Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, TN, 37205, USA.
Interdisciplinary Graduate Program, Vanderbilt University School of Medicine, Nashville, TN, 37205, USA.
STAR Protoc. 2021 May 19;2(2):100530. doi: 10.1016/j.xpro.2021.100530. eCollection 2021 Jun 18.
Transcriptional changes happen within minutes; however, RNAi or genetic deletion requires days to weeks before transcription networks can be analyzed. This limitation has made it challenging to distinguish direct from indirect targets of sequence-specific transcription factors. This inability to define direct transcriptional targets hinders detailed studies of transcriptional mechanisms. This protocol combines rapid degradation of endogenous transcription factors with nascent transcript analysis to define the earliest, and likely direct, regulatory targets of transcription factors. For complete details on the use and execution of this protocol, please refer to Stengel et al., 2021).
转录变化发生在数分钟内；然而，RNAi 或基因缺失需要数天到数周的时间才能分析转录网络。这一限制使得区分序列特异性转录因子的直接和间接靶标变得具有挑战性。这种无法定义直接转录靶标的能力阻碍了对转录机制的详细研究。该方案将内源转录因子的快速降解与新生转录本分析相结合，以定义转录因子的最早且可能直接的调控靶标。有关该方案使用和执行的完整详细信息，请参见 Stengel 等人，2021）。
