中蜂囊状幼虫病病毒结构蛋白的表达及其抗血清的制备  

Expression and antiserum preparation of structural protein of Chinese sacbrood virus

作　　者：沈克飞[1] 张邑帆[1] 曹兰[1] 王瑞生[1] 杨金龙[1] 杨柳[1] 戴荣国[1] 

机构地区：[1]重庆市畜牧科学院,重庆402460

出　　处：《中国生物制品学杂志》2013年第1期89-92,共4页Chinese Journal of Biologicals

基　　金：现代农业产业技术体系建设专项(CARS-45-SYZ 15);重庆市畜牧科学院基本科研业务费项目(10615)

摘　　要：目的原核表达中蜂囊状幼虫病病毒(Chinese sarbood virus,CSBV)结构蛋白,并制备其抗血清。方法采用RT-PCR法扩增CSBV结构蛋白基因片段,克隆至pGEX-4T-1载体中,转化大肠杆菌BL21(DE3),IPTG诱导表达。表达的重组蛋白经Gluthathione-Sepharose 4B亲和层析纯化后,免疫小鼠,制备抗血清,并进行Western blot鉴定。结果重组表达质粒经菌落PCR鉴定证明构建正确;表达的重组蛋白相对分子质量约52 000,以可溶性形式表达;纯化后纯度可达95%以上;制备的抗血清可识别天然CSBV结构蛋白。结论成功在大肠杆菌中表达了CSBV结构蛋白,并制备了其抗血清,为进一步研究囊状幼虫病病毒与宿主的相互作用及开发血清学诊断方法奠定了基础。Objective To express the structural protein of Chinese sacbrood virus（CSBV） in prokaryotic cells and prepare its antiserum.Methods The gene fragment encoding structural protein of CSBV was amplified by RT-PCR and cloned into vector pGEX-4T-1.The constructed recombinant plasmid pGEX-CSBV was transformed to E.coli BL21（DE3） for expression under induction of IPTG.The expressed recombinant protein was purified by Gluthathione-Sepharose 4B affinity chromatography and used for immunization of mice.The prepared antiserum as identified by Western blot.Results Colony PCR proved that recombinant plasmid pGEX-CSBV was constructed correctly.The expressed recombinant protein,with a relative molecular mass of about 52 000,existed in a soluble form and reached a purity of more than 95% after purification.The prepared antiserum recognized the structural protein of natural CSBV.Conclusion The structural protein of CSBV was successfully expressed in E.coli,which laid a foundation of further study on interaction between sacbrood virus and its host as well as development of serological diagnostic method.

关 键 词：囊状幼虫病病毒 病毒结构蛋白质类 基因表达 抗血清 

分 类 号：S895.133[农业科学—特种经济动物饲养] R392-33[农业科学—畜牧兽医]