戊型肝炎病毒ORF_2基因在人胚肾293细胞中的表达

Expression of ORF_2 protein from hepatitis E virus in 293 cell

目的探讨戊型肝炎病毒(hepatitis E virus,HEV)ORF2基因在人胚肾293细胞的表达。方法利用聚合酶链反应(PCR)技术,以HEV ORF2为模板扩增其部分基因片段,克隆入真核表达载体pcDNA 3．1中,构建真核表达载体质粒,转染人胚肾293细胞,用G418(Geneticin,遗传霉素)加压筛选得到稳定表达株。用western-blot方法对表达产物进行免疫学特性初步研究。结果含ORF2结构基因部分片段的重组质粒在人胚肾293细胞中得到表达。结论表达的ORF2重组蛋白具有抗HEV抗体识别的抗原表位和一定的抗原性,为进一步研制HEV的诊断试剂奠定了基础。

Objective To study the immunoreaction of the recombinant proteins encoded by the fragments of ORF2 （ second open readingframes） gene of hepatitis E virus （ HEV ）. Methods The aimed sequence from the full-length ORF2 in clone PEH2, which was derivedfrom a Chinese strain of HEV,was amplified and cloned it into vector pcDNA3.1 which was then transfected to 293 cell. Results TheORF2 protein was present in the soluble fractions of the cell lysate. The expressed protein of HEV ORF2 in 293 cells by using a plasmidpcDNA3.1-based system showed positive on immunoblots probed against antibodies raised in BALB/c mice. Conclusion The experiental results laid a foundation for developing diagnostic reagent to detect HEV by using the expressed products of ORF~ protein.