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【目的】观察小分子干扰RNA(siRNA)沉默细胞周期素依赖性蛋白激酶(CDK2)基因后,细胞周期相关基因RB、CyclinE、E2F1在肝癌细胞SMMC7721中mRNA的表达。【方法】将前期研究中已构建成功并筛选出的最有效干扰抑制CDK2基因的siRNA序列片段,采用Lipofectamine TM2000脂质体转染法转染肝癌细胞株SMMC7721后分6组:重组质粒组190、重组质粒组191、SMMC7721肝癌组、转染试剂组、阴性对照组、空质粒组。采用实时荧光定量PCR法检测RB、CyclinE、E2F1 mRNA水平。【结果】CDK2的siRNA转染SMMC7721细胞后,RB基因mRNA表达上调,CyclinE、E2F1基因mRNA表达下调。【结论】抑制CDK2基因的表达能使肝癌细胞SMMC7721中RB基因mRNA表达上调,CyclinE、E2F1基因mRNA表达下调,提示肝癌细胞中RB、CyclinE、E2F1基因的表达与CDK2基因的表达具有明显的相关性。
【Objective】 To observe the mRNA expression of cell cycle related genes RB, CyclinE and E2F1 in SMMC7721 hepatocellular carcinoma cells after silencing cyclin-dependent protein kinase (CDK2) by small interfering RNA (siRNA). 【Methods】 The siRNA fragment of CDK2 gene, which had been successfully constructed and screened in the previous study, was successfully cloned and transfected into SMMC7721 cell line by lipofectamine TM 2000 lipofectamine transfection method. The recombinant plasmid group 190, Recombinant plasmid group 191, SMMC7721 liver cancer group, transfection reagent group, negative control group, empty plasmid group. Real-time fluorescence quantitative PCR was used to detect the mRNA levels of RB, CyclinE and E2F1. 【Results】 The results showed that mRNA expression of RB gene was up-regulated in CDK2 siRNA transfected SMMC7721 cells, and mRNA expression of CyclinE and E2F1 was down-regulated. 【Conclusion】 Inhibition of CDK2 gene expression can upregulate the expression of RB gene mRNA and the expression of CyclinE and E2F1 mRNA in hepatocellular carcinoma cell line SMMC7721, which indicates that the expression of RB, CyclinE and E2F1 gene in hepatocellular carcinoma cells is significantly correlated with the expression of CDK2 gene .