DNA甲基转移酶1基因低表达的人支气管上皮细胞基因组甲基化水平的体外试验

The effect of DNA methyltransferase 1 low expression on the global genome DNA methylation status of 16HBE cell

目的观察人支气管上皮细胞（16HBE）的DNA甲基转移酶1（DNMT1）基因低表达细胞株细胞周期和基因组整体甲基化水平的改变。方法用慢病毒介导的RNA干扰方法将4个不同的短发夹RNA片段转染16HBE细胞，并用免疫印迹法（Western blotting）检测该细胞DNMT1蛋白表达水平，用流式细胞仪和5．甲基胞嘧啶（5-mC）免疫荧光法检测该细胞的细胞周期和细胞基因组整体甲基化水平。结果16HBE-shDNMT1—4细胞株的DNMT1蛋白表达与对照组相比平均降低约44％，差异有统计学意义（P〈0．05），但细胞周期和基因组整体甲基化水平无明显改变。结论成功建立DNMT1基因低表达的16HB细胞株，其细胞周期和基因组整体甲基化水平无明显改变。

Objective To construct DNA methyltransferase 1 （DNMT1） low expression 16HBE cell line and observe the variation of cell cycle and global genomic DNA methylation. Methods The method of Lenti-virus induced RNA interference was applied to introduce four different shRNA fragment into 16HBE cells. Flow cytometry and 5-mC immunofluorescence methods were used to observe the cell cycle and global DNA methylation status of DNMT1 low expression 16HBE cells. Results The DNMT1 protein relative expression level of 16HBE-shDNMT1-4 cell line was down regulated about 44%（P〈0.05 ） compared with the control. No obvious differences of cell cycle and global genome DNA methylation status were observed between the 16HBE and 16HBE-shDNMT1. Conclusion The DNMT1 gene low expression cell is successfully constructed, and there are no obvious changes happened on the cell cycle and global genomic DNA methvlation.