普氏立克次体120kDa表面抗原N段基因的克隆与表达

目的　克隆和表达普氏立克次体 (Rickettsiaprowazekii) 12 0kDa外膜蛋白N段基因。 方法　采用PCR方法 ,从普氏立克次体基因组中扩增其 12 0kDa表面抗原N段基因片段 ,将其与原核表达载体 pQE30连接 ,构建重组质粒 pQE30 /2 6kDa,将重组质粒转入大肠杆菌 ,用IPTG诱导大肠杆菌内的目的基因表达。结果　获得长为 717bp的PCR片段 ,序列分析结果与已知普氏立克次体 12 0kDa表面抗原基因一致 ;SDS -PAGE检测表达产物 ,在相对分子量 2 6 .3kDa处有一表达带 ;免疫印迹分析证明它能与普氏立克次体免疫兔血清反应 ;经双波长薄层扫描分析 ,目的蛋白占全菌体蛋白的 2 0 % ;提取表达菌体包涵体检查 ,证明目的蛋白主要存在于包涵体中。结论　普氏立克次体 12 0kDa表面抗原N段基因在大肠杆菌中获得了高效表达 ,该重组蛋白具有良好的免疫反应性。

Cloning and expression of gene fragment encoding the N region of 120kDa surface antigen of Rickettsia prowazekii in E.coli.

To Clone and express the gene fragment encoding the N region of 120kDa surface antigen of R.prowazekii in E.coli.A gene fragment encoding the N region of 120kDa surface antigen was amplified from the genome of R.prowazekii by PCR.The gene fragment was cloned into prokaryotic expression vector,pQE30;E.coli cells were transformed with the recombinant plasmid;the transformants were induced to express the fusion protein by IPTG.The fusion protein from the transformants was approximately 26.3 kDa in SDS-PAGE analysis and 20% of total cell protein in scanning analysis;the fusion protein reacted with the serum from a rabbit immunized with R.prowazekii cells.The gene fragment encoding the N region of 120kDa surface antigen of R.prowazekii can be efficiently expressed in E.coli and the recombinant protein is immunoreactive.