人微小病毒B19 VP1基因原核表达克隆的构建

①目的　构建人微小病毒B19VP1基因的原核高效表达系统。②方法　通过PCR扩增B19病毒主要抗原决定簇VP1区段基因 ,将目的基因克隆到原核表达载体pGEX 4T 2上 ,IPTG诱导表达融合蛋白 ,产物经亲和层析初步纯化 ,以Western Blot进行鉴定 ,并包被ELISA板 ,对临床血清进行检测。③结果　pGEX 4T 2 VP1可在大肠杆菌中高效表达 ,表达产物经纯化后可得单一目的蛋白条带。ELISA结果表明 ,其灵敏度、特异度等指标与德国Virion试剂盒有较好的符合率。④结论　该重组蛋白具有良好的抗原性 ,为研制和开发具有我国独立知识产权的B19病毒基因工程抗原ELISA诊断试剂盒提供了有价值的资料

CONSTRUCTION OF HUMAN PARV OVIRUS B19 VP1 GENE PROKARYOTIC EXPRESSION CLONES

Objective\ To construct human parvovirus B19 VP1 gene prokaryotic expression clones.\ Methods\ VP1 gene, the main antigenic determinants of B19 virus, was amplified by PCR technique and inserted into the prokaryotic expression plasmid pGEX 4T 2, the expressed fusion protein was induced with IPTG and purified by affinity chromatography and its antigenicity was identified by Western blot, also the ELISA coating with the recombinant protein was used to screen the clinical serums. Results\ pGEX 4T 2 VP1 plasmid expressed a fusion protein efficiently in E. coli., ELISA analysis showed that the purified expressed fusion protein had a good agreement with the Virion ELISA kit.\ Conclusion\ This expressed protein had strong antigenic characterization, it will provide valuable information for developing B19 recombinant antigen ELISA kit in our country.