【摘要】 目的预测并鉴定miR-138的靶基因人端粒酶反转录酶(human telomerase reverse transcriptase,hTERT)。方法采用生物信息学技术,预测miR-138与hTERT 3’UTR结合位点及结合后稳定性。分别采用RTqPCR、Western blot技术检测过表达miR-138后MKN-45细胞内hTERT在mRNA与蛋白水平的变化。采用双荧光素酶实验及突变实验,验证miR-138与hTERT mRNA的结合位点。结果生物信息学技术预测显示,miR-138可稳定结合于hTERT 3’UTR。miR-138过表达后可明显抑制MKN-45细胞内hTERT蛋白表达水平(P<0.01),但对hTERT mRNA水平无显著影响(P>0.05)。双荧光素酶实验证实miR-138可稳定结合于hTERT 3’UTR相应位点。结论 miR-138通过直接结合于靶基因hTERT 3’UTR抑制hTERT蛋白表达。miR-138可能成为基于miRNA的肿瘤基因治疗的新靶标。更多还原

【Abstract】 Objective To predict and identify the target gene human telomerase reverse transcriptase( hTERT) of miR-138. Methods The bioinformatics analysis was employed to predict miR-138 binding sites with hTERT 3’UTR. RT-qPCR and Western-blot were used to examine the inhibitory effect on hTERT in human gastric carcinoma MKN-45 cells by miR-138. Furthermore,the binding site of miR-138 to hTERT 3’ UTR was examined using dual luciferase reporter assay. Results The bioinformatics analysis showed stable binding site of miR-138 and hTERT 3’UTR. Over-expression of miR-138 in MKN-45 cells significantly suppressed the expression of hTERT protein( P < 0. 01),but not hTERT mRNA( P > 0. 05). The dual luciferase reporter assay verified the binding site of miR-138 to hTERT 3’UTR. Conclusion miR-138 can stably bind to hTERT 3’UTR and suppress the hTERT expression,which makes miR-138 a promising target in cancer gene therapy.更多还原