禽流感病毒NA基因稀有密码子分析及其原核表达  

Analysis of Rare Codons of NA Gene of AIV and Its Prokaryotic Expression

作　　者：任丽伟[1] 丁爱军[1] 杨海燕[1] 田智泉[1] 孙敏[1] 徐贵江[1] 李碧春[1] 

机构地区：[1]扬州大学动物科学与技术学院,江苏扬州225009

出　　处：《中国家禽》2009年第21期18-21,共4页China Poultry

基　　金：国家自然科学基金(30671509和30871791);高等学校博士学科点专项科研基金资助项目(20061117004);江苏省自然科学基金(BK2007077);江苏省高校自然科学重大基础研究项目(08KJA230001)

摘　　要：通过原核表达获取禽流感病毒(AIV)NA蛋白,为深入研究该蛋白的抗病性奠定基础。本试验以pMD19-T-NA为模板,PCR扩增NA基因完整开放阅读框(ORF),然后克隆到原核表达载体pET-32a(+)中,构建表达质粒pET-NA;转化E.coliBL21(DE3),IPTG诱导表达,SDS-PAGE分析检测;并利用稀有密码子在线分析软件对NA基因ORF进行分析。SDS-PAGE分析显示融合蛋白获得了表达,大小为65.4ku;稀有密码子统计表明,NA基因ORF中稀有密码子的比例高达13.4%,甚至还有多个稀有密码子串联成簇存在。重组质粒pET-NA构建正确,且在E.coliBL21(DE3)中成功诱导表达出目的蛋白。To understand the antiviral specificity of NA protein of avian influenza virus （AIV）,one of NA proteins has been obtained by prokaryotic expression. This experiment made pMD-19T-NA as a template,PCR amplificated of NA gene complete open reading frame,and then cloned into the prokaryotic expression vector pET-32a （＋）to construct the expression plasmid pET-NA,transformed into E.coli BL21 （DE3）,inducing expressed with IPIG,analyzed and detected with SDS-PAGE ;and used the rare codon on-line analysis software to analyze the NA gene ORF. The SDS-PAGE analysis showed that the fusion protein was expressed in the size of 65.4 ku. Rare codons statistics showed that the proportion of rare codons in NA gene ORF was as high as 13.4%,and even a number of tandem rare codons clusters existed. We had correctly constructed the recombinant plasmid pET-NA,and expressed the fusion protein by being induced in E.coli BL21（DE3）.

关 键 词：基因 大肠杆菌 原核表达 稀有密码子 

分 类 号：S852.659.5[农业科学—基础兽医学] Q78[农业科学—兽医学]