日本血吸虫双表膜抗原融合基因原核表达质粒的构建及表达  

Construction and Expression of Prokaryotic Expression Vector for Schistosoma japonicum Double Membrane Antigen Fusion Gene

作　　者：刘淼[1] 朱海玉[1] 彭强[2] 徐广文[2] 王盈[2] 沈际佳[1] 

机构地区：[1]安徽医科大学基础医学院病原生物学教研室,合肥230032 [2]安徽医科大学,七年制临床医学专业合肥230032

出　　处：《中国生物制品学杂志》2010年第2期146-149,共4页Chinese Journal of Biologicals

基　　金：安徽省自然科学基金(050430805);安徽省教育厅自然基金(2006KJ359B);安徽医科大学七年制临床医学专业学生"早期接触科研"训练项目

摘　　要：目的构建编码日本血吸虫表膜蛋白SjTsp2与Sj29融合基因的原核表达质粒,并表达重组蛋白。方法利用基因SOEing PCR技术得到SjTsp2与Sj29融合基因,将其克隆入原核表达载体pET32a中,转化E.coli BL21(DE3),IPTG诱导表达,SDS-PAGE和Western blot进行鉴定。结果序列分析显示,融合基因与两个目的基因的同源性达100%;SDS-PAGE显示表达的重组蛋白相对分子质量为43000,表达量占菌体总蛋白的20%以上;Western blot显示其分别能与SjTsp2和Sj29蛋白的单克隆抗体结合。结论已成功构建了SjTsp2与Sj29融合基因原核表达质粒,并在大肠杆菌中表达了重组蛋白,为研制血吸虫病疫苗提供了材料。Objective To construct a recombinant prokaryotic expression vector for fusion gene encoding the membrane proteins SjTsp2 and Sj29 of Schistosoma japonicum （Sj）and express the recombinant protein. Methods SjTsp2 and Sj29 fusion gene was amplified by SOEing PCR and cloned into prokaryotic expression vector pET32a. The constructed recombinant plasmid was transformed to E. coli BL21（DE3）for expression under induction of IPTG. The expressed product was identified by SDS-PAGE and Western blot. Results Sequencing proved that both the homologies of the fusion gene to two target genes were 100%. SDS-PAGE proved that the expressed product, with a relative molecular mass of 43 000, contained more than 20% of total somatic protein. Western blot showed specific bindings of the expressed product to the McAbs against SjTsp2 and Sj29 proteins. Conclusion A recombinant prokaryotic expression vector for SjTsp2 and Sj29 fusion gene was successfully constructed and expressed in E. coli, which provided a material for development of Sj vaccine.

关 键 词：日本血吸虫 SjTsp2 Sj29 融合基因 原核表达 

分 类 号：R383.24[医药卫生—医学寄生虫学] Q786[医药卫生—基础医学]