Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA.
Nat Protoc. 2012 Jan 19;7(2):256-67. doi: 10.1038/nprot.2011.444.
Eviction or destabilization of nucleosomes from chromatin is a hallmark of functional regulatory elements in eukaryotic genomes. Historically identified by nuclease hypersensitivity, these regulatory elements are typically bound by transcription factors or other regulatory proteins. FAIRE (formaldehyde-assisted isolation of regulatory elements) is an alternative approach to identify these genomic regions and has proven successful in a multitude of eukaryotic cell and tissue types. Cells or dissociated tissues are cross-linked briefly with formaldehyde, lysed and sonicated. Sheared chromatin is subjected to phenol/chloroform extraction and the isolated DNA, typically encompassing 1-3% of the human genome, is purified. We provide guidelines for quantitative analysis by PCR, microarrays or next-generation sequencing. Regulatory elements enriched by FAIRE have high concordance with those identified by nuclease hypersensitivity or chromatin immunoprecipitation (ChIP), and the entire procedure can be completed in 3 d. FAIRE has low technical variability, which allows its usage in large-scale studies of chromatin from normal or diseased tissues.
核小体从染色质上的逐出或去稳定化是真核基因组中功能调节元件的一个标志。这些调节元件最初是通过核酸酶敏感性来鉴定的，通常由转录因子或其他调节蛋白所结合。FAIRE（甲醛辅助分离调节元件）是一种识别这些基因组区域的替代方法，已在多种真核细胞和组织类型中得到成功验证。用甲醛短暂交联细胞或分离的组织，然后裂解和超声处理。用酚/氯仿抽提得到的断裂染色质，然后纯化分离的 DNA，通常包含人类基因组的 1-3%。我们提供了通过 PCR、微阵列或下一代测序进行定量分析的指南。FAIRE 富集的调节元件与通过核酸酶敏感性或染色质免疫沉淀（ChIP）鉴定的那些具有高度一致性，整个过程可以在 3 天内完成。FAIRE 具有较低的技术变异性，使其可以用于正常或患病组织中染色质的大规模研究。
