恶性疟原虫裂殖子表面蛋白MSP1-42基因的合成及其表达

目的 利用大肠杆菌表达系统表达可溶性的恶性疟原虫(3D7株)裂殖子表面蛋白1C末端MSP1-42蛋白.方法 采用两步PCR法拼接合成全长msp1-42基因,将测序正确的msp1-42基因构建至pET32a(+)表达载体,获得pET32amsp表达质粒,热激转化至宿主菌Rosetta gami(DE3)中进行诱导表达,用免疫印迹法检测表达产物.结果 全合成了恶性疟原虫3D7株msp1-42基因,合成基因在Rosetta gami(DE3)中以可溶性形式高水平表达,所表达的重组蛋白能与识别MSP1-19构象表位的单克隆抗体mAb5.9、PfCP2.9兔血清及恶性疟患者血清发生免疫反应.结论 合成的msp1-42基因能在大肠杆菌表达系统中以可溶性的形式表达,所表达的重组蛋白MSP1-42具有抗原性.

Synthesis and expression of 42 kDa C-terminal region of the major merozoite surface protein1 (MSP1-42) of Plasmodium falciparum in Rosetta gami

Objective To produce soluble recombinant MSP1-42 protein of Plasmodium falciparum strain 3D7 in Rosetta gami. Methods Two-step PCR method was utilized to synthesize the msp1-42 gene.The synthetic gene was then inserted into pET32a(+) to construct expressing plasmid pET132 amsp.Rosetta gami(DE3) was transformed with pET32 amsp and used for expression of recombinant MSP1-42 protein.SDS-PAGE and Westem blot were applied to detect the MSP1-42 protein. ResuIts The synthetic msp1-42 gene Was generated and expressed to produce soluble MSP1-42 recombinant protein in Rosetta gami.The recombinant protein could be recognized by conformational specific monoclonal antibody mAb5.9, rabbit sera immunizedwith PfCP2.9 and serum of patient infected with P. falciparum. Conclusion The synthetic msp1-42 3D7 gene could be expressed in Rosetta gami as soluble recombinant protein with antigenicity.