375 | NW_021318890.1 6027…43817 | 369 | 40.74 | 6.73 |
DcPP2C44 | LOC110114968 | NW_021540301.1 77…542 | 80 | 8.86 | 4.13 |
DcPP2C45 | LOC110103345 | NW_021319127.1 148186…155682 | 379 | 41.74 | 8.38 |
DcPP2C46 | LOC110099265 | NW_021319673.1 21670…23848 | 386 | 42.73 | 5.75 |
DcPP2C47 | LOC110095534 | NW_021319289.1 235158…239974 | 361 | 40.80 | 6.16 |
DcPP2C48 | LOC110096184 | NW_021320128.1 576030…582730 | 382 | 43.03 | 6.90 |
DcPP2C49 | LOC110102425 | NW_021319836.1 605740…611552 | 364 | 40.36 | 8.09 |
DcPP2C50 | LOC110094144 | NW_021319833.1 29275…42783 | 332 | 36.68 | 5.63 |
DcPP2C51 | LOC110098988 | NW_021387661.1 202…910 | 136 | 14.80 | 6.13 |
DcPP2C52 | LOC110104643 | NW_021319518.1 7586364…7608658 | 963 | 106.49 | 5.56 |
DcPP2C53 | LOC110107952 | NW_021318744.1 279285…290373 | 956 | 106.29 | 6.10 |
DcPP2C54 | LOC110101653 | NW_021319798.1 157820…165707 | 351 | 38.27 | 5.27 |
基因名称 Gene name | 基因 ID Gene ID | 基因位置 Genome location | 氨基酸/aa Amino acid | 分子量/kD Molecular weight | 等电点 Isoelectric point |
DcPP2C55 | LOC110104152 | NW_021320112.1 176838…213807 | 1 075 | 119.59 | 5.12 |
DcPP2C56 | LOC110094641 | NW_021319416.1 6703…36842 | 322 | 36.03 | 8.71 |
DcPP2C57 | LOC110113113 | NW_021319434.1 271374…304456 | 356 | 39.15 | 5.21 |
DcPP2C58 | LOC110105369 | NW_021319475.1 45167…49612 | 285 | 31.31 | 8.28 |
DcPP2C59 | LOC110102674 | NW_021318663.1 311998…323025 | 216 | 23.05 | 5.94 |
DcPP2C60 | LOC110106056 | NW_021319959.1 659303…671684 | 392 | 43.74 | 7.70 |
DcPP2C61 | LOC110108569 | NW_021319385.1 917516…937291 | 480 | 52.41 | 5.89 |
DcPP2C62 | LOC110099044 | NW_021320094.1 1386063…1409016 | 512 | 56.12 | 5.32 |
DcPP2C63 | LOC110092171 | NW_021319127.1 3301272…3324232 | 625 | 69.12 | 6.31 |
DcPP2C64 | LOC110098867 | NW_021319985.1 222186…229368 | 384 | 42.55 | 9.36 |
DcPP2C65 | LOC110108709 | NW_021318852.1 2065548…2077099 | 289 | 30.98 | 4.77 |
DcPP2C66 | LOC110112047 | NW_021318824.1 229329…231733 | 340 | 37.92 | 7.03 |
DcPP2C67 | LOC110115916 | NW_021319683.1 571454…602635 | 380 | 41.94 | 6.16 |
表2 铁皮石斛PP2C家族成员理化性质
Table 2 Physical and chemical properties of PP2C gene family in Dendrobium catenatum
基因名称 Gene name | 基因 ID Gene ID | 基因位置 Genome location | 氨基酸/aa Amino acid | 分子量/kD Molecular weight | 等电点 Isoelectric point |
---|---|---|---|---|---|
DcPP2C1 | LOC110092834 | NW_021319682.1 12353861…12363554 | 429 | 46.53 | 4.79 |
DcPP2C2 | LOC110093201 | NW_021319838.1 79636…86716 | 357 | 38.54 | 5.92 |
DcPP2C3 | LOC110097041 | NW_021445045.1 22930…89839 | 376 | 41.44 | 5.73 |
DcPP2C4 | LOC110092981 | NW_021479630.1 566050…569127 | 437 | 48.44 | 5.73 |
