陆地棉GAI基因原核表达与多克隆抗体制备  

Expression of a GAI Gene of Cotton (Gossypium hirsutum) and Preparation of Its Antibody

作　　者：郑银英[1,2,3] 崔百明[2,3] 祝建波[3] 廖文斌[2] 张根发[4] 彭明[1,2,3] 

机构地区：[1]海南大学,海口570228 [2]中国热带农业科学院热带生物技术研究所,海口571101 [3]石河子大学生命科学学院,农业生物技术重点实验室,石河子832003 [4]北京师范大学生命科学学院,北京100875

出　　处：《棉花学报》2009年第6期520-523,共4页Cotton Science

基　　金：国家973计划资助项目(2004CB117302);国家自然科学基金资助项目(30760099)

摘　　要：DELLA蛋白是GA信号响应的关键负调节因子,本文采用基于EST的电子克隆方法,从棉花中克隆了DELLA蛋白家族的一个成员GhGAI。根据电子克隆序列,从陆地棉花胚珠cDNA中扩增得到GhGAI全长基因片段。将其在原核中表达,得到了分子量(Mr)为62000的蛋白质条带。表达蛋白经His-Tag亲和层析纯化后,对兔子进行免疫,制备的抗血清通过间接ELISA检测,具有较高的效价和特异性。DELLA protein is the key negative regulator of GA signal response. In order to study the effect of GA on development of cotton fiber, we cloned GhGAI gene, a member of DELLA gene family from the upland cotton EST database by electronic cloning method based on EST. According to the electronic cloning sequence, an integrate coding region was amplified from cotton（Gossypium hirsuturn） ovule cDNA. A protein band with molecular weight （Mr） of 62000 was obtained from the prokaryotic expression products by SDS-PAGE. The protein was then purified by His-tag affinity chromatography and used for mutiple-cloned antibody preparation by rabbit immunization. Indirect ELISA test and the Western blot analysis showed that the anti-serum was in high immunocompetence and high specificity.

关 键 词：陆地棉 GhGAI基因 多克隆抗体 蛋白质印迹检测 

分 类 号：S562.035[农业科学—作物学] Q785[生物学—分子生物学]