水疱性口炎病毒糖蛋白的原核表达及间接ELISA诊断技术的试验  

Development of an Indirect ELISA for the Detection of Antibodies Against VSV Based on G Protein Prokaryotic Expression

作　　者：郭金玉[1] 张鹤晓[2] 高志强[2] 林祥超 臧京帅[1] 王麒文 牛建蕊 张乐萃[1] 

机构地区：[1]青岛农业大学动物科技学院,山东青岛266109 [2]北京出入境检验检疫局,北京朝阳100026 [3]北京森康生物技术开发有限公司,北京怀柔101400

出　　处：《中国兽医杂志》2015年第7期75-78,共4页Chinese Journal of Veterinary Medicine

基　　金：青岛农业大学研究生创新计划资助项目(QYC201302)

摘　　要：本研究利用重组质粒p ET-32a-NJ-G666(针对新泽西型水疱性口炎病毒截断的G蛋白基因片段约666 bp),将其转化至Rosetta宿主菌,用0.6 mmol/L IPTG在30℃诱导4 h,经10%聚丙烯酰胺凝胶电泳显示,表达的重组蛋白分子质量与预期的30 k Da相符。在此基础上,利用纯化的重组蛋白作为抗原包被酶标板,建立了可检测新泽西型VSV抗体的间接ELISA方法。该方法检测东部马脑脊髓炎和马西尼罗病毒的阳性血清均为阴性,且板内和板间变异系数均小于10%。试验结果表明,建立的间接ELISA方法特异性强、重复性好,为新泽西型VSV血清抗体的检测提供了依据。Vesicular stomatitis is a viral disease caused by vesicular stomatitis virus（VSV）.Indirect ELISA can be used effec-tively for the detection of VSV antibodies.In this study,the recombinant plasmid p ET-32a-NJ-G666（truncated 666 bp G proteingene fragment of VSV New Jersey type）was transformed into host bacteria Rosetta,induced 4 h at 30 ℃ by 0.6 mmol/L IPTG.10%SDS PAGE showed that molecular weight of the recombinant protein was consistent with the expected 30 k Da.Then,we establishedan indirect ELISA for testing antibodies against VSV-NJ based on the recombinant protein as a coating antigen. This method hadno cross reactions with eastern equine encephalomyelitis virus and equine west nile virus.The intra-plate and inter-plate variationcoefficients（CV）were less than 10%. The results indicated that the established indirect ELISA has good specificity and reproduc-ibility,and provided a basis for the detection of New Jersey type VSV antibodies in serum.

关 键 词：水疱性口炎病毒 原核表达 间接ELISA 

分 类 号：Q939.47[生物学—微生物学]