布鲁菌Omp31基因的克隆、表达及其蛋白的生物信息学分析  

Cloning,expression of the Omp31 gene and bioinformatics analysis of its protein of Brucella melitensis

作　　者：郭莳雨[1] 成鹰[1] 贾晓晓[1] 史巧芸[1] 荣辉[1] 张珈宁[1] 朱华培[1] 杜丽[1] 焦寒伟[1] 王凤阳[1] 

机构地区：[1]海南大学农学院海南省热带动物繁育与疫病研究重点实验室/海口市动物基因工程重点实验室,海口570228

出　　处：《黑龙江畜牧兽医》2013年第9期8-10,共3页Heilongjiang Animal Science And veterinary Medicine

基　　金："十二五"农村领域国家科技计划课题项目(2011AA100302)

摘　　要：为了进一步研究Omp31蛋白的功能,试验利用DNAMan软件设计上、下游引物,通过PCR技术从马耳他布鲁菌M5-90株中克隆获得Omp31基因,构建原核表达重组质粒pET28a-Omp31,并转化E.coli BL21(DE3)感受态;再以IPTG诱导重组蛋白表达,进行SDS-PAGE和Western-blot检测,并运用生物信息学软件对所获得的核苷酸序列进行分析。结果表明:成功克隆了马耳他布鲁菌外膜蛋白Omp31基因,并构建出重组质粒pET28a-Omp31;经IPTG诱导,得到与预期大小相符的目的蛋白;生物信息学分析发现,Omp31蛋白核苷酸序列包括1个723 bp的开放读码框,编码240个氨基酸;该蛋白二级结构中存在大量无规则卷曲和少量α螺旋。To further develop the research for the function of Omp31 protein, the DNAMan software was used to design the up - and downstream primers, and then the outer membrane protein 31 （Omp31 ） gene was amplified and obtained from Brucella melitensis M5 -90 by PCR. The 0rap31 gene was ligated with pET28a to construct the recombinant expression plasmid pET28a - Omp31, and then transformed into the E. coli BL21 （DE3） strain. The recombinant expression was induced by IPTG, and the products were analyzed by SDS -PAGE and Western -blot. The obtained nucleotide sequence was analyzed by the bioinformatics software. The results showed that Brucella melitensis Omp31 gene was cloned, the expression plasmid pET28a - Omp31 was successfully constructed. The target protein was obtained with a molecular weight same as the expected size using IPTG induction. Based on the biolnformatics analysis, it was found that the nucleotide sequence of the Omp31 protein in- cluded a 723 bp of the open reading frame encoding 240 amino acids, there were many random coils and few α - helices in the secondary structure of the protein.

关 键 词：布鲁菌Omp31 克隆 原核表达 生物信息学分析 

分 类 号：S852.61[农业科学—基础兽医学]