水疱性口炎病毒糖蛋白基因主要抗原表位区在大肠埃希氏菌中的高效表达及纯化  

Highly efficient expression and purification of truncated glycoprotein gene of vesicular stomatitis virus in Escherichia coli

作　　者：祁会彩[1] 龚振华[2] 杨增岐[1] 李葳[2] 刘俊辉[2] 潘康锁 王若聪[2] 蒋正军[2] 郑增忍[2] 

机构地区：[1]西北农林科技大学动物科技学院,陕西杨凌712100 [2]农业部动物检疫所,山东青岛266032 [3]陕西省畜牧兽医总站,陕西西安710016

出　　处：《中国兽医科技》2005年第10期757-760,共4页Chinese Journal of Veterinary Science and Technology

基　　金：国家"十五"重大专项(2001BA804A22)

摘　　要：利用PCR技术扩增编码水疱性口炎病毒印第安纳型(VSV-Ind)糖蛋白基因的主要抗原表位区,通过引入其两端的EcoRⅠ和XhoⅠ酶切位点,定向插入到pET30c质粒T7启动子下游的多克隆区,构建重组质粒pET30c-vsvG,并转化至宿主菌BL21(DE3)株中。用1 mmol/L IPTG诱导后,SDS-PAGE电泳表明,重组蛋白在大肠埃希氏菌中得到了高效表达,经过4 h表达即可达到高峰;融合蛋白的相对分子质量为57 ku。重组蛋白含有六聚组氨酸尾,主要以包涵体形式存在,Ni柱纯化后的蛋白经Western-blotting鉴定,具有良好的抗原性和特异性。The glycoprotein gene of vesicular stomatitis virus Indiana （VSV-Ind） without signal and transmembrane region was amplified by PCR. The DNA sequenced EcoR I and Xho Ⅰ restricted core fragment was cloned in a frame downstream of T7 promoter in the pET30c vector, then the recombinant plasmid pET30c-vsvG was transformed into E. coli BL21（DE3 ）. The recombinant fusion protein was expressed highly in E. coli BL21 four hours later induced by 1 mmol/L of IPTG and the molecular weight of the fusion protein was about 57 ku. The recombinant protein containing a six-histidine tag aggregated into the inclusion body was purified by Ni-column, subsequently examined and analysed by SDS-PAGE and Western-blotting tests. The results showed the purified protein was of good antigenicity and specificity.

关 键 词：水疱性口炎病毒 糖蛋白 抗原表位 表达 纯化 

分 类 号：S852.659.5[农业科学—基础兽医学] Q786[农业科学—兽医学]