轮状病毒VP4基因和VP7基因原核表达载体的构建及表达  

Expression vector construction and protocaryon expression of rotavirus VP4 gene and VP7 gene

作　　者：王鹏雁[1] 陈创夫[1] 余兴龙[2] 徐兴然[2] 涂长春[2] 张高轩[1] 

机构地区：[1]石河子大学动物科技学院,新疆石河子832003 [2]解放军军需大学军事兽医研究所,吉林长春130062

出　　处：《中国兽医科技》2003年第1期5-10,共6页Chinese Journal of Veterinary Science and Technology

摘　　要：经PCR从含有VP4基因、VP7基因片段的重组质粒 pT VP4和 pT VP7中扩增VP4、VP7基因 ,连接至 pGEM 5zf(+ )。经筛选和鉴定正确后同时酶切目的基因和表达质粒 pET 2 8a(+ ) ,连接、转化受体菌 ,经PCR、酶切和序列分析证明 ,连接向位和阅读框架是正确的 ,从而构建了轮状病毒保护性抗原VP4基因、VP7基因片段的原核表达载体 pET2 8a VP4、pET2 8a VP7。重组质粒转化表达菌在IPTG诱导下 ,用SDS PAGE、薄层凝胶扫描分析表达的蛋白。结果表明 ,表达的目的蛋白以包涵体的形式存在 ,蛋白的分子量分别为 37.6 9和 36 .2 0ku ,表达量占菌体总蛋白的比例分别为 17.1%和 14 .4 %。The VP4 gene and VP7 gene were amplified from recombinant plasmids pT VP4 and pT VP7 by PCR and ligated with plasmid pGEM 5zf(+).The recombinant plasmids and expression vector pET 28a(+) were digested by restriction endonuclease.The two genes were ligated and transformed into E.coli.By PCR,RE digestion and sequencing the orientation and coding frame were correct.The result showed that the recombinant plasmids pET28a VP4?pET28a VP7were constructed. The transformed recombinant plasmids were induced by IPTG and the expressed proteins were valued by SDS PAGE with expected molecular size of 37.69 ku and 36.20 ku and the expression efficiency were respectively 17.1% and 14.4%.

关 键 词：轮状病毒 VP4基因 VP7基因 原核表达载体 载体构建 

分 类 号：S855.99[农业科学—临床兽医学]