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作物学报 ›› 2022, Vol. 48 ›› Issue (10): 2533-2545.doi: 10.3724/SP.J.1006.2022.14183

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

马铃薯PYL基因家族的全基因组鉴定及表达分析

贾小霞1,2(), 齐恩芳1,2, 马胜1,2, 黄伟1,2, 郑永伟1,2, 白永杰1,2, 文国宏1,2,*()   

  1. 1甘肃省农业科学院马铃薯研究所 / 甘肃省马铃薯种质资源创新工程实验室, 甘肃兰州 730070
    2国家种质资源渭源观测实验站, 甘肃渭源 748201
  • 收稿日期:2021-10-11 接受日期:2022-01-05 出版日期:2022-10-12 网络出版日期:2022-02-09
  • 通讯作者: 文国宏
  • 作者简介:第一作者联系方式: E-mail: jiaxx0601@163.com
  • 基金资助:
    甘肃省自然科学基金重点项目(21JR7RA728);甘肃省农业科学院科技创新专项(2020GAAS16);甘肃省农业科学院生物技术育种专项(2020GAAS10);国家自然科学基金项目(31560412);国家自然科学基金项目(31860401);财政部和农业农村部国家现代农业产业技术体系建设专项(CARS-09-P06)

Genome-wide identification and expression analysis of potato PYL gene family

JIA Xiao-Xia1,2(), QI En-Fang1,2, MA Sheng1,2, HUANG Wei1,2, ZHENG Yong-Wei1,2, BAI Yong-Jie1,2, WEN Guo-Hong1,2,*()   

  1. 1Potato Research Institute of Gansu Academy of Agricultural Sciences / Gansu Engineering Laboratory of Potato Germplasm resources Innovation, Lanzhou 730070, Gansu, China
    2National Germplasm Resources Agricultural Experimental Station, Weiyuan 748201, Gansu, China
  • Received:2021-10-11 Accepted:2022-01-05 Published:2022-10-12 Published online:2022-02-09
  • Contact: WEN Guo-Hong
  • Supported by:
    Key Project of Natural Science Foundation of Gansu Province(21JR7RA728);Science and Technology Support Program of Gansu Academy of Agricultural Sciences(2020GAAS16);Special Project of Biotechnology Breeding of Gansu Academy of Agricultural Sciences(2020GAAS10);National Natural Science Foundation of China(31560412);National Natural Science Foundation of China(31860401);China Agriculture Research System of MOF and MARA(CARS-09-P06)

摘要:

作为关键信号分子, 脱落酸(ABA)通过其核心信号通路PYLs-PP2Cs-SnRK2s广泛调控植物的生长发育和逆境响应过程, PYLs蛋白作为ABA信号传导的核心组件, 在ABA信号传导中发挥着不可替代的作用。为探究PYLs (PYR/PYL/RCARs) 基因在马铃薯中的进化以及表达模式, 本研究从马铃薯全基因组‘DM-v 6.1’共鉴定到17个StPYLs基因, 并对其分布、蛋白理化性质、系统进化、基因结构特征以及基因表达模式进行了分析。结果表明, 17个StPYLs基因不均匀分布在8条染色体上, 其氨基酸大小在163~231 aa之间, 等电点在4.5~8.6之间, 相对分子量在18.71~25.29 kD之间。根据基因结构和蛋白的系统发育特征, StPYL家族成员共分为3个亚组, motif 1存在于本家族所有基因中, 说明它在StPYLs的进化过程中较为保守。基因表达模式分析表明, StPYL家族成员具有明显的组织表达特异性, 且除StPYL1在外源激素(BAP、ABA和IAA)和非生物胁迫(高温、盐和干旱)下均上调表达外, 其余基因存在功能分化, 不同胁迫下的表达模式各异。本研究结果为进一步阐明StPYLs基因在马铃薯中的功能奠定了理论基础。

关键词: 马铃薯, PYL基因家族, 脱落酸, 基因表达分析

Abstract:

As a key signalling molecule, abscisic acid (ABA) extensively regulates plant growth and development, and stress response processes through its core signalling pathway PYLs-PP2Cs-SnRK2s. As a core component of ABA signalling transduction, PYLs protein plays an irreplaceable role. In order to explore the evolution and expression pattern of PYL (PYR/PYL/RCARs) genes in potato, 17 PYL genes were identified from the whole potato genome ‘DM-v 6.1’, and their distribution, protein physical and chemical properties, system evolution, structural characteristics, and the relative expression patterns were analysed. The results showed that the 17 StPYL genes were unevenly distributed on 8 chromosomes, with amino acids of 163-231 aa, isoelectric points of 4.5-8.6, and relative molecular weights of 18.71-25.29 kD. According to the gene structure and protein phylogenetic characteristics, the members of the StPYL family were divided into three subgroups. Motif 1 existed in all genes in this family, indicating that it was relatively conserved in the evolution of StPYLs. Gene expression patterns showed that the members of the StPYL family had obvious tissue expression specificity, and except for the up-regulation of StPYL1 under exogenous hormones (BAP, ABA, and IAA) and abiotic stress (high temperature, salt and drought), the rest of the StPYL genes were functionally differentiated and their relative expression patterns were different under different stresses. The results of this study lay a theoretical foundation for further elucidating the function of StPYLs gene in potato.

