阴道毛滴虫黏附蛋白65-3基因原核表达质粒的构建及表达  

Construction of Prokaryotic Expression Plasmid and Expression of Adhension Protein 65-3 Gene of Trichomonas vaginalis

作　　者：朱晓燕[1] 王雅静[1] 杨树国[2] 毕世樑[3] 廖琳[1] 

机构地区：[1]四川大学华西基础医学与法医学院寄生虫学教研室,成都610044 [2]郧阳医学院寄生虫学教研室 [3]四川大学华西第二医院妇产科

出　　处：《四川动物》2008年第2期273-276,共4页Sichuan Journal of Zoology

摘　　要：质粒pMD-18T-ap65-3经XhoI和BamHI双酶切,1.0%凝胶电泳后纯化回收阴道毛滴虫黏附蛋白65-3基因,定向克隆至原核表达载体pET-32a(+),构建原核表达质粒pET32a-ap65-3。重组质粒转化大肠埃希菌JM109感受态细胞中,筛选阳性克隆,进行PCR、酶切及测序鉴定正确后,将重组质粒pET32a-ap65-3转化于大肠埃希菌BL21中,异丙基-β-D硫代半乳糖苷(IPTG)诱导重组蛋白表达后利用十二烷基磺酸钠一聚丙烯酰胺凝胶电泳(SDS-PAGE)和免疫印迹(Western blot)对重组蛋白进行分析鉴定。本实验为研究阴道毛滴虫病的致病机制及蛋白生物学功能奠定了基础。The plasmid pMD-18T-ap65-3 was digested by Xho I and BamH I, the ap65-3 gene was subeloned into the pET-32a（＋） expression vector. The recombinant plasmid pET32a-ap65-3 was identified by PCR technique and restriction analysis, the insert fragment was sequenced. Then the correctly constructed recombinant plasmid was transferred into E. coli BL21 and induced to express by IPTG. The expressed recombinant protein was identified by SDS-PAGE and Western blot. Prokaryotie expression plasmid pET32a-ap65-3 was constructed and expressed in E. eoli BL21 successfully, which can be useful for the further studies on the pathogenesis mechanism of Triehomonas vaginalis and biological function of the protein.

关 键 词：阴道毛滴虫 黏附蛋白65-3 原核表达质粒 基因表达 

分 类 号：Q78[生物学—分子生物学]