放线杆菌外膜蛋白酵母双杂交诱饵质粒的构建及自激活与毒性检测  

Construction of Actinobacillus pleuropneumoniae Omp bait plasmid for yeast two-hybrid system and detection of its self-activation and toxic effect

作　　者：倪宏波[1,2] 刘思国[1] 杜艳芬[1] 林靖凯[1] 刘慧芳[1] 王春来[1] 司薇[1] 常月红[1] 彭玮[1] 王聃[1] 赫明雷[1] 

机构地区：[1]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室动物细菌病研究室,黑龙江哈尔滨150001 [2]黑龙江八一农垦大学动物科技学院,黑龙江大庆163319

出　　处：《中国兽医科学》2010年第6期561-564,共4页Chinese Veterinary Science

基　　金：中国农业科学院哈尔滨兽医研究所中央级公益性科研院所基本科研业务费专项(2008-14)

摘　　要：为了筛选胸膜肺炎放线杆菌外膜蛋白在肺泡上皮细胞上的作用受体,应用酵母双杂交技术构建诱饵质粒载体pGBKT7-omp(胸膜肺炎放线杆菌外膜蛋白),从胸膜肺炎放线杆菌提取基因组DNA,用PCR方法获得omp,克隆于pGBKT7载体上,进行酶切鉴定及序列分析,并检测pGBKT7-omp在酵母双杂交系统中的自激活和毒性作用。结果表明,成功构建了诱饵质粒载体pGBKT7-omp,并证明其在酵母双杂交系统中无自激活及毒性作用。表明pGBKT7-omp可应用在酵母双杂交系统中,用于寻找猪肺泡上皮细胞cDNA文库中与omp相互作用的蛋白质。To construct the bait vector pGBKT7-omp of Matchmaker GAL4 two-hybrid system,omp fragment was amplified by PCR from A.pleuropneumoniae and cloned into pGBKT7.Self-activation and toxic action of the recombination vector pGBKT7-omp were tested.The pGBKT7-omp vector was successfully constructed and proved to be no self-activation and toxic action.The pGBKT7-omp could be applied for the screening of proteins interacting with the omp in porcine alveolar epithelial cell cDNA library in the yeast two-hybrid system.

关 键 词：胸膜肺炎放线杆菌 外膜蛋白 诱饵质粒 自激活作用 毒性作用 

分 类 号：S852.619[农业科学—基础兽医学]