Key words: potato, PYL gene family, abscisic acid (ABA), gene expression analysis

表1

用于qRT-PCR表达验证的基因及其引物"

基因编号
Gene ID
正向引物
Forward primer (5′-3′)
反向引物
Reverse primer (5′-3′)
EF1α ATTGGAAACGGATATGCTCCA TCCTTACCTGAACGCCTGTCA
Soltu.DM.05G022460 TCCTCAAGAGCTGCCACG GCCGCCGACTATACTAATGC
Soltu.DM.10G022490 CTCCCGTCTCCACCGTATG CCCCACCAACAACGCTAAA
Soltu.DM.03G013340 CGTCATTAGTTTCAGCGTCATC CGTTTCTTCCTTTGTATTCCCT
Soltu.DM.10G019340 CAATCCCACCAATCACTACAGC TTTGTACGCTTGAGGGTTGTC
Soltu.DM.02G024820 ATCGAACCTGTTTATCCTCCTG AAATAGCCATCTGGCAAGTCA
Soltu.DM.02G001580 TGAAGACGATGTTTTCCCAGAC TGAAGGCAGTGAAGCGGTAT
Soltu.DM.04G027470 GCATTGCTTCCTCCTCATTACT TGAACTCTTCTGCCCTATCCC
Soltu.DM.01G033740 GTTGCCTGACGGGACATTT CATATCCTTGGACCAGCATTAA

表2

马铃薯PYL基因家族成员的序列特性"

基因名称
Gene name
基因编号
Gene ID
染色体定位
Chromosome localization
亚组分类Subgroup 氨基酸数量
Number of amino acids (aa)
等电点
Isoelectric point (pI)
相对分子量
Molecular
weight (kD)
StPYL1 Soltu.DM.01G034800 Chr.01: 74327768-74330316 I 186 6.30 20.79
StPYL2 Soltu.DM.03G013340 Chr.03: 35693624-35694662 II 201 5.95 22.16
StPYL3 Soltu.DM.03G022780 Chr.03: 47692175-47694992 I 185 5.61 20.96
StPYL4 Soltu.DM.05G022460 Chr.05: 50931814-50932956 II 208 7.15 22.89
StPYL5 Soltu.DM.06G010300 Chr.06: 31336857-31338048 II 218 6.40 23.99
StPYL6 Soltu.DM.06G018570 Chr.06: 45115651-45116827 III 213 4.98 23.72
StPYL7 Soltu.DM.08G013560 Chr.08: 39726166-39727508 III 188 6.65 21.08
StPYL8 Soltu.DM.08G014660 Chr.08: 41607872-41608584 I 183 4.93 20.17
StPYL9 Soltu.DM.08G014670 Chr.08: 41614500-41615633 I 176 4.50 19.78
StPYL10 Soltu.DM.08G014680 Chr.08: 41634425-41635105 I 163 5.32 18.71
StPYL11 Soltu.DM.08G022790 Chr.08: 52187476-52188465 III 231 5.12 25.29
StPYL12 Soltu.DM.08G028930 Chr.08: 58181068-58185195 I 189 6.75 21.57
StPYL13 Soltu.DM.09G009100 Chr.09: 10398942-10400047 II 214 8.60 23.39
StPYL14 Soltu.DM.10G019340 Chr.10: 50673639-50674710 II 207 7.10 22.82
StPYL15 Soltu.DM.10G022490 Chr.10: 54650751-54652125 II 213 6.79 23.49
StPYL16 Soltu.DM.12G004400 Chr.12: 3523014-3524941 III 190 5.12 21.24
StPYL17 Soltu.DM.12G008390 Chr.12: 7357759-7362992 I 181 5.97 20.68

图1

StPYLs的进化关系、基因结构和保守基序分析 A: StPYLs进化树; B: StPYLs基因结构; C: StPYLs中保守基序的分布。"

图2

PYL基因家族系统发育进化树 At: 拟南芥; St: 马铃薯; Bn: 甘蓝型油菜; Os: 水稻。"

图3

马铃薯PYLs基因启动子序列结合转录因子"

表3

StPYL、AtPYL、OsPYL基因进化选择压力分析"

