布鲁菌wzt基因的原核表达及生物信息学分析

Prokaryotic expression and bioinformatics analysis of Brucella wzt gene

利用PCR技术克隆wzt基因并对其进行了原核表达,采用SDS-PAGE和Western Blotting进行了检验,运用Clustalx1.83和Mega5.0进行了同源性分析,在此基础上,利用Discovery Studio 2.5软件进行了结构预测,探讨了wzt基因表达产物的分子基本特征.结果表明:本研究克隆了756bp的目的片段,经原核表达得到了相对分子质量约28 000的目的蛋白,该蛋白经HisTrapTMFF亲和层析柱纯化以及Western Blotting鉴定,证明为具有His标签的外源蛋白;系统树分析表明,布鲁菌wzt蛋白属内高度保守;结构分析表明,其具有8个α螺旋、7个β折叠片层结构域.

In this paper, the wzt gene was cloned by PCR method, and linked to the pEASY-E1, and thenexpressed in E. coli and detected by SDS-PAGE and western blotting. The predicted amino chainsequence polymorphism was analyzed by Clustalxl. 83 and MegaS. 0 homology analysis,and structureprediction was by using Discovery Studio 2.5 software for the characteristics of product of wzt gene.The results showed that 756 bp fragment was cloned and relative molecular 28 000 proteins wasexpressed. The protein was purified by HisTrapTM FF affinity chromatography and the protein wasidentified by Western Blotting. Polymorphism analysis showed that the wzt protein was highlyconserved,and the protein was with eight a helices and seven pleated sheet regions.