Biology Department, Indiana University, Bloomington, IN, USA.
Methods Mol Biol. 2021;2346:165-172. doi: 10.1007/7651_2020_307.
Transcription factors are ubiquitous proteins that associate with promoter DNA and regulate gene expression through a variety of mechanisms. Understanding transcriptional control mechanisms requires in-depth investigation of the binding of transcription factors to the promoters they regulate. There are many in vivo and in vitro methods for testing the binding of a known protein to a promoter, such as chromatin immunoprecipitation and electrophoretic mobility shift assays. However, for these experiments, one must have a protein candidate to test and is not able to identify unknown proteins bound to a particular promoter. Thus, the promoter pull-down assay was developed to fill this void. This method uses DNA as bait to capture proteins that bind to a specific promoter, such as transcription factors, from cellular lysates. Coupled with other experiments, the promoter pull-down assay vastly improves the repertoire of methods available for defining regulatory complexes that influence transcription.
转录因子是普遍存在的蛋白质，它们与启动子 DNA 结合，并通过多种机制调节基因表达。要了解转录控制机制，需要深入研究转录因子与它们调节的启动子的结合。有许多体内和体外的方法可以测试已知蛋白质与启动子的结合，如染色质免疫沉淀和电泳迁移率变动分析。然而，对于这些实验，人们必须有一个要测试的蛋白质候选物，而不能鉴定与特定启动子结合的未知蛋白质。因此，启动子下拉测定法就是为了填补这一空白而开发的。这种方法使用 DNA 作为诱饵，从细胞裂解物中捕获与特定启动子（如转录因子）结合的蛋白质。与其他实验相结合，启动子下拉测定法极大地扩展了可用于定义影响转录的调节复合物的方法组合。
