猪肠球菌溶血素cylA基因原核表达载体的构建  

Construction of Prokryotic Expression Vector of Hemolysin cylA Gene from Swine-derived Enterococus faecalis

作　　者：刘磊[1] 王亚宾[1] 程金平[1] 王中明[1] 段志刚[1] 蔡京雷[1] 崔保安[1] 

机构地区：[1]河南农业大学牧医工程学院兽医微生物实验室,河南郑州450002

出　　处：《江西农业学报》2009年第1期95-96,99,共3页Acta Agriculturae Jiangxi

摘　　要：以猪致病性粪肠球菌染色体DNA为模板,PCR扩增溶血素cylA基因,相应酶切后,克隆到原核表达载体pET32a中,构建pET32a-cylA重组质粒,并将pET32a-cylA质粒转化入大肠杆菌BL21(DE3)。经HindⅢ、XhoI双酶切及测序鉴定,并与已发表的肠球菌溶血素cylA基因序列比较,同源性达99.8%。表明成功构建猪肠球菌溶血素cylA基因的重组表达质粒,为制备单抗、开发疫苗及其致病机制研究奠定了基础。cylA gene was amplified from the chromosomal DNA of swine - derived Enterococus faecalis by PCR. After digestion with restrictive enzymes, the PCR products were cloned into prokaryotic expression vector pET32a and then were transformed to E. coli BL21 （DE3）. The recombinant plasmid pET32a- cylA was identified by Hind Ⅲ, Xho Ⅰ restrictive endonuclease digestion and DNA sequencing. DNA sequence of the amplified fragment was compared with the published sequence of cylA gene, it showed 99.8% nucleotide homology. Results showed that cylA gene of 1239 bp was amplified from swine -derived Enterococusfaecalis and the recombinant plasimid pET32a - cylA was successfully constructed. These results provide the basis for preparation of monoclonal antibody and development of enterococcus vaccine as well as further study on pathogenetie mechanisms.

关 键 词：猪 粪肠球菌 溶血素 原核表达 

分 类 号：S855[农业科学—临床兽医学]