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作物学报 ›› 2019, Vol. 45 ›› Issue (2): 267-275.doi: 10.3724/SP.J.1006.2019.84075

• 耕作栽培·生理生化 • 上一篇    下一篇

青海高原耐旱蚕豆品种青海13号响应干旱胁迫蛋白质组学分析

李萍1,2,侯万伟1,2,刘玉皎1,2,*()   

  1. 1 青海大学农林科学院, 青海西宁 810000
    2青海大学省部共建三江源生态与高原农牧业国家重点实验室, 青海西宁 810000
  • 收稿日期:2018-05-29 接受日期:2018-10-08 出版日期:2019-02-12 网络出版日期:2018-11-03
  • 通讯作者: 刘玉皎
  • 基金资助:
    本研究由国家自然科学基金项目(31460377);国家现代农业产业技术体系建设专项资助(CARS-09)

Proteomic analysis of drought stress response on drought resistance for Vicia faba L. variety ‘Qinghai 13’ in Qinghai Plateau of China

Ping LI1,2,Wan-Wei HOU1,2,Yu-Jiao LIU1,2,*()   

  1. 1 Academy of Agriculture and Forestry Science of Qinghai University, Xining 810016, Qinghai, China
    2 State Key Laboratory of Plateau Ecology and Agriculture, Qinghai University, Xining 810016, Qinghai, China
  • Received:2018-05-29 Accepted:2018-10-08 Published:2019-02-12 Published online:2018-11-03
  • Contact: Yu-Jiao LIU
  • Supported by:
    This study was supported by the National Natural Science Foundation of China(31460377);and the China Agriculture Research System(CARS-09)

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摘要：

蛋白质组学研究在功能基因组时代发挥着越来越重要的作用, 利用双向电泳技术和质谱鉴定技术, 可大量研究作物逆境胁迫后蛋白质组的变化, 增加作物响应干旱胁迫机制的认识和理解。为探讨一种抗旱性蚕豆青海13号品种的耐旱机制, 本研究对其幼苗期进行干旱胁迫处理, 应用上述技术进行差异蛋白质组分析, 经t检验发现32个差异表达蛋白点, 部分呈现上调表达, 部分呈现下调表达, 还有7个消失蛋白点和1个新增蛋白点。采用MALDI-TOF/TOF鉴定和生物信息学分析发现, 成功鉴定的21个蛋白点按其所参与的代谢途径和生化功能可分为七大类, 参与信号转导的2个, 参与自由基清除的1个, 参与防卫反应的1个, 参与代谢的8个, 参与蛋白加工的1个, 参与光合的5个, 未知功能蛋白3个。22 kD干旱诱导蛋白、应激诱导蛋白、17.5 kD一级热激蛋白、超氧化物歧化酶是与抗旱性有直接关联的蛋白点, 其相对表达量的上调可能是青海13号蚕豆具有较强抗旱性的重要原因。

关键词: 蚕豆, 干旱胁迫, 蛋白质组, 差异表达, 功能分类

Abstract:

Proteomics is playing an increasingly important role in the functional genomics era. Two-dimensional electrophoresis and mass spectrometry can be used to study the changes of proteome of crop under stress and increase the recognization and comprehension of crop response to drought stress. In order to explore the mechanism of drought resistance of Vicia faba L. variety ‘Qinghai 13’, we treated the seedlings with three days for water stress and analyzed by two-dimensional gel electrophoresis combined with mass spectrometry analysis. By t-test, 32 differentially expressed proteins spots were detected between normal and drought-stress treatments, respectively, including up- and down-regulated proteins, seven disappear protein spots and a new protein spot. Twenty-one differentially expressed proteins in seven function categories were identified and confirmed by MALDI-TOF/TOF. Among them, two participated in information transfer, one in oxygen radical scavenging, one in protective response, eight in energy metabolism, one in protein processing, five in photosynthesis, and three unknown in function. These results indicate that drought-inducible 22 kD protein, stress-inducible protein, superoxide dismutase and 17.5 kD class HSP are directly related with drought resistance, which may be the important reasons for strong drought resistance of ‘Qinghai 13’.

