Furlan-Magaril Mayra, Recillas-Targa Félix
Nuclear Dynamics Programme, The Babraham Institute, Cambridge, CB22 3AT, UK.
Instituto de Fisiología Celular, Departamento de Genética Molecular, Universidad Nacional Autónoma de México, Mexico City, USA.
Methods Mol Biol. 2015;1334:205-18. doi: 10.1007/978-1-4939-2877-4_13.
DNA regulatory elements nucleate the interaction of several transcription factors in conjunction with ubiquitous and/or tissue-specific cofactors in order to regulate gene expression making it relevant to determine the profiles of cohabitation of several proteins on the chromatin fiber. Chromatin immunoprecipitation (ChIP) has been broadly used to determine the profile of several histone posttranslational modifications as well as transcription factor occupancy in vivo. However, individual ChIP does not resolve whether the epitope under study is present at the same time on a given genomic location. Here we describe the ChIP-re-ChIP assay that represents a direct strategy to determine the in vivo co-localization of proteins or histone posttranslational modifications in a chromatinized template on the basis of double and independent rounds of immunoprecipitation with high-quality ChIP-grade antibodies.
DNA调控元件促使几种转录因子与普遍存在的和/或组织特异性辅因子相互作用，从而调节基因表达，因此确定几种蛋白质在染色质纤维上的共定位情况具有重要意义。染色质免疫沉淀（ChIP）已被广泛用于确定几种组蛋白翻译后修饰的情况以及体内转录因子的占有率。然而，单个ChIP无法确定所研究的表位是否同时存在于给定的基因组位置。在此，我们描述了ChIP再ChIP分析方法，该方法是一种直接策略，用于在基于两轮独立且高质量的ChIP级抗体免疫沉淀的染色质化模板中，确定蛋白质或组蛋白翻译后修饰的体内共定位情况。
