小麦TaCP3基因的克隆及其参与非生物胁迫的表达分析  

Cloning of TaCP3 Gene and Its expression in Abiotic Stress

作　　者：张炳慧 高翔[1,2] 杨明明 王宪国[1] 何庆梦[1] 姚晓露 李晓燕 赵万春[1,2] 董剑[1,2] ZHANG Binghui;GAO Xiang;YANG Mingming;WANG Xianguo;HE Qingmeng;YAO Xiaolu;LI Xiaoyan;ZHAO Wanchun;DONG Jian(College of Agronomy,Northwest A&F University,Yangling,Shaanxi 712100,China;Wheat Engineering Research Center of Shaanxi Province/Engineering Research Centre Wheat of New Varieties Cultivation of Wheat,Yangling,Shaanxi 712100,China)

机构地区：[1]西北农林科技大学农学院,陕西杨凌712100 [2]陕西省小麦工程技术研究中心/陕西省小麦新品种培育工程研究中心,陕西杨凌712100

出　　处：《麦类作物学报》2019年第5期513-519,共7页Journal of Triticeae Crops

基　　金：“十二五”农村领域国家科技计划项目(2011AA100501);国家现代农业产业技术体系建设专项(CARS-3-2-47);国家自然科学基金项目(31801443);中国博士后基金项目(2016M602871);中央高校基本科研业务费资助项目(Z109021623);作物遗传种质创新国家重点实验室开放课题(ZW201702)

摘　　要：半胱氨酸蛋白酶(cysteine protease,CP)是植物中重要的蛋白酶家族之一,广泛参与植物的各种生理过程;TaCP3属于papain-like(木瓜蛋白酶)家族的半胱氨酸蛋白酶,在小麦的生长发育及抗逆中起到重要作用。为研究TaCP3基因的结构特征,以及在干旱、高盐、低温和高温胁迫下的表达情况,从小麦抗旱品种西农538中克隆了TaCP3基因,该基因仅含1个1125bp的开放阅读框,编码374个氨基酸,在蛋白氨基端有一个28个氨基酸残基组成的信号肽,羧基端具有木瓜蛋白酶亚家族的保守结构域。生物信息学分析表明,TaCP3与大麦和山羊草半胱氨酸蛋白酶基因相似性最高。同时,本研究构建了pcold-TF/TaCP3原核表达载体,通过转化大肠杆菌BL21(DE3),成功表达了TaCP3重组蛋白,分子量约为40kD。qRT-PCR结果表明,TaCP3的表达对干旱、高盐、低温和高温胁迫均有响应,初始都呈先降后升的趋势;且对干旱胁迫有强烈的正向响应。Cysteine protease(CP) is one of the most important protease families in plants.It is widely involved in various physiological processes of plants.TaCP3 belongs to the papain-like family of the cysteine protease.It also plays an important role in wheat growth and development and stress resistance.In order to study the structural characteristics of TaCP3 gene and its expression under drought stress,high salt stress,low and high temperature stresses,the TaCP3 gene was cloned from the common wheat variety Xinong 538.The gene contains only a 1 125 bp open reading frame and encodes a protein containing 374 amino acids.There is a signal peptide consisting of 28 amino acid residues at the N-terminus of the protein,and the C-terminus has a conserved domain of the papain subfamily.Bioinformatics analysis showed that TaCP3 had the highest similarity with cysteine protease genes in barley and Aegilops.We constructed the pcold-TF/TaCP3 prokaryotic expression vector and successfully expressed TaCP3 recombinant protein by transforming E.coli BL21(DE3) with molecular weight of approximately 40 kD.The results of qRT-PCR showed that the expression of TaCP3 was all responded to drought stress,high salt stress,low and high temperature stresses,and the initial changes showed a trend of 'first decrease and then increase'.There was a strong positive response under drought stress.

关 键 词：小麦 TaCP3 基因克隆 原核表达 非生物胁迫 

分 类 号：S512.1[农业科学—作物学] S330