论文部分内容阅读
目的:应用RNA干扰(RNAi)技术抑制人膀胱癌细胞(BIU-87)EGFR基因的表达,探索采用该技术干预膀胱癌的可行性。方法:制备3个针对人EGFR基因的shRNA表达质粒,转染BIU-87细胞48 h后,RT-PCR及蛋白质印迹法检测EGFR基因的表达。结果:酶切及测序鉴定证实,3个shRNA表达质粒构建成功,将其转染BIU-87细胞后,第3号表达质粒抑制效果最为明显,使EGFR基因表达在mRNA水平抑制〔抑制率(51.43±4.09)%〕,在蛋白水平抑制〔抑制率(46.37±5.25)%〕。结论:针对EGFR基因设计的shRNA在体外有效地抑制了膀胱癌EG-FR基因表达,且实现RNA干扰具有序列选择性。
OBJECTIVE: To use RNA interference (RNAi) technology to inhibit the expression of EGFR gene in human bladder cancer cells (BIU-87) and to explore the feasibility of using this technique to interfere with bladder cancer. Methods: Three shRNA expression plasmids targeting human EGFR gene were prepared and transfected into BIU-87 cells for 48 h. The expression of EGFR gene was detected by RT-PCR and Western blotting. Results: The three shRNA expression plasmids were successfully constructed and confirmed by restriction enzyme digestion and sequencing. The expression of plasmid 3 was the most obvious after transfected into BIU-87 cells, and the expression of EGFR gene was inhibited at the mRNA level [inhibition rate (51.43 ± 4.09)%] at the protein level [inhibition rate (46.37 ± 5.25)%]. Conclusion: shRNA targeting EGFR gene effectively inhibits the expression of EG-FR gene in bladder cancer and has sequence selectivity for RNA interference.