BVDV-2 E2基因的原核表达及重组蛋白反应原性的研究  

Prokaryotic Expression E2 gene of Bovine Viral Diarrhea Virus Genotype 2 and Reactionogenicity of Recombinant Protein

作　　者：刘昱成[1] 孟庆玲[1,2] 乔军[1] 王国超[1] 张倩[1] 贺志昊[1] 杨海波[1] 陈创夫[1] 

机构地区：[1]石河子大学动物遗传改良与疾病控制新疆自治区重点实验室,石河子832003 [2]中国农业科学院兰州兽医研究所家畜疫病病原生物学国家重点实验室,兰州730046

出　　处：《石河子大学学报（自然科学版）》2013年第6期680-683,共4页Journal of Shihezi University(Natural Science)

基　　金：新疆兵团国际科技合作计划项目(2012BC006);国家农业公益行业专项子课题(201303037-5)

摘　　要：为了对BVDV进行诊断,根据GenBank登录的BVDV-2E2基因序列,设计特异性引物,对BVDV-2新疆分离株SW E2基因进行扩增,克隆入T载体中测序,然后亚克隆入BL21(DE3)表达载体pET-28a中,构建重组表达载体pET-28a-E2,并转化入大肠埃希氏菌中。用IPTG进行诱导重组菌,SDS-PAGE和Western blot分析表达产物。SDSPAGE分析证实表达的重组融合蛋白相对分子量为32ku;Western blot分析表明该重组蛋白可与BVDV-2多克隆抗体发生特异性血清学反应,证实表达的重组E2蛋白具有良好的反应原性,为BVDV-2诊断试剂研发奠定基础。According to BVDV-2 E2 gene sequences accessed in GenBank,a pair of specific primers was designed to amplify E2 gene of SW isolate. The amplified product was cloned into T vector and sequenced. And then E2 gene was subcloned into BL21 （DE3） expression vector pET-28a for expression. Recombinant plasmid pET-28a-E2 was transformed into E. coli, Analysis of SDS-PAGE and Western blot were conducted for expression products after IPTG induction. SD^PAGE confirmed that the re- combinant fusion protein was expressed with a relative molecular weight of 32 ku. Western blot analysis showed that the recom binant protein can specifically be reactive to anti-BVDV-2 polyclonal antibody,which confirms that the recombinant E2 protein is endowed with good reactionogenicity. The study lays a foundation for developing diagnostic reagents for BVDV-2 infection.

关 键 词：BVDV E2基因 载体构建 原核表达 

分 类 号：S852.65[农业科学—基础兽医学]