猪繁殖与呼吸综合征病毒Hn-1/06株M基因的原核表达与重组蛋白的复性研究  

Prokaryotic expression and the recombinant protein renaturation of the M gene of porcine reproductive and respiratory syndrome virus Hn-1/06 strain

作　　者：刘明莉[1] 张凤华[1] 范旭[1] 赵琳[1] 刘玉松[1] 徐红运[1] 夏平安[1] 崔保安[1] 

机构地区：[1]河南农业大学牧医工程学院,河南郑州450002

出　　处：《河南农业大学学报》2009年第5期521-525,共5页Journal of Henan Agricultural University

基　　金：河南省科技攻关项目(0623011800)

摘　　要：采用PCR方法从重组质粒pTG19-T-M扩增得到缺失N端疏水区序列的基因片段tM.将tM与原核表达载体pET-32 a进行连接并转化,重组质粒经鉴定并测序.结果表明,目的基因在大肠杆菌BL21细胞中成功地表达了醉繁殖与呼吸病毒重组蛋白pET-tM,表达量为31%.表达产物经SDS-PAGE分析,得到分子量约为29 kD的重组蛋白且以包涵体形式存在.8 mol.L-1尿素变性溶解包涵体,再用稀释与透析相结合的方法对重组蛋白进行复性.复性蛋白经W estern-b lot检测,结果证明复性蛋白可被PRRSV阳性血清识别用.The gene segments tM deleting hydrophobic region sequence were successfully amplified from recombinant plasmids pTG19-T-M by PCR.The fragment was inserted into the prokaryotic expression vector pET-32a to construct the recombinant plasmid pET-tM,and expressed in E.Coli BL21.The result indicated that the fusion protein pET-tM was successfully expressed.SDS-PAGE analysis demonstrated the molecular weight of reorganization protein existing in cytorrhyctes type was 29 kD and amountedto 31% of the mass of bacterial proteins in cvtorrhvctes. The renaturation of reorganization protein was carried on by dilution and the dialysis after treatment with the 8 mol·L^-1 urea. The renaturation reorganization protein was analyzed by Western-blot,the result demonstrated that the renaturation reorganization protein was recognized by the PRRSV masculine blood serum.

关 键 词：PRRSV M蛋白 原核表达 

分 类 号：S852.2[农业科学—基础兽医学]