猪Viperin基因丙酸诱导型载体构建及表达  

Porcine viperin gene vector construction and expression of inducible propionate

作　　者：王晓婷[1] 樊文杰[1] 查光明[1] 郭豫杰[1] 钟凯[1] 李和平[1] 

机构地区：[1]河南农业大学农业部动物生化与营养重点开放实验室,河南郑州450002

出　　处：《中国兽医学报》2016年第5期864-868,共5页Chinese Journal of Veterinary Science

基　　金：农业部转基因重大专项基金资助项目(2014ZX0801015B);郑州市科技攻关计划基金资助项目(141PPTGG407)

摘　　要：本试验以猪十二指肠cDNA为模板,通过PCR、酶切鉴定克隆Viperin基因,利用非酶连接技术将其连接丙酸诱导型表达载体pEX5,经丙酸钠诱导表达,SDS-PAGE和Western-blotting进行表达蛋白鉴定。结果显示,成功克隆了猪Viperin基因,包括完整开放阅读框ORF,其长度为1 046bp,将扩增得到的序列成功连接到载体pEX5,构建了pEX5-Viperin;SDS-PAGE和Western-blotting。6×His-NusA-TEV-LIC-Viperin融合蛋白主要以包涵体形式表达,其相对分子质量约96 500。结果表明,pEX5-Viperin的成功构建,为后续猪Viperin蛋白的纯化,Viperin基因与病毒互作关系及信号通路调控机制的研究奠定了基础。Viperin is an important antiviral innate immune molecules,has the function of broadspectrum antivirus,can be induced expression by a variety of dsRNA,DNA virus and pathogen.The experiment used porcine duodenum cDNA as template,through PCR,enzyme cutting identification to clone viperin gene,the use of non enzymatic connection technology connects the propionic acid inducible expression vector pEX5,the expressed protein induced by sodium propionate was identified by SDS-PAGE and Western-blotting.The successful cloning the porcine viperin gene,included the complete open reading frame of ORF with the length of 1 046 bp,the sequence amplified was successfully connected to the vector pEX5,constructed pEX5-Viperin,SDS-PAGE and Western-blotting results showed that 6×His-NusA-TEV-LIC-viperin fusion protein was expressed mainly in the form of inclusion body,the relative molecular mass of about 96 500.The resust lays a foundation for subsequently research on porcine viperin protein purification,the interaction relationship between virus and viperin gene,and regulation of signal pathway.

关 键 词：猪 Viperin 丙酸诱导型 载体构建 表达 

分 类 号：Q78[生物学—分子生物学] S828[农业科学—畜牧学]