猪细小病毒NS1蛋白主要抗原表位区的原核表达

Expression of Major Epitope Domain of NS1 of PPV in E.coli

目的:用大肠杆菌表达猪细小病毒NS1基因的主要抗原表位区。方法:通过对GenBank发表的猪细小病毒(Porcine Par-vovirus,PPV)中国株非结构蛋白NS1的氨基酸序列分析,确定了其中抗原性较高的区域,针对此区域设计并合成了一对特异性引物,扩增出包含NS1主要抗原表位区的843bp的片段。酶切后连接到原核表达载体pET30a(+)上,构建的原核表达载体pET30a-NS1s转化大肠杆菌BL21(DE3)感受态细胞,诱导表达。结果:重组蛋白大小约为43kD,与预期大小相符。Westernblot分析表明,该蛋白能与标准阳性血清发生特异性反应,证明该蛋白具有生物学活性。结论:NS1蛋白主要抗原表位区在大肠杆菌中成功表达,具有免疫原性。可作为鉴别诊断抗原用于PPV的临床检测。

Objective：Express major epitope domain of NS1 of PPV in E.coli.Method：According to the amino acid sequence of porcine parvovirus china strain in GenBank,We found the major epitope domain in NS1.A pair of specific primer was designed to amplify the major epitope domain of NS1.The size of PCR product is 843 base pairs.This gene was cloned into the multiple cloning site of pET30a（＋） vector after double enzyme digested.The recombinant of pET30a-NS1s was transformed into E.coli BL21（DE3） competent cells and induced to express.Result：The size of recombinant protein is 43 kDa accord with anticipate.The results of Western blot showed that the protein was specifically reacted with porcine parvovirus positive serum,indicating the recombinant protein had wonderful specificity.Conclusion：Major epitope domain of NS1 protein was expressed in E.coli and showed good specificity.It can be used as antigen in diagnostic assay for detection antibodies of PPV in practice.