DcPP2C5 | LOC110106882 | NW_021319908.1 1599017…1626996 | 656 | 73.43 | 6.05 |
DcPP2C6 | LOC110093401 | NW_021319500.1 2605723…2631661 | 355 | 39.03 | 5.44 |
DcPP2C7 | LOC110096102 | NW_021318918.1 327845…329678 | 349 | 38.17 | 5.57 |
DcPP2C8 | LOC110092255 | NW_021319550.1 589338…598830 | 289 | 31.62 | 6.90 |
DcPP2C9 | LOC110102146 | NW_021318673.1 106681…130525 | 283 | 31.37 | 6.13 |
DcPP2C10 | LOC110101792 | NW_021437127.1 239…864 | 169 | 18.38 | 5.19 |
DcPP2C11 | LOC110116114 | NW_021319083.1 7218367…7229170 | 363 | 39.77 | 5.22 |
DcPP2C12 | LOC110103211 | NW_021319536.1 1067495…1070936 | 342 | 37.17 | 5.22 |
DcPP2C13 | LOC110108169 | NW_021320036.1 164362…168605 | 403 | 44.84 | 6.30 |
DcPP2C14 | LOC110101987 | NW_021319486.1 1718938…1721588 | 394 | 43.43 | 4.77 |
DcPP2C15 | LOC110097073 | NW_021319809.1 112607…115681 | 404 | 44.61 | 5.01 |
DcPP2C16 | LOC110094890 | NW_021319875.1 884026…895795 | 433 | 46.69 | 8.12 |
DcPP2C17 | LOC110101160 | NW_021320015.1 317054…320264 | 463 | 51.24 | 5.47 |
DcPP2C18 | LOC110103650 | NW_021319857.1 597472…599426 | 521 | 58.42 | 5.78 |
DcPP2C19 | LOC110107612 | NW_021319184.1 52038…60613 | 536 | 57.64 | 4.97 |
DcPP2C20 | LOC110092282 | NW_021319843.1 1140274…1141088 | 377 | 40.61 | 5.67 |
DcPP2C21 | LOC110092686 | NW_021318775.1 1151130…1158463 | 391 | 43.31 | 6.47 |
DcPP2C22 | LOC110100546 | NW_021542761.1 641641…655285 | 306 | 33.64 | 6.32 |
DcPP2C23 | LOC110116041 | NW_021369505.1 23106…26698 | 217 | 23.13 | 5.31 |
DcPP2C24 | LOC110105480 | NW_021319083.1 17250298…17253552 | 645 | 71.49 | 5.58 |
DcPP2C25 | LOC110093956 | NW_021319004.1 5426598…5430388 | 597 | 65.52 | 5.66 |
DcPP2C26 | LOC110111364 | NW_021319303.1 345392…366805 | 432 | 46.00 | 5.27 |
DcPP2C27 | LOC110095027 | NW_021320057.1 2721345…2737862 | 399 | 43.97 | 8.90 |
DcPP2C28 | LOC110116281 | NW_021319063.1 1121923…1141845 | 376 | 42.47 | 6.60 |
DcPP2C29 | LOC110111869 | NW_021319891.1 154900…158247 | 387 | 42.31 | 5.57 |
DcPP2C30 | LOC110111777 | NW_021319083.1 28116466…28125329 | 285 | 31.08 | 6.18 |
DcPP2C31 | LOC110102478 | NW_021319127.1 3797515…3799946 | 408 | 44.11 | 5.81 |
DcPP2C32 | LOC110110966 | NW_021318758.1 3751…4653 | 270 | 29.36 | 5.99 |
DcPP2C33 | LOC110092340 | NW_021319405.1 21754…29592 | 613 | 67.67 | 5.92 |
DcPP2C34 | LOC110098931 | NW_021319048.1 1082370…1091816 | 440 | 47.25 | 5.98 |
DcPP2C35 | LOC110114396 | NW_021319770.1 96534…111295 | 445 | 48.21 | 6.28 |
DcPP2C36 | LOC110115466 | NW_021319945.1 1826754…1859103 | 438 | 47.35 | 8.10 |
DcPP2C37 | LOC110101385 | NW_021319178.1 7021686…7023466 | 408 | 44.08 | 6.47 |
DcPP2C38 | LOC110109810 | NW_021319281.1 570006…587525 | 564 | 60.50 | 4.75 |
