中国药理学通报2012，Vol.28Issue(7)：1023-1026,4.DOI:10.3969/j.issn.1001-1978.2012.07.032
PPARγ基因靶向shRNA真核表达载体的构建及表达
Construction and expression of eukaryotic expression plasmids of shRNA targeting the PPARγ gene
摘要
Abstract
Aim To construct eukaryotic expressing plasmids of short hairpin RNA ( shRNA) targeting the PPAR γ gene and to evaluate their inhibitory effect on PPARγ expression in human umbilical veinendothelial cells ( HUVECs). Methods Three pairs of complementary shRNA oligonucleotides targeting the PPARγ gene were designed , synthesized, annealed and inserted into the pGPU6/GFP/Neo plasmid. The recombinant plasmids were identified by restriction enzyme analysis and sequence anal -ysis. The inhibitory effect of recombinant plasmids on PPAR γ expression in HUVECs was detected by Western blot. ResultsAfter restriction enzyme analysis and sequence analysis , three eukaryotic expression plasmids of shRNA targeting the PPAR γ gene were successfully constructed. Western blot analysis showed that pGPU6/GFP/NeoshRNAPPARγ3 reduced PPARγ expression by 63.2% in HUVECs induced with high glucose. Conclusion Three eukaryotic expression plasmids of shRNA targeting the PPARγ gene are successfully constructed. These recombinant plasmids can efficiently inhibit PPARγ expression in HUVECs.关键词
过氧化物酶体增殖物激活受体γ/胰岛素抵抗/短发夹RNA/RNA干扰/真核表达载体/糖尿病/磷脂酰肌醇3激酶Key words
PPARs/ insulin resistance/ small hairpin RNA/ RNA interfereence/ recombinant plasmid/diabetes/PI3K分类
医药卫生引用本文复制引用
高艳,占日新,刘丽丽,陈芳辉,李宙雪,张盈,孔滢,黄起壬..PPARγ基因靶向shRNA真核表达载体的构建及表达[J].中国药理学通报,2012,28(7):1023-1026,4.基金项目
国家自然科学基金资助项目(No 81070633) （No 81070633）
江西省研究生创新专项资金资助项目(No YC 09A012) （No YC 09A012）
