猪繁殖与呼吸综合征病毒河北地方株GF1107株ORF7基因的克隆与表达  

Cloning and ExpressioninE.colifor ORF7 Gene of Porcine Reproductive and Respiratory Syndrome Virus Isolated in Hebei Province

作　　者：李鹏[1] 郑其升[1] 李斐[1] 周斌[1] 曹瑞兵[1] 陈溥言[1] 

机构地区：[1]南京农业大学动物疫病诊断与免疫重点开放实验室

出　　处：《中国兽医学报》2006年第5期468-470,共3页Chinese Journal of Veterinary Science

摘　　要：根据GenBank公布的猪繁殖与呼吸综合征病毒(PRRSV)ATCC VR2332株的核衣壳蛋白基因的核苷酸序列,设计并合成1对特异性引物。应用RT-PCR方法扩增PRRSV GF1107株(GenBank登陆号:AY684124)的ORF7片段。将扩增片段克隆入pMD18-T载体中获得含有相应片段的重组质粒pMDHB-ORF7。对质粒中的插入片段进行测序,应用DNAstar序列分析软件对所测序列与GenBank中PRRSV毒株进行同源性比较。将ORF7基因克隆入原核表达载体pET-32a(+)中,在IPTG的诱导下成功获得表达,经Western blotting分析表明,表达蛋白能够与PRRSV的阳性血清发生特异性反应,为PRRSV新型诊断试剂的研究奠定了基础。Using a pair of specific primers designed according to the relevant sequence from GenBank（Accession number GF1107/AY684124） ,ORF7 gene of porcine reproductive and respiratory syndrome virus isolated in Hebei province from suspected pigs was amplified with RT-PCR method. The RT-PCR product was cloned into the pMD18-T vector and the positive clone was named pMDHB-ORF7. After sequencing,the ORF7 gene was blasted with PRRSV genome sequence downloaded from NCB I with DNAstar software. Subsequently,after being cloned into the prokaryotic expression vector pET-32a, the ORF7 gene was successfully obtained with the induction of 1.0 mmol/L IPTG. Western blotting analysis indicated that the ex- pressed recombinant protein can vigorously react against the positive serum of PRRSV. Conclusions：the recombinant ORF7 protein can be applied in the detection for PRRSV antibodies in swine serum.

关 键 词：猪繁殖与呼吸综合征病毒 0RF7基因 克隆 原核表达 

分 类 号：S858.28[农业科学—临床兽医学] S852.65[农业科学—兽医学]