人PPARδ受体cDNA全长序列的克隆及测序  

Cloning and sequencing of the full-length of human PPARδ cDNA

作　　者：章涛[1] 袁野[1] 李苌清[1] 杨俊卿 周元国[2] 周岐新[1] 

机构地区：[1]重庆医科大学基础医学院药理学教研室,重庆400016 [2]第三军医大学野战外科研究所分子生物学中心,重庆400042

出　　处：《重庆医科大学学报》2007年第8期789-791,795,共4页Journal of Chongqing Medical University

基　　金：国家自然科学基金资助项目(30572353);重庆医科大学博士启动基金(2004)

摘　　要：目的:对人过氧化物酶体增殖物激活受体δ(hPPARδ)全长cDNA序列进行克隆并测序,为构建含hPPAR δ基因真核表达载体及其受体功能研究奠定基础。方法:采用逆转录-聚合酶链式反应(RT-PCR)从HepG2细胞总RNA克隆hPPARδ全长cDNA序列,胶回收目的条带后与pMD19-T载体连接并转化DH5α感受态菌,用BamHI、SalI双酶切及基因测序筛选、鉴定阳性克隆pMD19-hPPARδ-T载体中hPPARa基因的完整性和忠实性。结果:经酶切和测序证实pMD19-hPPARδ-T载体中插入的hPPARδ基因序列与GeneBank中提交的序列(AY919140)一致。结论:成功克隆了hPPARδ基因,构建了pMD19-hPPARδ-T中间载体,为含hPPARδ基因真核表达载体的构建和受体功能研究打下了良好的基础。Objective：To clone and sequence the full-length of human PPAR8 cDNA in order to further construct the eukaryotic expression vector carrying hPPAR8 gene. Methods：hPPAR8 cDNA was cloned from HepG2 cells total RNA by RT-PCR. The PCR product recovered from gel were ligated with pMD19-T vector and transformed into DH5ct competent cell. The integrity and fidelity of hPPAR8 cDNA sequence inserted in T vector were verified by BamH Ⅰ and So/ Ⅰ double excising and DNA sequencing assays. Results：The positive clone T vector plasmid containing correct sequence of hPPAR8 cDNA were verified by enzyme digestion as well as sequence analysis and was named as pMD19-hPPARB-T vector. The sequence of inserted hPPAR8 cDNA was in accordance with the corresponding sequence in GeneBank database（AY919140）. Conclusion： hPPAR8 gene is successfully cloned and the pMD19-hPPARB-T intermediate vector is constructed,which provide a good basis for further constructing the eukaryotic expression vector carrying hPPAR 8 gene and studying the receptor＇s function.

关 键 词：过氧化物酶体增殖物激活受体Δ RT-PCR CDNA克隆 T载体 测序 

分 类 号：R383.24[医药卫生—医学寄生虫学]