恶性疟原虫PfRh5F2片段pTGub21b表达载体的构建及融合蛋白表达鉴定  

Cloning,expression and identification of recombinant reticulocyte-binding protein homologue 5 of Plasmodium falciparum

作　　者：程莎莎[1] 郝文波[1] 罗树红[1] 廖小青[1] 

机构地区：[1]南方医科大学生物技术学院抗体工程研究所,广东广州510515

出　　处：《热带医学杂志》2014年第4期422-425,共4页Journal of Tropical Medicine

基　　金：国家自然科学基金(81171608)

摘　　要：目的克隆表达恶性疟原虫网状细胞结合蛋白同源体5(PfRh5)F2片段,并评价其抗原性。方法 PCR扩增目的基因片段,克隆到表达载体pTGub21b中,构建PfRh5F2/pTGub21b原核表达载体。IPTG诱导表达目的基因,SDS-PAGE电泳分析表达产物,并用Western blot检测其抗原性。结果成功构建了PfRh5F2/pTGub21b原核表达系统,并在大肠杆菌中可溶性高效表达,表达产物能被恶性疟患者血清识别,而重组抗原免疫鼠血清也能识别恶性疟患者血样中的相应抗原。结论恶性疟原虫PfRh5 F2片段在大肠杆菌中获得高效表达且表达产物具有良好的抗原性和免疫原性。Objective To clone and express reticulocyte- binding protein homolog 5 （PfRh5） of Plasmodium falciparum in order to evaluate the antigenicity of PfRh5. Methods The 507 bp（571-1 077） PfRh5 gene was specifically amplified by polymerase chain reaction, and was cloned into pTGub21b vector . The recombinant plasmid was transformed and then induced to express in the E. coli Rosetta. The expressed product was analyzed by SDS-PAGE and Western blot. Results The size of the expressed protein was about 80.4x103 Mr as predicted. Also it exhibited a specific reaction with immune sera obtained from patients with Pf malaria, and there is a strong reaction between antiserum against purified recombinant protein and the blood of patient infected with Pf. Conclusion These results demonstrate that the PfRh5 F2 fragment has been successfully expressed and the recombinant protein has certain antigenicity and immunogenicity.

关 键 词：恶性疟原虫 网状细胞结合蛋白同源体5 克隆 表达 

分 类 号：R382.31[医药卫生—医学寄生虫学]