人瘦素基因的克隆及在酿酒酵母中的表达和生物信息学分析  

Cloning and Expression and Bioinformatic Analysis of Human Leptin Protein in Saccharomyces Cerevisia

作　　者：韩晋云[1] 郑文岭[1,2] 宋艳斌[1] 姜茂竹[1] 王妮莎[1] 马文丽[1] 

机构地区：[1]南方医科大学基因工程研究所,广州510515 [2]华南基因组研究中心,广州510800

出　　处：《热带医学杂志》2010年第5期516-519,F0002,共5页Journal of Tropical Medicine

基　　金：广东省重点实验室基金(No.960327)

摘　　要：目的构建人瘦素基因和蛋白转导肽Tat基因的融合表达载体,转入酿酒酵母表达系统进行诱导表达,并对融合瘦素蛋白进行生物信息学分析。方法脂肪组织中提取总RNA,逆转录为cDNA,以此cDNA为模板扩增瘦素基因,构建重组质粒pYES2-Tat-leptin,转入酿酒酵母表达菌株INVSc1诱导表达Tat-leptin融合蛋白,利用相关在线软件对融合瘦素蛋白相关的生物学功能进行分析。结果经SDS-PAGE电泳和Western blotting鉴定,酿酒酵母成功表达融合蛋白Tat-leptin。经生物信息学分析后获得了融合瘦素蛋白的相关生物学特性。结论 Tat-leptin融合蛋白在酿酒酵母中成功表达,并对融合瘦素蛋白的生物学特征进行了预测,为消化道基因治疗肥胖的研究奠定了基础。Objective To construct recombinant yeast plasmid harbouring fusion gene Tat-Leptin,and express the recombinant leptin protein in Saccharomyces cerevisia,and conduct bioinformatic analysis of recombinant leptin protein. Methods RNA isolated from adipose tissue was reverse transcribed to cDNA,and then leptin gene was amplified by PCR. The recombinant plasmid pYES2-tat-leptin was constructed,was and transfected into INVSc1 and induced by galactose to express the fusion protein. The bioinformatic analysis of the fusion protein was conducted with the help of related online software. Results The Saccharomyces cerevisia expressed recombinant human leptin protein,which could be identified by SDS-PAGE and Western blot. Related biological characteristics of recombinant leptin were obtained after the online software analysis. Conclusion The recombinant protein was successfully expressed in Saccharomyces cerevisia. It＇s biological characteristics were predicted. This study lays foundation for further studies on gene therapy targeting alimentary tract for obesity.

关 键 词：瘦素 酿酒酵母 消化道 基因治疗 

分 类 号：Q786[生物学—分子生物学]