MiR-148a-3p靶基因的预测与验证

目的:预测和验证miR-148a-3p的靶基因.方法:利用生物信息学方法预测miR-148a-3p的靶基因;在神经胶质瘤细胞U87/U251中建立miR-148a过表达及抑制表达稳定细胞系.利用qPCR实验验证miR-148a-3p与靶基因的靶向关系;利用Western Blot实验检测靶蛋白表达水平.结果:生物信息学方法预测结果显示dynein light chain LC8-type 2(DYNLL2)为miR-148a-3p靶基因之一;qPCR实验结果显示DYNLL2在神经胶质细胞HA1800与神经胶质瘤细胞U87/U251中的表达有差异,在神经胶质瘤细胞miR-148a过表达及抑制表达的细胞中表达有差异;Western Blot实验结果显示DYNLL2的蛋白表达水平在神经胶质细胞HA1800与胶质瘤细胞U87/U251中有差异.结论:miR-148a-3p与DYNLL2有确切的靶向关系,DYNLL2在神经胶质细胞与胶质瘤细胞中均有表达,与正常胶质细胞相比,DYNLL2在神经胶质瘤细胞中表达下调,包括基因水平和蛋白水平,miR-148a-3p能够负向调控DYNLL2的表达.

Prediction and Verification of MiR-148a-3p Target Gene

Objective To predict and verify the target gene of miR-148a-3p. Methods The putative target genes of miR-148a-3p were predicted by bioinformatics approach. Establishment of miR-148a overexpression and suppression of stable ex-pression in glioma cell line U87/U251. QPCR experiments were used to verify the targeting relationship between miR-148a-3p and target genes. The target protein expression level was detected by Western Blot. Result Bioinformatics methods predict that dynein light chain LC8-type 2(DYNLL2) is one of the miR-148a-3p target genes, QPCR experiments showed that there were differences in the DYNLL2 expression of glioma cell line U87/U251 between glial cell line HA1800, There were differences in the over expression and inhibitory expression of miR-148a in glioma cells, The results of Western Blot showed that the protein expression of DYNLL2 was different in glial cell line HA1800 and glioma cell line U87/U251. Conclusion DYNLL2 was a target gene of miR-148a-3p, DYNLL2 is expressed both in glial cells and glioma cells. Compared with normal glial cells, the expression of DYNLL2 was down regulated in glioma cells including gene levels and protein levels, and miR-148a-3p could reversely regulate DYNLL2 expression.