【摘要】 根据GenBank中发表的猪圆环病毒2型(PCV2)ORF2基因序列,设计合成1对特异性引物,用PCR方法从接种PCV2吉林株(JL01)PK-15细胞中,扩增出PCV2毒株的ORF2基因。将扩增片段克隆于pMD18-T载体,进行序列测定。结果表明,PCV2 JL01毒株的ORF2基因核苷酸长度为702bp。将重组质粒用SalⅠ和XhoⅠ酶切后与同样处理的pET-32a载体连接,转化BL21细胞后,挑取阳性克隆。经PCR和酶切鉴定后,用IPTG诱导,细菌裂解液经SDS-PAGE和Western-blot分析,表明ORF2基因在大肠杆菌中得到了表达,并能被PCV2阳性血清所识别。更多还原

【Abstract】 According to the nucleotide sequences of porcine circovirus type 2(PCV2) published in GenBank,a pair of primers specific to the ORF2 gene of PCV2 was designed and synthesized.ORF2 gene was amplified by polymerase chain reaction(PCR) from PCV2 Jilin strain(JL01) which was isolated from PK-15 cell line and had been characterized.The PCR product was cloned into the pMD 18-T vector and then sequenced.The recombinant plasmid pMD-ORF2 digested by Sall and XhoⅠ was subcloned into prokaryotic expression vector pET-32a.The resultant pET-ORF2 was transformed into E.coli BL21 cells and induced by IPTG.The result of SDS-PAGE and Western-blot indicated that the ORF2 gene had been expressed and could be recognized by the antiserum against PCV2.更多还原