胃腺癌细胞系MGC803中miR-23a靶基因IL-6R的鉴定

Identifcation of the miR-23a targeted gene-IL-6R in gastric adenocarcinoma cell line MGC803

目的建立双荧光蛋白报告基因分析系统,并用来验证miR-23a的靶基因白细胞介素-6受体(IL-6R)。方法选取表达绿色荧光蛋白的质粒pcDNA3/EGFP,将IL-6R3′UTR的一段特异性序列插入该质粒中,并与miR-23a及表达红色荧光蛋白质pDsRed2-N1共同转染胃腺癌细胞系MGC803,转染后的细胞和提取的蛋白样品,分别用荧光显微镜和荧光分光光度计进行定性和定量检测。结果共转miR-23a和pcDNA3/EGFP-IL-6R3′UTR质粒后,绿色荧光蛋白的表达量明显低于pcDNA3和pcD-NA3/EGFP-IL-6R3′UTR共转组。结论研究表明IL-6R可能是miR-23a的直接靶基因。

Purpose To establish a dual fluorescent protein reporter assay system, and to identify the miR-23a targeted gene-IL-6R using this method. Methods A sequence of IL-6R3 UTR （ untranslated region） was inserted into the plasmid which expressed green fluorescent protein （pcDNA3/EGFP）. This plasmid （pcDNA3/EGFP-IL-6R3 UTR） and miR-23a and the plasmid expressed red fluorescent protein （ pDsRed2-N1 ） were cotransfeeted into MGC803 cells. The cells and the extracted protein had been detected under fluorescence microscope and fluorescence spectrophotometer, respectively. Results After miR-23a and the plasmid of pcDNA3/EGFP-IL-6R3 UTR being cotransfected, the intensity of green fluorescent protein was significantly lower than that group of eotransfeeted peD-NA3/EGFP-IL-6R3UTR with pcDNA3. Conclusion This study demonstrates that IL-6R may be a direct target gene of miR-23a.