MiR-148a-3p靶基因筛选及验证

Screening and verification of miR-148a-3p target gene

目的 :筛选miR-148a-3p的靶基因,并进行验证。方法 :利用生物信息软件miRanda、TargetScan和miRBase筛选miR-148a-3p潜在靶基因;运用荧光定量PCR检测靶基因在正常胶质细胞与神经胶质瘤与mRNA表达水平,并在miR-148a不同表达条件下各神经胶质瘤细胞株中靶基因的mRNA表达水平;再利用荧光素酶实验对靶基因检验。结果 :生物信息学网站预测ST18为miR-148a-3p靶基因;RT-PCR结果显示神经胶质瘤细胞系(U87、U251、T98G)中ST18的mRNA表达水平较正常胶质细胞低;过表达miR-148a的胶质瘤细胞中ST18mRNA与对照组比较表达量明显下调;抑制miR-148a表达的胶质瘤细胞中ST18 mRNA与对照组比较表达量明显增加;荧光素酶实验结果显示ST18WT+mimics NC与ST18WT+Mir-148a比较,ST18WT+Mir-148a荧光强度明显下降,有统计学差异。结论 :ST18是miR-148a-3p的靶基因。

Objective Screening and validation of miR-148 a-3 p target genes.Methods Bioinformatics software(Targetscan,miRanda and miRBase)was used to predict the target genes of miR-148 a-3 p;Fluorescence quantitative PCR was used to compare the mRNA expression level of target genes in glioma and normal glioma cells,and to detect the expression level of target genes in glioma cell lines under MiR-148 a over expression and inhibition.Detection of target genes by luciferase test.Results ST18 is one of MiR-148 a-3 p target genes;RT-PCR results showed that the expression level of MRNA in ST18 in glioma cell lines(U87,U251,T98 G)was lower than that in normal glioma cells,and in glioma cell lines(U87,U251);The expression of ST18 mRNA in glioma cells with hyperexpression miR-148 a was significantly reduced in the control group;The expression of ST18 mRNA in glioma cells inhibited by miR-148 a increased significantly compared with control group;Fluorescent enzyme experimental results show that compared with ST18 WT+mimics NC and ST18 WT+Mir-148 a,the fluorescence intensity of ST18 WT+Mir-148 a decreased significantly,with a statistical difference.Conclusion ST18 is the target gene for iR-148 a-3 p.