羊布鲁菌16M VjbR蛋白酵母双杂交诱饵表达载体的构建及其毒性和自激活效应检测  

Construction of a bait plasmid containing Brucella melitensis 16M VjbR gene in a yeast two-hybrid system and evaluation of its toxicity and self-activation

作　　者：屈海龙[1] 王景龙[1] 李晓燕[1] 张瑞[1] 王秀然[1] 陈思[1] 钱晶[1] 王兴龙[1] 

机构地区：[1]军事医学科学院军事兽医研究所吉林省人兽共患病预防与控制重点实验室,吉林长春130122

出　　处：《中国兽医学报》2012年第6期852-856,共5页Chinese Journal of Veterinary Science

基　　金：国家自然科学基金资助项目(30972198/C180503);国家科技支撑计划资助项目(2010BAD04B03)

摘　　要：利用Sos招募系统(SRS),通过聚合酶链反应(PCR)扩增羊布鲁菌16M VjbR基因的编码序列,定向克隆到酵母表达载体pSos中,构建诱饵重组质粒pSos-VjbR,经测序正确后其将转入酵母菌cdc25H(α)感受态细胞,检测其表达产物对酵母细胞有无毒性作用及对报告基因有无自激活作用。结果表明,经序列测定证实重组诱饵质粒pSos-VjbR构建成功。重组质粒转化入酵母细胞后,经检测其表达产物对cdc25H(α)酵母细胞无毒性作用,对报告基因亦无自激活作用。这为利用SRS来研究与羊布鲁菌16MVjbR蛋白相互作用的蛋白奠定了基础。To construct a yeast expression vector of Brucella melitensis 16M VjbR gene using the Sos-recruitment system （SRS）, the coding sequence of Brucella rnelitensis 16M VjbR was amplified by PCR and cloned into the yeast expression plasmid pSos. The recombinant bait plasmid pSos-VjbR was verified by sequencing before transformation into competent yeast cells. The effects of the expression product on the yeast cell growth and activation of the report- er gene were evaluated. The results show the yeast expression vector of Brucella melitensis 16M VjbR gene was con- structed sucessfully. The recombinant bait plasmid showed no toxic effect on yeast cdc25H cells without a self-acti vation of the reporter gene. The SRS can be used to study the proteins interacting with Brucella melitensis 16M VjbR protein and provides a means for obtaining insight into the pathogenic mechanism of Brucella melitensis 16M.

关 键 词：vjbR蛋白 羊布鲁茵16M 自激活作用 酵母双杂交 

分 类 号：S852.614[农业科学—基础兽医学]