用四环素调控启动子调控表达恶性疟原虫裂殖子表面蛋白1全合成基因

目的 ]用含四环素调控启动子 (PLtetO 1 启动子 )的pZE11质粒表达恶性疟原虫MSP1全合成基因及其C末端 42kDa片段。 方法 ]将MSP1全合成基因和MSP1C末端 42kDa片段基因分别克隆入受四环素诱导调控的质粒pZE11上 ,转化大肠杆菌DH5αZ1,质粒经酶切、SDS PAGE电泳和Westernblotting反应鉴定重组质粒的构建和重组质粒在DH5αZ1中的表达。 结果 ]成功地构建了重组pZE11 MSP1质粒和pZE11 MSP1 42质粒 ,SDS PAGE电泳和免疫印迹 (Westernblotting)反应证明两个重组质粒在DH5αZ1中表达了 190和 42kDa蛋白。 结论 ]含PLtetO 1 启动子的新型表达载体成功地表达了恶性疟原虫MSP1蛋白 ,降低表达产物对宿主细胞的毒性 ,并可为构建受四环素诱导的疟疾———伤寒口服疫苗株提供依据。

INDUCIBLE EXPRESSION OF MSP1 GENE OF PLASMODIUM FALCIPARUM BY A TETRACYCLINE-CONTROLLED PROMOTER

OBJECTIVE: To express the entire MSP1 gene of Plasmodium falciparum and its C-terminal 42 kDa fragment using a tetracycline-controlled PLtetO-1 promoter. METHODS: The entire MSP1 gene and 42 kDa fragment gene were cloned into the plasmid of pZE11, and transformed into E. coli DH5 alpha Z1. Restriction enzyme analysis, SDS-PAGE and Western blotting were used to examine two recombinant plasmids and their expression in E. coli DH5 alpha Z1. RESULTS: The recombinant plasmids of pZE11/MSP1 and pZE11/MSP1-42 were constructed successfully. The expressive products about 190 kDa and 42 kDa of two genes in E. coli DH5 alpha Z1 were identified by SDS-PAGE and Western blotting. CONCLUSION: Tightly controlling expression of the MSP1 gene in E. coli is essential to reduce the toxicity of the product to its host cells as well as to provide a feasibility to construct Salmonella vaccine strain which can inducibly express the important malarial vaccine candidate gene.