【摘要】 目的研究人肝再生增强因子(ALR)对细胞周期素依赖性激酶1启动子(CDK1p)转录调节的作用,探讨ALR的生物学作用机制。方法根据GenBank中CDK1p序列的分析设计引物,应用聚合酶链反应(PCR)扩增人CDK1基因启动子基因序列,并克隆至真核报告载体pCAT3-Basic中,构建CDK1启动子报告基因表达载体pCAT3-CDK1p,以该质粒转染HepG2细胞系,并同时与pcDNA3.1(-)-ALR共转染HepG2细胞系,用酶联免疫吸附法(ELISA)检测报告基因氯霉素乙酰转移酶(CAT)的表达活性。结果 pCAT3-CDK1p组CAT表达活性明显高于pCAT3-basic组,差异有统计学意义,pCAT3-CDK1p与pcDNA3.1(-)-ALR共转染组的CAT表达活性明显低于pCAT3-CDK1p组,差异有统计学意义。结论 CDK1启动子有顺式激活下游基因的活性,ALR对CDK1基因有下调作用。更多还原

【Abstract】 Objective To investigate the effects of human augmenter of liver regeneration (ALR) on the transcription of cyclin-dependent kinase 1 gene promoter (CDK1p), and to explore the biological function mechanism of ALR. Methods Polymerase chain reaction (PCR) technique was employed to amplify the sequence of CDK1 promoter from HepG2 genomic DNA, named CDK1p, and the PCR product was cloned into pCAT3-basic, named pCAT3-CDK1p. The HepG2 cells were transfected with pCAT3-CDK1p, and then co-transfected with pCAT3-CDK1p and pcDNA3.1(-)-ALR. The chloramphenicol acetyltransferase (CAT) activity was detected by an enzyme linked immunosorbent assay (ELISA) kit. Results The CAT activity of the HepG2 transfected by pCAT3-CDK1p was significantly higher than that of the negative control group. The CAT activity of the HepG2 co-transfected by pCAT3-CDK1p and pcDNA3.1(-)-ALR was significantly lower than that of the HepG2 transfected by pCAT3-CDK1p. Conclusions The CDK1p has transcription activity. It is suggested that ALR can down-regulate CDK1 gene.更多还原