PRRSV Hn-1/06株GP5蛋白基因表达载体的构建及高效表达  

Construction of Prokaryotic Expression Vector and High-level Expression of GP5 Gene of PRRSV Hn-1/06 Strain

作　　者：卢高峰[1] 夏平安[1] 刘明莉[1] 李伟娟[1] 邱璜[1] 李素平[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：《江西农业学报》2009年第1期91-94,共4页Acta Agriculturae Jiangxi

摘　　要：通过对PRRSV Hn-1/06株ORF5基因序列分析,设计删除信号肽和跨膜功能区的ORF5序列的3对引物,应用重叠延伸PCR以PTG19-T-ORF5质粒为模板进行扩增目的片段,得到长度为410 bp的片段。然后将此基因亚克隆到原核表达载体PET32a,经筛选获得了阳性重组质粒PET32a-ORF5abc,进而在IPTG的诱导下成功表达,获得了大小约为34 kD的融合蛋白GP5abc-His,Western blotting检测结果证实表达的融合蛋白具有良好的生物学活性。According to analysis of the ORF5 gene order in PRRSV Hn - 1/06 strain, three pairs of primers for the deletion of signal peptide and ORF5 sequence in transmembrane domain function area were designed, and PTGI9 -T -GP5 was used as a template, 410 bp segment was obtained by overlap extension PCR amplification. Then after being sub - cloned into the prokaryotie expression vector PET32a, the ORF5abc gene was successfully obtained with the inducement of 1.0 mmol/L IPTG. Western -blotting was performed to confirm that the expressed fusion protein GP5abc - His （about 34 kD） could speeifically react with antiserum against PRRSV.

关 键 词：PRRSV ORF5基因 RT—PCR 原核表达 

分 类 号：Q782[生物学—分子生物学]