Braveman Melissa W, Chen-Plotkin Alice S, Yohrling George J, Cha Jang-Ho J
Mass General Institute for Neurodegenerative Disease and Department of Neurology, Massachusetts General Hospital, Charlestown, USA.
Methods Mol Biol. 2004;277:261-76. doi: 10.1385/1-59259-804-8:261.
Transcriptional dysregulation has emerged as an important pathologic mechanism underlying the pathogenesis of Huntington's disease (HD). The control of transcription depends on appropriate binding of transcription factor proteins to specific promoter regions of genes. Chromatin immunoprecipitation (ChIP) is a technique that has been used to study the association of transcription factors with DNA. To address the hypothesis that there is altered transcription factor-DNA association in HD, we have recently adapted the ChIP technique to the study of transgenic mouse brain. Here, we describe our method of performing ChIP in intact mouse brain. We have optimized conditions for formaldehyde crosslinking, antibody immunoprecipitation, and quantitative real-time polymerase chain reaction detection. Using ChIP, one can measure the association of transcription factors with specific genes and determine if this association is altered in transgenic HD mouse models. ChIP applied to whole-mouse brain can thus offer a window into mechanisms of transcriptional dysregulation.
转录失调已成为亨廷顿舞蹈病（HD）发病机制的重要病理机制。转录的调控取决于转录因子蛋白与基因特定启动子区域的适当结合。染色质免疫沉淀（ChIP）是一种用于研究转录因子与DNA关联的技术。为了验证HD中转录因子与DNA的关联发生改变这一假说，我们最近将ChIP技术应用于转基因小鼠脑的研究。在此，我们描述在完整小鼠脑中进行ChIP的方法。我们优化了甲醛交联、抗体免疫沉淀和定量实时聚合酶链反应检测的条件。使用ChIP，人们可以测量转录因子与特定基因的关联，并确定这种关联在转基因HD小鼠模型中是否发生改变。因此，应用于全小鼠脑的ChIP可为转录失调机制提供一个窗口。
