副溶血弧菌tlh基因的克隆、表达及功能的研究

目的　构建融合表达载体pET32a+-tlh ,获得具有生物活性功能的蛋白。方法　用PCR法扩增tlh全序列 ,克隆入 pET32a +高效表达载体进行诱导表达 ,亲和层析纯化目的蛋白 ,采用逐步透析的复性方法 ,并对复性产物进行功能检测。结果与结论　成功构建了融合表达载体pET32a +-tlh ,在大肠杆菌DE3 中表达产物以包涵体形式存在 ,复性后的蛋白在卵磷脂存在下具有溶血活性。

Molecular cloning,expression and functions of the tlh gene of Vibrio parahaemolyticus

To construct the expression vector of pET32a -tlh to obtain the protein with biological activities,the complete sequence of the tlh gene of Vibrio parahemolyticus was amplified by PCR and cloned into high efficient expression vector pET32a The expression product was purified by using an affinity chromatographic method and renatured by gradient dialysis.The expression vector pET32a -tlh was successfully constructed by the above mentioned methods and the gene product was expressed in E.coli DE3 as an inclusion body.The renatured protein was activated in the addition of phosphatidycholine and showed the hemolytic activity.