Key words: faba bean, drought stress, proteomic, differential expression, functional classification

图1

不同梯度干旱胁迫下土壤含水量(SWC)和蚕豆叶片的相对含水量(RWC)变化 大写字母表示不同胁迫处理间0.01水平上显著性差异, 小写字母表示0.05水平上差异显著。DS1~DS9为不同梯度干旱胁迫处理, 从DS1到DS9干旱胁迫程度逐渐加强。"

图2

干旱胁迫下蚕豆叶片蛋白质组双向电泳图谱"

表1

干旱胁迫下蚕豆叶片中差异表达蛋白的MALDI-TOF/TOF鉴定结果"

蛋白点
Spot No.		 登录号
Accession No.		 分子量/等电点
Theoretical Mr (kD)/pI		 序列覆盖率
Sequence coverage (%)		 评分
Score		 蛋白名称
Protein name		 物种来源
Source of species
A, up-regulated protein spots
2705 gi|15667623 15.923/5.78 18 230 Drought inducible 22 kD protein 甘蔗Saccharum officinarum
3106 gi|380005612 20.731/5.79 31 717 Superoxide dismutase 蚕豆Vicia faba
3208 gi|83776798 16.256/5.82 19 332 17.5 kDa class I HSP, partial 花生Arachis hypogaea
3809 gi|308810206 85.783/8.69 1 54 Shikimate dehydrogenase substrate binding,N-terminal, partial (ISS) 绿藻类Ostreococcus tauri
5605 gi|357508933 39.616/5.91 9 158 O-acetylserine (thiol) lyase 蒺藜状苜蓿Medicago truncatula
5807 gi|357481949 65.668/5.7 5 237 Stress-inducible protein, putative 蒺藜状苜蓿Medicago truncatula
7206 gi|75220301 23.908/6.16 13 212 Full=Kunitz-type trypsin inhibitor-like 2 protein; 豌豆Pisum sativum
408 gi|571556750 21.914/4.36 35 493 Nascent polypeptide-associated complex subunit alpha-like protein 1 大豆Glycine max
B, down-regulated protein spots
1815 gi|3913031 56.446/5.34 13 387 Full=1,4-alpha-D-glucan maltohydrolase 紫花苜蓿Medicago sativa
2306 gi|115788 28.692/5.47 25 411 Full=Chlorophyll a-b binding protein AB80, chloroplastic; 豌豆Pisum sativum
2309 gi|115788 28.692/5.47 25 415 Full=Chlorophyll a-b binding protein AB80, chloroplastic; 豌豆Pisum sativum
2813 gi|1045394920 86.972/5.46 13 643 Hypothetical protein TSUD_183880 三叶草Trifolium subterraneum
3815 gi|528749836 55.812/5.22 20 659 ATP synthase CF1 alpha subunit (plastid) 蚕豆Vicia faba
3816 gi|528749836 55.812/5.22 20 760 ATP synthase CF1 alpha subunit (plastid) 蚕豆Vicia faba
4707 gi|75308025 43.565/5.5 17 323 Full=S-adenosylmethionine synthase 2; Short=AdoMet synthase 2; 茱萸Elaeagnus umbellata
5202 gi|217071344 31.249/6.59 8 309 Unknown 蒺藜状苜蓿Medicago truncatula
5314 gi|729390274 29.386/8.65 16 426 Chlorophyll a-b binding protein 8, chloroplastic-like 醉蝶花Tarenaya hassleriana
5808 gi|357445031 80.087/6 11 480 Plastid transketolase 蒺藜状苜蓿Medicago truncatula
5809 gi|357445031 80.087/5.78 11 532 Plastid transketolase 蒺藜状苜蓿Medicago truncatula
6303 gi|502090577 37.493/7.04 25 570 Carbonic anhydrase, chloroplastic isoform X1 鹰嘴豆Cicer arietinum
7005 gi|132097 20.402/9.24 39 354 Full=Ribulose bisphosphate carboxylase small chain 3C, chloroplastic; Short=RuBisCO small subunit 3C; AltName: Full=PSS15; Flags: Precursor 豌豆Pisum sativum
7307 gi|502090577 37.493/7.04 31 723 Carbonic anhydrase, chloroplastic isoform X1 鹰嘴豆Cicer arietinum
7311 gi|502090577 37.493/7.04 31 735 Carbonic anhydrase, chloroplastic isoform X1 鹰嘴豆Cicer arietinum
7608 gi|20729 43.696/8.93 24 847 Unnamed protein product 豌豆Pisum sativum
8004 gi|132097 20.402/9.24 31 448 Full=Ribulose bisphosphate carboxylase small chain 3C, chloroplastic; Short=RuBisCO small subunit 3C; AltName: Full=PSS15; Flags: Precursor 豌豆Pisum sativum
8010 gi|502082899 19.055/9.93 20 219 Photosystem II repair protein PSB27-H1, chloroplastic 鹰嘴豆Cicer arietinum
1404 gi|502133626 37.498/7.63 29 541 Haloacid dehalogenase-like hydrolase domain-containing protein At3g48420 鹰嘴豆Cicer arietinum

图3

蚕豆叶片中鉴定的21个差异蛋白的功能分类"

图4

差异蛋白GO功能分类"

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