序号
No.
基因1
Gene 1
基因2
Gene 2
非同义替换率
Ka
同义替换率
Ks
非同义替换率/同义替换率
Ka/Ks
1 AtPYL1 StPYL11 0.252113 2.46901 0.102111
2 AtPYL10 StPYL1 0.209896 2.62041 0.0801006
3 AtPYL10 StPYL3 0.203876 3.47713 0.0586336
4 AtPYL2 StPYL16 0.199507 3.38436 0.0589498
5 AtPYL2 StPYL7 0.202235 3.41170 0.0592768
6 AtPYL4 StPYL14 0.277640 3.19545 0.0868861
7 AtPYL4 StPYL15 0.251724 3.78848 0.0664447
8 AtPYL5 StPYL2 0.337324 3.20802 0.10515
9 AtPYL5 StPYL5 0.358874 3.08254 0.116422
10 AtPYL6 StPYL14 0.391640 2.85503 0.137175
11 AtPYL6 StPYL2 0.335979 3.04837 0.110216
12 AtPYL6 StPYL5 0.310522 3.18218 0.0975815
13 AtPYL7 StPYL1 0.228439 2.12794 0.107352
14 AtPYL7 StPYL12 0.214102 3.49394 0.0612782
15 AtPYL8 StPYL1 0.148691 4.52157 0.0328847
16 AtPYL8 StPYL12 0.176819 3.71724 0.0475675
17 AtPYL8 StPYL3 0.130553 4.14104 0.0315267
18 AtPYL9 StPYL1 0.163537 1.84526 0.0886251
19 AtPYR1 StPYL11 0.196102 1.34623 0.145668
20 AtPYR1 StPYL6 0.204242 3.81927 0.0534767
21 OsPYL1 StPYL11 0.347476 3.45055 0.100702
序号
No.
基因1
Gene 1
基因2
Gene 2
非同义替换率
Ka
同义替换率
Ks
非同义替换率/同义替换率
Ka/Ks
22 OsPYL12 StPYL1 0.330709 2.89213 0.114348
23 OsPYL12 StPYL12 0.285345 2.94688 0.0968295
24 OsPYL4 StPYL13 0.347064 3.44965 0.100609
25 OsPYL4 StPYL5 0.355825 3.12683 0.113797
26 OsPYL5 StPYL13 0.362471 3.22157 0.112514
27 OsPYL5 StPYL14 0.360063 3.06688 0.117404
28 OsPYL5 StPYL5 0.356826 3.18746 0.111947
29 OsPYL6 StPYL13 0.384219 3.18292 0.120713
30 OsPYL6 StPYL14 0.376110 3.09537 0.121507
31 OsPYL6 StPYL5 0.359420 3.20874 0.112013
32 OsPYL7 StPYL1 0.364590 3.24765 0.112262
33 OsPYL7 StPYL3 0.321310 3.17375 0.10124
34 OsPYL8 StPYL12 0.273630 2.71313 0.100854

图4

StPYLs基因在不同胁迫下的表达量热图"

图5

StPYLs基因在不同组织(A)和不同干旱胁迫下(B)的表达模式分析 图5-B中, T75-1、T75-2、T75-3代表75%~80%的3次生物学重复, T65-1、T65-2、T65-3代表65%~70%的3次生物学重复, T55-1、T55-2、T55-3代表55%~60%的3次生物学重复, T45-1、T45-2、T45-3代表45%~50%的3次生物学重复。"

表4

不同基因在不同干旱胁迫下的FPKM值分析"

基因名称
Gene name
处理Treatment
T75 T65 T55 T45
StPYL1 82.75±3.26 b 105.73±9.28 a 110.58±13.47 a 101.73±7.90 a
StPYL3 84.46±3.52 a 73.15±4.16 a 73.57±2.17 a 78.06±3.83 a
StPYL12 11.47±2.07 a 15.16±1.26 a 12.43±0.29 a 15.89±2.72 a
StPYL17 38.49±6.22 a 39.70±0.44 a 34.52±1.45 a 27.13±4.26 a
StPYL8 1.53±0.46 ab 1.96±0.04 a 0.99±0.19 b 0.97±0.16 b
StPYL9 2.72±0.32 a 2.91±0.33 a 3.17±0.52 a 3.07±0.09 a
StPYL10 0.46±0.09 b 0.70±0.11 a 0.53±0.15 b 0.52±0.13 b
StPYL2 72.03±3.26 a 58.73±1.45 b 28.23±2.78 c 21.69±5.01 c
StPYL4 17.93±5.37 a 14.44±1.41 a 6.24±1.51 b 5.11±2.19 b
StPYL5 25.90±2.86 a 26.34±4.12 a 17.34±3.35 b 14.46±3.55 b
StPYL13 4.07±0.43 a 2.94±0.48 ab 2.87±0.27 ab 2.54±0.45 b
基因名称
Gene name
处理Treatment
T75 T65 T55 T45
StPYL14 22.86±2.94 a 18.82±1.06 ab 15.00±1.31 bc 9.44±1.92 c
StPYL15 146.99±4.30 a 121.88±7.18 b 105.61±8.19 b 66.12±9.62 c
StPYL6 40.26±5.09 a 48.10±0.99 a 44.86±3.91 a 48.78±3.15 a
StPYL11 1.00±0.08 c 1.42±0.04 ab 1.47±0.11 a 1.10±0.14 bc
StPYL7 1.68±0.28 b 2.03±0.34 a 1.12±0.20 b 1.14±0.28 b
StPYL16 0 0 0 0

图6

RNA-seq数据的qRT-PCR验证 A: 干旱胁迫下差异表达基因在T75 vs T45 和T75 vs T55组中的相对表达量分析, qRT-PCR的数据是3次重复的平均值, 误差条代表标准误。B: qRT-PCR和RNA-seq间的相关性。"

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