爪蟾卵母细胞中PPARγ-PPRE调控系统的建立和应用  

Studies on a PPARγ-PPRE regulatory systerm in xenopus oocyte and its application

作　　者：严瑾 沃兴德[1] 范春雷[1] 高丽萍[1] 钱颖[1] 宋家梁[1] 

机构地区：[1]浙江中医学院分子医学研究所

出　　处：《中国药学杂志》2005年第7期550-553,共4页Chinese Pharmaceutical Journal

基　　金：国家自然科学基金资助项目(30070932)

摘　　要：目的在爪蟾卵母细胞中建立人类PPARγ-PPRE调控系统,为PPARγ配体类药物筛选提供一个可靠的实验平台方法将调节蛋白PPAR基因构建在爪蟾卵母细胞启动子XOE下游,构成pXOE-PPARγ重组质粒;将PPRE反应元件构建在增强型绿色荧光蛋白d2EGFP报告基因的上游,得到pPPRE-d2EGFP重组质粒。将pXOE-PPARγ质粒和pPPRE-d2EGFP质粒共注射入爪蟾Ⅴ,Ⅵ期卵母细胞中,建立PPRE调控通路的表达系统,分别在含前列腺素E1(PGE1)、吡咯列酮的培养液中培养3d,对照组共注射pXOE-PPARγ质粒和pTAL-d2EGFP质粒,用荧光显微镜观察绿色荧光蛋白的表达情况。结果在含PGE1,吡咯列酮的卵母细胞中观测到绿色荧光蛋白;对照组未见到荧光。结论PGE1,吡咯列酮可以通过影响PPRE反应元件上调其下游基因的表达;通过核内共注射的方法在爪蟾卵母细胞中建立的PPARγ-PPRE调控系统。OBJECTIVE: To seek for a new cellular model for the quick selection of PPARγ ligand, and to establish the PPARγ-PPRE pathway regulatory system in xenopus oocytes. METHODS: PPARγ expression vectors pXOE-PPARγ was constructed by inserting the human PPARγ-cDNA into the pET17B-XOE vector. The d2EGFP gene was constructed in the downstream of PPRE promoter, and a plasmid named pPPRE-d2EGFP was obtained. The two plasmids (pXOE-PPARγ, pPPRE-d2EGFP) were injected simultaneously into the nucleus of the Xenopus oocytes. The oocytes were treated with prostaglandin E1, pioglitazone, respectively. In control group, two plasmids (pXOE-PPARγ, pTAL-d2EGFP) were co-injected into the nucleus of the oocytes. RESULTS: The expression of d2EGFP was induced by Prostagalandin E1, Pioglitazone, and no expression of d2EGFP was detected in the control group. CONCLUSION: A PPARγ-PPRE regulation system was developed in xenopus oocytes for quick selection of drugs.

关 键 词：PPRE PPARΓ 爪蟾卵母细胞 绿色荧光蛋白 基因表达 

分 类 号：Q813[生物学—生物工程]