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纸质出版:2012
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周维,谢肇,许建中.miR-125b调节C3H10T1/2间充质干细胞中Cbfβ的表达[J].陆军军医大学学报(原第三军医大学学报),2012,34(11):1048-1051.
Zhou Wei,Xie Zhao,Xu Jianzhong.Effect of miR-125b on expression of Cbfβ in C3H10T1/2 cells[J].J Amry Med Univ (J Third Mil Med Univ),2012,34(11):1048-1051.
目的 研究miR-125b对鼠间充质干细胞株C3H10T1/2中核心结合因子-β(core-binding factor beta
Cbfβ)表达的调节作用。 方法 将microRNA阴性对照、miR-125b mimic和anti-miR-125b用脂质体转染小鼠间充质干细胞系C3H10T1/2, Western blot及RT-PCR测定细胞中Cbfβ蛋白及mRNA表达;以PicTar数据库预测的Cbfβ3′-非翻译区上miR-125b靶位点附近约30个碱基为基础合成3组插入序列(阳性对照、野生型序列、突变序列),连接到经HindⅢ、SpeⅠ双酶切后的PmiR-report载体质粒上,然后与miRNA 阴性对照和miR-125b mimic共同转染293T细胞,双荧光素酶报告基因检测荧光素酶活性。 结果 过表达miR-125b 抑制Cbfβ蛋白及mRNA表达(P0.05),转染anti-miR-125b可促进Cbfβ蛋白及mRNA表达(P0.05);miR-125b 抑制Cbfβ3′-非翻译区荧光素活性。 结论 外源性miR-125b可作用于Cbfβ3′非翻译区从而负性调节C3H10T1/2细胞中Cbfβ的表达。
Objective To study the effect of miR-125b on expression of core-binding factor beta (Cbfβ) in mouse C3H10T1/2 cells. Methods Mouse C3H10T1/2 cells were transfected with microRNA negative control
miR-125b mimics and anti-miR-125b
respectively. Expressions of Cbfβ protein and mRNA in C3H10T1/2 cells were detected by Western blotting and RT-PCR
respectively. Inserting sequences of miRNA negative control
wild type and mutation were synthesized according to the 30 bases adjacent to the miR-125b target on the Cbfβ 3′UTR in PicTar database and linked to the PmiR-report plasmids digested by HindⅢ and SpeⅠ. The 293 T cells were co-transfected with miRNA negative control and miR-125b mimics. The activity of luciferase was detected by dual luciferase report assay. Results The over-expression of miR-125b inhibited the expression of Cbfβ protein and mRNA in C3H10T1/2 cells (P0.05)
while the anti-miR-125b-transfected C3H10T1/2 cells promoted the expression Cbfβ protein and mRNA (P0.05). The miR-125b suppressed the activity of luciferase on the Cbfβ 3′UTR. Conclusion Exogenous miR-125b can act on the Cbfβ 3′UTR
and thus down-regulate the expression of Cbfβ in C3H10T1/2 cells.
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