发现小鼠Bcrp1基因替代启动子使用和转录调控的方法。
Methods to Discover Alternative Promoter Usage and Transcriptional Regulation of Murine Bcrp1.
作者信息
Natarajan Karthika, Xie Yi, Nakanishi Takeo, Moreci Rebecca S, Jeyasuria Pancharatnam, Hussain Arif, Ross Douglas D
机构信息
Greenebaum Cancer Center, University of Maryland School of Medicine; Pharmaceutical Sciences, University of Maryland School of Pharmacy.
Greenebaum Cancer Center, University of Maryland School of Medicine; Baltimore VA Medical Center.
出版信息
J Vis Exp. 2016 May 27(111):53827. doi: 10.3791/53827.
Gene expression in different tissues is often controlled by alternative promoters that result in the synthesis of mRNA with unique - usually untranslated - first exons. Bcrp1 (Abcg2), the murine orthologue of the ABC transporter Breast Cancer Resistance Protein (BCRP, ABCG2), has at least four alternative promoters that are designated by the corresponding four alternative first exons produced: E1U, E1A, E1B, and E1C. Herein, in-silico protocols are presented to predict alternative promoter usage for Bcrp1. Furthermore, reporter assay methods are described to produce reporter constructs for alternative promoters and to determine the functionality of putative promoters upstream of the alternative first exons that are identified.
不同组织中的基因表达通常受可变启动子调控，这些可变启动子会导致合成具有独特（通常是非翻译的）首个外显子的mRNA。AB C转运蛋白乳腺癌耐药蛋白（BCRP，ABCG2）的小鼠同源物Bcrp1（Abcg2）至少有四个可变启动子，它们由产生的相应四个可变首个外显子指定：E1U、E1A、E1B和E1C。本文介绍了用于预测Bcrp1可变启动子使用情况的计算机模拟方案。此外，还描述了报告基因检测方法，用于构建可变启动子的报告基因构建体，并确定所鉴定的可变首个外显子上游推定启动子的功能。
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