以PRDM1基因启动子为靶点的药物筛选模型的建立研究

发布时间：2018-06-21 15:49

本文选题：PRDM基因 + 双荧光素酶报告基因　；参考：《重庆医学》2017年11期

【摘要】：目的以PRDM1基因启动子为靶点构建双荧光素酶报告基因载体,建立体外药物筛选细胞模型,并对中草药小分子化合物进行筛选。方法将人PRDM1基因启动子序列(267、1 257bp)克隆入荧光素酶报告基因载体pGL3-Basic中,构建重组质粒pGL3-PRDM1并与内参质粒pRL-TK瞬时共转染工具细胞,通过检测荧光素酶报告基因表达水平的变化反映PRDM1基因启动子的启动转录活性,并对共转染质粒比例、工具细胞选择等条件进行探索和优化,相关药物处理进行验证。结果成功构建了荧光素酶报告载体pGL3-PRDM1。用0.5、1、2、4μmol/L的5-Aza-CdR处理重组的U266细胞筛选模型,与0μmol/L 5-Aza-CdR相比,增加5-Aza-CdR的刺激,荧光强度及PRDM1基因启动子的活性呈剂量依赖性增强。与0μmol/L相比,青蒿琥酯从5μmol/L浓度开始对PRDM1基因启动子活性呈明显降低(P0.05),在20μmol/L浓度时降到最低;白芍总苷呈现小剂量促进PRDM1启动子活性,高剂量抑制PRDM1基因启动子活性。青蒿琥酯对细胞生长增殖影响较小,白芍总苷对细胞生长增殖的影响较复杂。结论成功建立了以PRDM1基因启动子为靶点的药物筛选模型,并筛选出青蒿琥酯能抑制PRDM1基因启动子活性。
[Abstract]:Objective to construct a double luciferase reporter gene vector with PRDM1 gene promoter as the target, to establish a drug screening cell model in vitro and to screen small molecular compounds of Chinese herbal medicine. Methods the promoter sequence of human PRDM1 gene was cloned into luciferase reporter gene vector pGL3-basic. The recombinant plasmid pGL3-PRDM1 was cotransfected with pRL-TK. The change of luciferase reporter gene expression level reflected the priming transcriptional activity of PRDM1 gene promoter, and the conditions of co-transfection plasmid ratio and tool cell selection were explored and optimized, and the related drug treatment was verified. Results the luciferase report vector pGL3-PRDM1 was successfully constructed. Compared with 0 渭 mol / L 5-Aza-CdR, 5-Aza-CdR increased 5-Aza-CdR stimulation, fluorescence intensity and PRDM1 promoter activity in a dose-dependent manner. Compared with 0 渭 mol / L, artesunate decreased the activity of PRDM1 gene promoter from 5 渭 mol / L to the lowest at 20 渭 mol / L. Total glucoside of Paeoniae Paeoniae Paeoniae Alba increased PRDM1 promoter activity at low dose and inhibited PRDM1 promoter activity at high dose. Artesunate had little effect on cell growth and proliferation, while total glucosides of paeony had more complex effects on cell growth and proliferation. Conclusion A drug screening model targeting PRDM1 gene promoter was successfully established and artesunate could inhibit the activity of PRDM1 gene promoter.
【作者单位】：郑州大学第一附属医院检验科;河南中医学院第一附属医院检验科;
【基金】：河南省卫生厅计划指导项目(201304015) 河南省教育厅科技研究重点项目(14A320026) 郑州市科技局资助项目(141PPTGG437)
【分类号】：R96

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