高致病性猪蓝耳病病毒GST-N融合蛋白的原核表达及纯化  

Prokaryotic expression and purification of GST-N fusion protein of highly pathogenic Porcine reproductive and respiratory syndrome viruses

作　　者：李晓辉[1] 何建斌[1] 刘金玲[1] 罗芳 

机构地区：[1]沈阳农业大学畜牧兽医学院,辽宁沈阳110161 [2]公安部昆明警犬基地,云南昆明650204

出　　处：《黑龙江畜牧兽医》2010年第4期30-32,共3页Heilongjiang Animal Science And veterinary Medicine

基　　金：国家科技攻关项目(413010209);沈阳农业大学校青年基金项目(200705)

摘　　要：为了构建高致病性猪蓝耳病病毒(HPPRRSV)GST-N融合蛋白,试验采用RT-PCR技术扩增ORF7基因片段,对其胶回收产物进行测序分析,再将目的片段连接到质粒PGEX-6P-1上,得到重组质粒PGEX-6P-1-N,并转化到E.coliBL21(DE3)感受态细胞中,筛选阳性菌落,并用IPTG进行诱导表达,应用SDS-PAGE和Western-blot对表达产物进行检测与鉴定。结果表明:该序列与GenBank公布的HN2007的同源性为97%;重组质粒在大肠杆菌中获得了表达,表达产物为42ku的GST-N融合蛋白。In order to construct prokaryotic expression of GST - N fusion protein of highly pathogenic porcine reproductive and respiratory syndrome viruses（ HPRRSV）, ORF7 gene was amplified by RT- PCR and was sequenced. Its homology compared to HN2007 （published Gen- Bank） was 97%. The fragment was connected to the plasmid PGEX -6P - 1 ,and got the recombinant plasmid PGEX -6P - 1 - N, and then was transformed into competent cells E. coli BI21 （DE3）, and induced with IPTG. The expression of products was detected and identified using SDS - PAGE and Western - blot. The results showed that the recombinant plasmid was successful expression of the 42 ku product and was purified.

关 键 词：ORF7基因 原核表达 纯化 GST—N融合蛋白 

分 类 号：S852.659.6[农业科学—基础兽医学]