CAJ | 학술논문

目的:分析miR-25在胃癌组织中的表达水平并鉴定其新的靶基因.方法:选取10例胃癌患者的胃癌及癌旁正常组织样本,用qRT-PCR法检测miR-25的表达水平.利用miRNA靶基因预测数据库预测miR-25的靶基因.构建荧光素酶载体(pMIR-TOB1)及绿色荧光蛋白报告载体(pMIR-GFP-TOB1)(含有miR-25结合位点的TOB1的3'UTR片段),利用荧光素酶活性实验和GFP荧光强度报告实验鉴定miR-25的预测靶基因(TOB1).qRT-PCR法和Western blot法分别检测TOB1的mRNA和蛋白质的表达水平.结果:miR-25在80%(8/10)的胃癌组织样本中表达上调.miR-25模拟物显著抑制了荧光素酶的活性(P<0.01)和GFP荧光强度.miR-25模拟物显著下调了AGS细胞中TOB1的mRNA(P<0.05)和蛋白质的表达.结论:miR-25在人胃癌组织中表达上调,miR-25能结合TOB1的3'UTR并下调TOB1的mRNA和蛋白质的表达.
목적:분석miR-25재위암조직중적표체수평병감정기신적파기인.방법:선취10례위암환자적위암급암방정상조직양본,용qRT-PCR법검측miR-25적표체수평.이용miRNA파기인예측수거고예측miR-25적파기인.구건형광소매재체(pMIR-TOB1)급록색형광단백보고재체(pMIR-GFP-TOB1)(함유miR-25결합위점적TOB1적3'UTR편단),이용형광소매활성실험화GFP형광강도보고실험감정miR-25적예측파기인(TOB1).qRT-PCR법화Western blot법분별검측TOB1적mRNA화단백질적표체수평.결과:miR-25재80%(8/10)적위암조직양본중표체상조.miR-25모의물현저억제료형광소매적활성(P<0.01)화GFP형광강도.miR-25모의물현저하조료AGS세포중TOB1적mRNA(P<0.05)화단백질적표체.결론:miR-25재인위암조직중표체상조,miR-25능결합TOB1적3'UTR병하조TOB1적mRNA화단백질적표체.
Objective:This study aims to analyze the miR-25 expression profile in gastric cancer (GC) tissue and identify its novel target gene. Methods:Ten paired GC tissues and adjacent normal tissues were included in this study. MiR-25 expression levels were measured via quantitative reverse transcription polymerase chain reaction (qRT-PCR). The target genes of the miR-25 were predicted in the miRNA Target Database. Luciferase (pMIR-TOB1) and GFP reporters (pMIR-GFP-TOB1), which contain the binding site of miR-25 to TOB1 3'UTR, were constructed. The predicted target gene (TOB1) of miR-25 was identified via luciferase activation and GFP expression assays. The mRNA or protein expression of TOB1 was detected via qRT-PCR or Western blot, respectively. Results:Increased expression of miR-25 was observed in eight GC tissues (80%). miR-25 mimics significantly inhibited luciferase activation (P<0.01) and GFP expression in the HEK293 cells, and significantly decreased the mRNA (P<0.05) and protein expression levels of TOB1 in the AGS cells. Conclusion:miR-25 is significantly increased in human GC tissue. Moreover, MiR-25 can significantly repress the mRNA and protein expression of TOB1 by directly targeting 3'UTR.