猪戊型肝炎病毒ORF2 C-端主要抗原区的克隆及原核表达

为研究猪戊型肝炎病毒(HEV)ORF2C-端主要抗原区的原核表达产物是否具有抗原性,用PCR技术对猪戊型肝炎病毒ORF2C-端基因进行扩增,经克隆与序列分析,发现该基因片段编码289个氨基酸残基。将该目的片段连接到原核表达载体pET-28a(+)中,转化表达菌BL21(DE3),成功构建了重组质粒pET-HEV-ORF2-C,其重组菌于37℃、0.5mmol/L IPTG诱导表达4h后,菌体裂解物经SDS-PAGE分析,在分子质量约为34ku处出现一新蛋白带,与预期的目的蛋白分子质量相符。质谱鉴定表明,成功地表达了HEV-ORF2C-端蛋白。Western-blot和抗体效价检测分析结果表明该重组蛋白具有良好的抗原性。

Cloning and prokaryotic expression of main antigenic area in the C-terminal of ORF2 peptide of swine hepatitis E virus

To study the antigenicity of prokaryotic expression of main antigenic area in the C-terminal of ORF2 peptide of hepatitis E virus(HEV),the C-terminal of the ORF2 peptide,which contains 289 amino acid residues,was amplified with PCR,sequenced and cloned into the vector pET-28a(+).The recombinant,named pET-HEV-ORF2-C,was expressed in Escherichia coli BL21(DE3) with 0.5 mmol/L IPTG induction at 37 ℃.The expressed product was analyzed by SDS-PAGE,which indicated that there was a new 34 ku-electrophoretic band corresponding to the presumptive interest protein.The result of mass spectro-metry showed that the C-terminal of HEV-ORF2 gene was expressed successfully.Western-blot and ELISA showed that the protein had good antigenicity.