高致病性猪繁殖与呼吸综合征病毒ORF3基因的原核表达

Prokaryotic Expression of Highly Pathogenic PRRSV ORF3 Gene

为进一步研究猪繁殖与呼吸综合征病毒(PRRSV)GP3蛋白的结构、功能及免疫学特性,根据GenBank中已发表的高致病性PRRSV HuN4株ORF3基因序列,设计合成扩增PRRSV ORF3基因的引物。将扩增的基因克隆到表达载体pProEx HTb中,转化到大肠埃希菌BL21进行表达。Western blot分析表明,GP3蛋白在大肠埃希菌中获得正确表达,且具有良好的反应活性。将纯化的GP3蛋白免疫小鼠制备抗GP3多克隆血清,酶联免疫吸附试验(ELISA)测定其效价,结果在小鼠体内产生较高滴度的特异性抗体,证明PRRSV GP3蛋白具有良好的抗原性。

To further study the structure,function and immunogenicity of highly pathogenic porcine reproductive and respiratory syndrome virus（PRRSV） GP3.A pair of primers for ORF3 gene of PRRSV were designed according to the PRRSV（HuN4 strain） sequence available in GenBank.The ORF3 gene of PRRSV was cloned into the expression vector pProEx HTb and the recombinant plasmid was transformed into E.coli competent cells BL21.Expression of fusion protein in E.coli was observed by Western blot.The purified GP3 was inoculated into Balb/c mice to prepare anti-GP3 serum,the specificity and titer of the antiserum were evaluated by ELISA.Specific high titer antibodies were detected in the vaccinated mice.The results showed that the GP3 expressed in E.coli could induce high antibody titer against GP3,and had strong antigenicity.