黄瓜花叶病毒2b基因在大肠杆菌中的表达及其自激活特性检测  

Expression and Self-activation Analysis of Cucumber Masaic Virus 2b Gene in E.coli

作　　者：王小明[1] 王健华[2] 冯团诚[2] 张雨良[2] 吴育鹏[1] 刘志昕[2] 

机构地区：[1]海南大学环境与植物保护学院,海南儋州571737 [2]中国热带农业科学院热带生物技术研究所,海南海口571101

出　　处：《热带作物学报》2010年第7期1115-1119,共5页Chinese Journal of Tropical Crops

基　　金：国家科技支撑计划课题(2007BAD48B01);中央级公益性科研院所基本科研业务费专项(ITBBZD0754)资助

摘　　要：2b蛋白是黄瓜花叶病毒与植物寄主相互作用的一种重要多功能蛋白。用PCR方法扩增目的基因2b,经酶切、连接将其克隆至带有λcI基因的pBT载体上,构建与λcI基因融合的表达载体pBT-CMV 2b,转化大肠杆菌XL1-Blue MRF′报告菌株,用IPTG诱导表达黄瓜花叶病毒2b基因。Western blot分析结果表明,2b-λcI融合蛋白已成功表达,大小约为38 ku,与预期大小一致。pBT-CMV 2b与空质粒pTRG共转化XL1-Blue MRF′报告菌株,同时以pBT空质粒和pTRG-Gal11p共转化作为对照。结果显示,CMV 2b蛋白未产生自激活作用,从而为利用大肠杆菌双杂交系统筛选与2b蛋白相互作用的蛋白质研究奠定基础。CMV 2b protein is a very important multifunctional protein interacting with the host factor. In this paper, 2b gene was obtained by PCR amplification and was subcloned to pBT vector fused to λcI gene. DNA sequencing result showed that the recombinant plasmid was constructed successfully. And then it was transformed into XL1-Blue MRF＇ report strain, 2b-λcI fused protein was expressed with IPTG inducing and identificated by Western blot. The expression protein was about 38kDa and consistent with expected molecular weight of the fused protein, when pBT-CMV 2b and pTRG were co-transformed into XL1-Blue MRF＇report strain, self-activation of 2b was not happened. This result showed that bait plasmid pBT-CMV 2b could be used to screen interacting proteins by BacterioMatchⅡ Two-Hybrid System.

关 键 词：黄瓜花叶病毒 2b基因 自激活 大肠杆菌双杂交 

分 类 号：S432.41[农业科学—植物病理学]