鸭坦布苏病毒E蛋白膜外区的原核表达与鉴定  

Prokaryotic expression of the ectodomain of E gene from duck Tembusu virus and identification of the recombinant protein

作　　者：刘宗梁[1] 付钰广[2] 吉艳红[2] 李郁[1] 魏建忠[1] 朱启运[2] 

机构地区：[1]安徽农业大学动物科技学院,安徽合肥230036 [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,甘肃兰州730046

出　　处：《中国兽医科学》2013年第8期850-854,共5页Chinese Veterinary Science

基　　金：甘肃省高层次人才科技创新创业扶持行动项目(1013JHTA008);中央级公益性科研院所基本科研业务费专项(0032011027)

摘　　要：为获得鸭坦布苏病毒(duck Tembusu virus,DTMUV)E蛋白膜外区的表达产物,以DTMUV安徽分离株(AH10)E基因序列设计特异性引物,扩增E基因,将其克隆至原核表达载体pET-SUMO中,构建重组表达质粒pET-SUMO-E,然后将其转化至感受态细胞E.coli Rosetta(DE3)中,并以IPTG诱导表达。经SDS-PAGE检测,获得了70ku的表达产物,与预期结果相符。Western-blot分析表明,目的蛋白能与兔抗DTMUV E蛋白多克隆血清及DTMUV鸭阳性血清发生特异性反应,显示出良好的反应活性。结果表明,成功表达了E蛋白膜外区,这一成果为DTMUV疫苗制备和诊断试剂盒的研发奠定了基础。According to the sequence of E gene of duck Tembusu virus AH10 strain,a primer pair was designed to amplify the ectodomain of E gene by PCR.The E gene was subsequently cloned into pET-SUMO vector.The resultant plasmid pET-SUMO-E was transformed into E.coli Rosetta（DE3） competent cells.The highly expressed product induced by IPTG was confirmed to be about 70 ku in size by SDS-PAGE,which was consistent with the expectation.Western-blot assay showed that it could react with both the rabbit anti-DTMUV polyclonal serum and the positive serum from DTMUV-infected duck. In summary,a high-efficient expression E gene in this study paves a way for development of new vaccines or diagnostic reagents.

关 键 词：鸭坦布苏病毒 E蛋白膜外区 原核表达 鉴定 

分 类 号：S852.65[农业科学—基础兽医学]