摘要
目的:构建DEK的pcDNA3-Flag表达载体,研究其对抑癌基因p53启动子活性的影响。方法:以乳腺文库为模板,PCR扩增DEK编码序列,克隆到pcDNA3-Flag载体,构建成pcDNA3-Flag-DEK,转染293T细胞,Western印迹鉴定pcDNA3-Flag载体介导的DEK的表达,萤光素酶报告基因活性实验研究DEK对p53启动子活性的影响。结果:双酶切实验证实得到pcDNA3-Flag-DEK阳性克隆;Western印迹实验发现DEK在293T细胞内表达;转录活性实验表明在ZR75-1乳腺癌细胞中,DEK呈剂量依赖性抑制p53启动子的活性。结论:构建了DEK的真核表达载体,并发现此表达载体能在ZR75-1乳腺癌细胞中抑制p53启动子活性。
Objective: To construct a eukaryotic expression vector for expression of DEK, and to study its effect on p53 promoter activity. Methods: The coding sequences of full length DEK was amplified from breast library by PCR and cloned into the pcDNA3-Flag vector. DEK expression was detected by Western blot after the recombi- nant plasmids were transfected into 293T cells. The activity of p53 promoter was detected in ZR75-1 cells. Re- sults: The full length DEK was expressed in the 293T cells. Compared to the empty vector, the activity of p53 promoter was decreased by DEK in a dose-dependent manner. Conclusion: We have constructed the eukaryotic ex- pression vector of DEK and found that p53 promoter activity was decreased by DEK in ZR75-1 cells.
出处
《生物技术通讯》
CAS
2013年第1期34-36,共3页
Letters in Biotechnology
基金
国家自然科学基金(31071174)