pax2启动子的克隆及其在人胚胎肾293T细胞中的转录活性

The Cloning of pax2 Promoter and Characterization of the Promoter Activity in 293T Cell

目的:克隆paired box2(pax2)基因的启动子,插入荧光素酶报告基因载体中,并检测其活性。方法:采用PCR技术从人乳腺癌细胞系MCF-7基因组中扩增出pax2启动子,插入荧光素酶报告基因载体pGL3-basic中,确定所扩增的DNA序列。将重组的报告基因瞬时转染人胚胎肾293T细胞,检测pax2启动子活性。结果:测序结果显示扩增的pax2启动子序列正确;活性实验表明构建的报告基因具有启动子活性,雌激素受体α(ERα)能以剂量依赖的方式升高pax2报告基因的转录。结论:克隆了pax2启动子,为ERα共调节子的功能研究提供了重要基础。

Objective： To clone pax2 promoter and insert it into a luciferase reporter gene vector, and to characterize the promoter activity. Methods： The promoter of pax2 was amplified from human breast cancer cell line MCF-7 genomic DNA by PCR and cloned into pGL3-basic. The resulting plasmid was determined by DNA sequencing. The promoter activity was analyzed in 293T cell line. Results： The result of DNA sequencing showed that the sequence of the cloned promoter was correct. The activity of the pax2 luciferase reporter gene was confirmed, and this activity was increased by estrogen receptor α（ERct） in a dose-dependent manner. Conclusion： pax2 promoter was cloned successfully, and this will be an important basis for the study of the functions of ERct coregulators.