伪狂犬病病毒gB基因的原核表达及间接ELISA方法的建立  

Expression of PRV gB Gene in E.coli and Establishment of Indirect ELISA

作　　者：王雨[1] 吉艺宽[1] 程艺[1] 张宝石[1] 向柯宇 琚春梅[1] 

机构地区：[1]华南农业大学兽医学院,广东广州510642

出　　处：《动物医学进展》2015年第6期19-23,共5页Progress In Veterinary Medicine

基　　金：国家自然科学基金项目(31001074)

摘　　要：为了建立评价伪狂犬病疫苗免疫效果的方法,应用PCR方法扩增gB基因的主要抗原表位区序列,大小为768bp,用BamHⅠ和HindⅢ双酶切后克隆至pET-32a(+)载体,构建重组表达质粒pETgB768,转化大肠埃希菌BL21(DE3)后,通过IPTG进行诱导表达,经SDS-PAGE和Western blot检测融合蛋白的表达情况及其反应原性。用纯化的gB768蛋白作为包被抗原建立间接gB768-ELISA检测方法,试验结果表明,该方法具有良好的敏感性、特异性、重复性。用该方法检测128份临床送检猪血清样本,并与商品化ELISA试剂盒检测结果进行比较,符合率为93.8%。该方法的建立为猪伪狂犬病疫苗免疫效果的评估奠定了基础。To establish the assay for evaluating the efficiency of PRV vaccines, the gB gene fragment of PRV containing the main antigen region was amplified by PCR and cloned into the prokaryotic expression vector pET32a （＋） to obtain the recombinant plasmid pET-gB768. The recombinant plasmid was trans- formed into E. coli BL21 （DE3）. After induction with IPTG, target protein was identified by SDS-PAGE and Western blot. Wells of ELISA plates were coated with purified recombinant protein gB768 to establish the indirect gB768-ELISA. This assay has good sensitivity, specificity and repeatability. 128 clinical por- cine sera were detected by this ELISA. Compared to commercial ELISA Kit, the coincidence rate was 93.8%. This method laid a fundation for assessing the immune efficiency of PRV vaccines.

关 键 词：伪狂犬病病毒 GB基因 主要抗原表位区 gB768-ELISA 

分 类 号：S852.655[农业科学—基础兽医学]