弓形虫截短SAG2基因的克隆表达与免疫活性分析

Prokaryotic Soluble Expression of Truncated SAG2 Protein of Toxoplasma gondii and Its Antigenic Activity

构建截短的弓形虫表面抗原2(SAG2)原核表达系统,并探讨其抗原活性。PCR扩增去掉信号肽和C-末端的SAG2基因片段,插入原核表达载体pGEX-4T-3后转化到大肠杆菌DH5α,并用IPTG诱导。SDS-PAGE和Western blot鉴定重组SAG2t的表达及其免疫反应原性。每升菌液约纯化出4mg rSAG2t蛋白;Western blot显示,ME49株感染的鼠血清和rSAG2t免疫的鼠血清均可强烈识别表达的截短SAG2蛋白。原核表达体系实现了截短的SAG2蛋白的可溶性表达,并具有良好的完全抗原活性。

To express SAG2 gene of Toxoplasma gondii by prokaryotic expression system and to identify its antigenicity,truncated SAG2 without the highly hydrophobic signal peptide and C-terminus were amplified by PCR,and recombinant prokaryotic expression plasmid(pGEX-4T-3-SAG2t)with SAG2 tprotein gene was constructed.Then the recombinant plasmids were transferred into E.coli DH5α,and the bacteria were induced by IPTG.The expression and immune-reactivity of SAG2 tprotein were detected by SDS-PAGE and Western blotting respectively.The yield of purified rSAG2 twas more than 4 mg per liter of culture medium.Western blot showed that rSAG2 tcan be strongly recognized by mice serum infected with T.gondii ME49 strain and mice serum immunized by rSAG2 t.The result showed that the soluble expression of SAG2 tprotein with favorable complete antigenic activity was implemented by prokaryotic expression system.