DcPP2C39 | LOC110102541 | NW_021319394.1 554440…558932 | 366 | 40.43 | 8.14 |
DcPP2C40 | LOC110113698 | NW_021319086.1 1277580…1305129 | 843 | 93.73 | 5.15 |
DcPP2C41 | LOC110110144 | NW_021319483.1 1927209…1943550 | 294 | 31.93 | 4.82 |
DcPP2C42 | LOC110113512 | NW_021320183.1 343309…347253 | 405 | 44.74 | 9.13 |
DcPP2C43 | LOC110109375 | NW_021318890.1 6027…43817 | 369 | 40.74 | 6.73 |
DcPP2C44 | LOC110114968 | NW_021540301.1 77…542 | 80 | 8.86 | 4.13 |
DcPP2C45 | LOC110103345 | NW_021319127.1 148186…155682 | 379 | 41.74 | 8.38 |
DcPP2C46 | LOC110099265 | NW_021319673.1 21670…23848 | 386 | 42.73 | 5.75 |
DcPP2C47 | LOC110095534 | NW_021319289.1 235158…239974 | 361 | 40.80 | 6.16 |
DcPP2C48 | LOC110096184 | NW_021320128.1 576030…582730 | 382 | 43.03 | 6.90 |
DcPP2C49 | LOC110102425 | NW_021319836.1 605740…611552 | 364 | 40.36 | 8.09 |
DcPP2C50 | LOC110094144 | NW_021319833.1 29275…42783 | 332 | 36.68 | 5.63 |
DcPP2C51 | LOC110098988 | NW_021387661.1 202…910 | 136 | 14.80 | 6.13 |
DcPP2C52 | LOC110104643 | NW_021319518.1 7586364…7608658 | 963 | 106.49 | 5.56 |
DcPP2C53 | LOC110107952 | NW_021318744.1 279285…290373 | 956 | 106.29 | 6.10 |
DcPP2C54 | LOC110101653 | NW_021319798.1 157820…165707 | 351 | 38.27 | 5.27 |
基因名称 Gene name | 基因 ID Gene ID | 基因位置 Genome location | 氨基酸/aa Amino acid | 分子量/kD Molecular weight | 等电点 Isoelectric point |
DcPP2C55 | LOC110104152 | NW_021320112.1 176838…213807 | 1 075 | 119.59 | 5.12 |
DcPP2C56 | LOC110094641 | NW_021319416.1 6703…36842 | 322 | 36.03 | 8.71 |
DcPP2C57 | LOC110113113 | NW_021319434.1 271374…304456 | 356 | 39.15 | 5.21 |
DcPP2C58 | LOC110105369 | NW_021319475.1 45167…49612 | 285 | 31.31 | 8.28 |
DcPP2C59 | LOC110102674 | NW_021318663.1 311998…323025 | 216 | 23.05 | 5.94 |
DcPP2C60 | LOC110106056 | NW_021319959.1 659303…671684 | 392 | 43.74 | 7.70 |
DcPP2C61 | LOC110108569 | NW_021319385.1 917516…937291 | 480 | 52.41 | 5.89 |
DcPP2C62 | LOC110099044 | NW_021320094.1 1386063…1409016 | 512 | 56.12 | 5.32 |
DcPP2C63 | LOC110092171 | NW_021319127.1 3301272…3324232 | 625 | 69.12 | 6.31 |
DcPP2C64 | LOC110098867 | NW_021319985.1 222186…229368 | 384 | 42.55 | 9.36 |
DcPP2C65 | LOC110108709 | NW_021318852.1 2065548…2077099 | 289 | 30.98 | 4.77 |
DcPP2C66 | LOC110112047 | NW_021318824.1 229329…231733 | 340 | 37.92 | 7.03 |
DcPP2C67 | LOC110115916 | NW_021319683.1 571454…602635 | 380 | 41.94 | 6.16 |
图3 铁皮石斛PP2C基因在不同组织的表达分析 LE:叶片;ST:茎;CA:蒴果;SE:花萼;Gy:合蕊柱;RO:根。
Fig. 3 Relative expression level of DcPP2C genes in different tissues LE:Leaf;ST:Stem;CA:Capsule;SE:Sepal;Gy:Gynostemium;RO:Root.
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