旋毛虫肌幼虫ES抗原的基因克隆及高效表达

Cloning and High Level Expression of Gene Encoding ES Antigen from Trichinella spiralis Muscle Larvae

作者对编码旋毛虫肌幼虫ES抗原的部分结构基因进行了克隆、鉴定和表达。用RNA PCR技术直接从旋毛虫肌幼虫总RNA中反转录并扩增出0.7kb的靶DNA,酶切分析后将其克隆到融合表达载体pEX31C中。SDS—PAGE电泳表明,含重组子的大肠杆菌能够表达出一分子量为37kDa的融合蛋白(P37),后者占菌体总蛋白的22％以上,并以包含体形式存在于菌体中。经对纯化后表达蛋白的ELISA检测,证明它能被猪旋毛虫病阳性血清和抗旋毛虫单克隆抗体识别。研究结果揭示,重组蛋白P37对于研制旋毛虫病诊断抗原和免疫抗原具有潜在的应用价值。

The partial structure gene encoding ES antigen derived from T. spiralis (TSP) muscle larvae was cloned,characterized,and expressed in E. colt. The target DNA (0. 7kb) was directly obtained by using RNA PCR technique from the TSP total RNA. Analysed it with the RE digestion, the fragment was cloned into the fusion expression vector pEX31C. It was shown that a kind of 37 kDa fusion proteins was expressed in E. colt containing the recombinant plasmid by SDS-PAGE electrophoresis. The expressed . proteins was over 22% of the total cell protein and it was aggregated in the form of inclusion bodies in E. coll. The purified protein could be recognized in ELISA both by sera from swine-infected with TSP and by the monoclonal antibody against TSP. These findings suggest that the recombinant protein is a potentially valuable antigen both for im-munodiagnosis and vaccine development of trichinellosis. This is the first demonstration of cloning and high level expression for structure gene encoding ES antigen in our country.