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目的 利用可被放射线激活而转录的早期生长反应基因 1 (Egr 1 )启动子驱动单纯疱疹病毒胸苷激酶基因 (tk)在肝癌细胞中高效表达。方法 构建以Egr 1为启动子 ,以tk为目的基因的重组质粒 pET ,经脂质体介导转染人肝癌细胞株SMMC 772 1 ,经G41 8抗性筛选、分别以 0、5、7.5、1 0、1 5、2 0Gy剂量γ射线照射、逆转录 聚合酶链反应 (RT PCR)半定量分析、比较tk基因的mRNA表达。结果 转染后的肝癌细胞株经射线照射后其tkmRNA的表达较未照射组 (55 .2±7.2 )有明显提高 ,以 1 5Gy最为显著 (1 1 7.2± 1 1 .1 ,P <0 .0 0 1 )。结论 经γ射线照射后 ,可被放射线转录激活的Egr 1启动子可引起tk基因在肝癌细胞中高效表达 ,从而为肝癌的基因 放射治疗的实验研究打下坚实的基础
OBJECTIVE: To efficiently express herpes simplex virus thymidine kinase (tk) gene in hepatoma cells by using the promoter of the early growth response gene 1 (Egr 1) that can be activated by radiation. Methods Recombinant plasmid pET with Egr 1 as promoter and tk as target gene was constructed and transfected into human hepatocellular carcinoma cell line SMMC 772 1 by lipofectamine 2000. After screening by G41 8, The mRNA expression of tk gene was analyzed by semi-quantitative RT-PCR with 1, 10, 15 and 20 Gy doses of γ-rays and reverse transcriptase polymerase chain reaction (RT PCR). Results After transfection, the expression of t-kappa-tRNA in hepatoma cell lines was significantly higher than that in the non-irradiated group (55.2 ± 7.2), most significantly at 15 Gy (11.2 ± 11.1, P <0. 0 0 1). Conclusion The Egr 1 promoter, which can be activated by radiation, can cause the high expression of tk gene in hepatoma cells after γ-ray irradiation, so as to lay a solid foundation for the experimental study of gene therapy for hepatocellular carcinoma