RNA干扰诱导DNA修复基因hMGMT表达沉寂的研究

doi:10.3969/j.issn.1004-616x.2006.01.003

癌变·畸变·突变 ›› 2006, Vol. 18 ›› Issue (1): 9-011.doi: 10.3969/j.issn.1004-616x.2006.01.003

RNA干扰诱导DNA修复基因hMGMT表达沉寂的研究

赖延东;吕嘉春;李秀英;宾晓农;吴建军

广州医学院化学致癌研究所，广东广州 510182

收稿日期:2005-03-10 修回日期:2005-05-20 出版日期:2006-01-30 发布日期:2006-01-30
通讯作者: 吕嘉春

Study of RNA Interference Induced Expression Silence of DNA Repaired Gene hMGMT

LAI Yan-dong;LU Jia-chun;LI Xiu-ying;BIN Xiao-nong;WU Jian-jun

Institute for Chemical Carcinogenesis, Guangzhou Medical College, Guangzhou 510182,Guangdong, China

Received:2005-03-10 Revised:2005-05-20 Online:2006-01-30 Published:2006-01-30
Contact: LU Jia-chun

摘要/Abstract

摘要： 背景与目的：构建能特异诱导人DNA修复基因hMGMT表达沉寂的RNA干扰载体。材料与方法：通过两步法聚合酶链反应（polymerase chain reaction，PCR）扩增hMGMT特异RNA干扰表达盒，连入质粒载体构建RNA干扰质粒pU6-MGMTi，与pGEFP-C1共转染人支气管上皮16HBE（Human bronchial epithelial）细胞，G418筛选后，采用RT-PCR和Western Blot分别检测mRNA和蛋白水平的表达抑制情况。结果：成功构建hMGMT特异RNA干扰质粒pU6-MGMTi，转染16HBE细胞的mRNA和蛋白表达受到显著的抑制。结论：应用RNA干扰表达盒以及共转染技术，构建MGMT基因表达沉寂的细胞株，为进一步阐明MGMT基因的功能创造条件。

关键词: hMGMT基因, RNA干扰, 16HBE

Abstract: BACKGROUND & AIM: To construct the RNA interference vector which can specially induce the expression silence of human DNA repair gene hMGMT. MATERIAL AND METHODS: The hMGMT specific siRNA expression cassette was made by two steps PCR, and then linked with pUC19 to get pU6-MGMTi. Co-transfected with pEGFP-C1 into 16HBE and screened by G418. The expression level of mRNA and protein was detected by RT-PCR and Western Blot respectively. RESULTS: hMGMT specific RNA interfere vector pU6-MGMTi was constructed successfully. Transfected 16HBE cell’s MGMT mRNA and protein expression level were dramatically suppressed. CONCLUSION: MGMT expression silence cell line was built by RNA interfere expression cassette and co-transfection technology, which offered condition for studying the gene function of MGMT.

Key words: hMGMT gene, RNA interfere, 16HBE

中图分类号:

赖延东, 吕嘉春, 李秀英, 宾晓农, 吴建军. RNA干扰诱导DNA修复基因hMGMT表达沉寂的研究[J]. 癌变·畸变·突变, 2006, 18(1): 9-011.

LAI Yan-dong, LU Jia-chun, LI Xiu-ying, BIN Xiao-nong, WU Jian-jun. Study of RNA Interference Induced Expression Silence of DNA Repaired Gene hMGMT[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2006, 18(1): 9-011.

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RNA干扰诱导DNA修复基因hMGMT表达沉寂的研究

Study of RNA Interference Induced Expression Silence of DNA Repaired Gene hMGMT

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