PRRSV变异株ORF5全基因的可溶性表达与重组蛋白的纯化  

Soluble Prokaryotic Expression of ORF5 Gene of PRRSV Variant and Purification of Recombinant Protein

作　　者：任颖超[1] 黄娟[1] 单虎[1] 

机构地区：[1]青岛农业大学山东省高校预防兽医学重点实验室,山东青岛266109

出　　处：《中国兽医杂志》2016年第4期30-32,共3页Chinese Journal of Veterinary Medicine

基　　金：国家科技基础性工作专项(2012FY111000);山东省现代农业产业技术体系生猪产业创新团队建设;青岛农业大学高层次人才科研基金(6630719)

摘　　要：为了获得猪繁殖与呼吸综合征病毒(PRRSV)变异株可溶性糖基化囊膜蛋白(GP5),从p MD18-T-ORF5(V)重组质粒中PCR扩增得到PRRSV变异株LX13(1)结构蛋白ORF5全基因编码区,并克隆至原核表达载体p Cold TF DNA上,得到重组表达质粒p Cold-TF-GP5(V),转化大肠埃希菌BL21中诱导表达,并通过SDS-PAGE和Western Blot鉴定分析。结果表明,目的基因插入位置和阅读框正确,成功表达的融合蛋白分子量约为75.2 ku,并以可溶性蛋白的形式存在,表达量约占菌体总蛋白含量的22.26%。该融合蛋白能与鼠抗His标签单克隆抗体发生特异性反应,说明重组蛋白具有反应原性。该可溶性重组蛋白的获得为研究PRRSV变异株GP5蛋白的抗原性奠定了基础。The coding region of ORF5 gene from PRRSV variant was successfully amplified from p MD18-T-ORF5（V）by PCR,and was cloned into the prokaryotic expression vector p Cold TF. The recombinant plasmid p Cold-TF-GP5（V）was transformed into E.coli BL21 and the induced protein was confired by SDS-PAGE and Western blot.The results showed the target gene was inserted into the exact location and the reading frame was correct.An 75.2 ku soluble protein was successfully induced with a content of 22.26% of total bacterial protein. The recombinant protein showed good reaction with the His-tag monoclonal antibody.Our results contribute to the antigenicity study of glycosylated envelope protein 5（GP5）from PRRSV variant.

关 键 词：猪繁殖与呼吸综合征病毒 变异株 ORF5基因 可溶性原核表达 

分 类 号：S852.651[农业科学—基础兽医学]