绿色荧光蛋白标记的人脂联素真核表达载体的构建  

Construction and Identification of Eukaryotic pcDNA3.1/ CT- GFP-TOPO-Adiponectin Expression Vector

作　　者：刘德敏[1] 杜春红[1] 孙颖[1] 张捷[1] 

机构地区：[1]天津医科大学代谢病医院,300070

出　　处：《天津医药》2007年第5期324-325,I0003,共3页Tianjin Medical Journal

基　　金：天津市教委基金项目(项目编号:2003025);天津市自然科学基金资助项目(项目编号:043609211)

摘　　要：目的:构建绿色荧光蛋白(GFP)标记的人脂联素(Adiponectin)真核表达质粒。方法:用本实验室已构建好的人脂联素克隆载体pMD18-T/Adiponectin为模板,结合已设计好的特异性引物,采用PCR方法克隆出Adiponectin目的片段,将该目的片段再连至真核表达载体pcDNA3.1/CT-GFP-TOPO上,转化入TOP10感受态细胞中,通过筛选得到融合有绿色荧光蛋白的重组质粒pcDNA3.1/CT-GFP-TOPO-Adiponectin。结果:PCR获得长度为736bp的目的片段,经pcDNA3.1/CT-GFP-TOPO真核表达载体连接、筛选及序列分析后,证实所插入的目的片段与genebank检索的人脂联素cDNA序列(AccessionNM-004797)100%匹配。结论:绿色荧光蛋白标记的人脂联素真核表达载体构建成功。Obiective： To construct the pcDNA3.1/CT -GFP-TOPO-Adiponectin expression vector. Methods： The encoding fragment of human Adiponectin gene was obtained from a recombinant plasmid pMD18-T/Adiponectin using PCR, then the gene was inserted into the vector pcDNA3.1/CT-GFP-TOPO after sequencing, which introduced into TOP10 competent ceils in order to choose and sequence, Results： A 736 bp fragment was obtained by PCR clone after bloting and sequencing. The results were matching to the cDNA of human adiponectin, which accession of the gene bank was NM-004797. Conclusion： The eukaryotic pcDNA3.1/CT-GFP-TOPO- Adipon ectin expression vector was constructed successfully.

关 键 词：脂类 遗传载体 质粒 绿色荧光蛋白质类 聚合酶链反应 

分 类 号：R512.6[医药卫生—内科学]