长武134小麦GADPH介导抗旱的调控途径

Water stress is one of important factors to decrease the products of wheat. To solve this question, many researchers were devoted to discovered the pathway of drought resistance in wheat. In drought resistance wheat Chanwu 134, the applicant has found a 39.5KDa drought inducible protein which is a member of 3-glyceraldehyde phosphate dehydrogenase (GAPDH) family. Therefore, the drought-regulated mechanism about GAPDH will be studied as follows: firstly, the dynamic expression profile of GAPDH will be detected using western blot method. Secondly, the promoters of GAPDH will be cloned from different drougt resistance wheats. Then their drought-responsive elements will be identified through element lack method. At last, according to the N-terminal amino acid sequence, the full length cDNA of GAPDH will be cloned. And then it will be transformed into Arabidopsis and water sensitive wheat for validating the drought resistance founction of GAPDH protein.this project will might to discovered a new drought resistance pathway in wheat. And it can also provide a new gene resource for cultivating drought resistance wheat through genetic engeneering techniques.

干旱是影响小麦产量的重要逆境因子，研究小麦的抗旱途径和机制具有重要意义。申请人在前期工作中从抗旱小麦品种长武134中发现39.5kDa的干旱诱导特异表达蛋白3-磷酸甘油醛脱氢酶（GAPDH）。在本项目中，将从三方面来探索GAPDH参与调控小麦抗旱的分子机理。1、将利用已制备的GAPDH抗体，通过Western blot技术确定GAPDH在干旱条件下的时空表达特征；2、从不同抗性小麦中克隆GAPDH启动子，通过元件缺失法，将缺失不同元件的GAPDH启动子与GUS基因融合后转化拟南芥，进而研究了解哪些元件具有干旱应答功能；3、根据已测定的目标蛋白N-端氨基酸序列,进一步克隆其基因全长cDNA序列,并将目的基因导入拟南芥和不抗旱小麦体内，确定该基因在抗旱中的作用。通过本项目，有望揭示一个全新的GAPDH介导的小麦抗旱调控途径，为利用基因工程方法获得转基因抗旱小麦品种提供新的基因资源和理论基础。

干旱等非生物胁迫是限制农作物生长的重要环境因素。在干旱等非生物逆境下，一些功能基因的诱导表达在植物响应非生物胁迫中至关重要。近年研究表明，GAPDH (glyceraldehyde-3-phosphate dehydrogenase)不仅是参与糖酵解的关键酶，而且在植物响应非生物胁迫中发挥重要作用。本课题以重要的粮食作物小麦入手，研究小麦中GAPDH基因对干旱等非生物胁迫的响应。结果表明，小麦中的GAPDH基因在干旱、低温胁迫下其mRNA和蛋白水平的表达量均显著升高；在转基因拟南芥和转基因小麦中，GAPDH基因的过量表达可有效提高拟南芥和水敏感小麦郑引1号的抗旱性，GAPDH基因的有效沉默会显著降低拟南芥和抗旱性小麦长武134的抗旱能力；进一步研究发现，GAPDH基因启动子中存在许多响应非生物胁迫的顺式作用元件，并且在非生物胁迫下，启动子区富含胞嘧啶序列的甲基化修饰可调节该基因的表达；此外，经生物信息学分析发现，小麦中存在13个GAPDH基因。以上数据表明，GAPDH基因在小麦响应干旱中发挥重要作用，并初步表明该基因的表达可能是通过其上游启动子序列上响应非生物胁迫的顺式作用元件及其甲基化修饰来调节的。因此，通过本项目的完成，我们初步揭示了GAPDH 基因介导干旱的调控途径。