胸膜肺炎放线杆菌体内表达基因的筛选  

Screening on the expression genes of Actinobacillus pleuropneumoniae in vivo

作　　者：刘慧芳[1] 刘思国[1] 王春来[1] 司微[1] 王聃[1] 杜艳芬[1] 赵海玲[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

出　　处：《中国兽医科学》2008年第10期847-850,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专项资金项目(2008-14)

摘　　要：利用体内诱导抗原技术对胸膜肺炎放线杆菌（APP）体内表达的基因进行了初步筛选。将收集到的5份感染了APP1型的猪血清混合．分别用体外培养的APP1和大肠杆菌BL21（DE3）的全菌及全菌裂解物进行吸附。用ELISA方法检测吸附效果，经吸附后血清D150nm分别由原来的0．927和0．239降低为0．196和0．078。同时,将APP1基因组DNA以Bsp143I进行酶切，回收500～2000bp的片段，与pET30a／b／c载体连接，构建了APP1基因组质粒表达文库，库容量（CFU）为2．0×10^5。用吸附后的血清通过菌落原位免疫杂交筛选该基因组表达文库，从3000个菌落中获得了12个阳性克隆。To screen the expression genes of Actinobacillus pleuropneumoniae （APP） in vivo by the technology of inducing antigen technology（IVIAT）,sera from 5 pigs with APP1 infection were first pooled and adsorbed with APPl,Escherichia coli BL21 （DE3）,and its lysates. To detect effect of absorption, D150nm, values of the pooled sera adsorbed successively with in vitro cultured APP1 or E. coli BL21 （DE3） were decreased from 0. 927 and 0. 239 to 0. 196 and 0. 078,respectively,by ELISA. The DNA fragments of 500 bp to 2 000 bp from the genomic DNA of APP1 digested with Bsp143 I, and ligated into the prokaryotic expression vectors pET30a/b/e to construct genornic expression library, and the constructed expression library（CFU） was 2. 0 × 10^5 in size. The genomic expression library was probed used the adsorbed-pooled sera,and 12 positive clones were obtained from 3 000 postulating colonies by immunological hybridization.

关 键 词：胸膜肺炎放线杆菌 体内诱导抗原技术 菌落原位免疫杂交 

分 类 号：S852.619[农业科学—基础兽医学]