催乳素对奶牛RANKL基因启动子活性的研究

Effect of prolactin on RANKL gene promoter activity in dairy cows

文章以奶牛乳腺上皮细胞为实验模型,探讨催乳素对RANKL基因启动子活性的调节。采用Western Blot方法检测催乳素对RANKL蛋白表达的影响,构建RANKL启动子荧光素酶载体,采用双荧光素酶报告基因检测系统检测催乳素对RANKL基因启动子活性影响,采用定点合成突变的方法,构建突变型RANKL启动子荧光素酶载体,运用双荧光素酶报告基因检测系统检测催乳素对突变型RANKL启动子活性影响。结果表明,催乳素能够显著提高乳腺上皮细胞RANKL启动子活性及RANKL蛋白表达,催乳素响应元件在奶牛RANKL基因启动子-826~-814 bp区域。

In this study,using mammary epithelial cells of dairy cows as a experimental model to investigate the regulation of prolactin on RANKL gene promoter activity.The expression of RANKL protein induced by prolactin was detected by Western Blot.The RANKL promoter luciferase vectors were constructed,and the the effect of prolactin on RANKL promoter activity was detected by dual luciferase assay.The mutant RANKL promoter luciferase vector was constructed by site-directed mutation synthesis,and the effect of prolactin on the activity of mutant RANKL promoter was detected by dual luciferase assay.The results showed that prolactin significantly increased the promoter activity and protein expression of RANKL in mammary epithelial cells.The prolactin response element was located in the-826~-814 bp region of dairy cows RANKL gene promoter.