Histone acid extraction assay is a popular method to determine histone modification levels in mammalian cells. It includes three steps: first, histones are released from chromatin by sulfuric acid; trichloroacetate (TCA) is then added to precipitate histones; and finally, histones are dissolved in double-distilled H2O (ddH2O). Here we present a detailed histone acid extraction assay in our laboratory using a colon cancer cell line, HCT116, as a model.
[Background] The nucleosome is the fundamental unit of eukaryotic chromatin, which is composed of a histone octamer (2 copies of H3, H4, H2A, H2B, respectively) wrapping by DNA (Strahl and Allis, 2000). The amino terminal of histone is subjected to a variety of post-translational modifications, such as methylation, acetylation, phosphorylation, ubiquitylation and sumoylation (Kouzarides, 2007). Although the function of these modifications has remained elusive, there is ever-growing studies suggest that histone modifications play vital roles in intracellular processes (Bannister and Kouzarides, 2011). Therefore, it is important to extract histones efficiently to detect histone modifications.
Histones can be extracted via different methods, in which histone acid extraction assay is one of the most popular procedures. It does not interrupt post-translational modifications of histones, and so it is very good for histone modification analysis. It has been tested that the extracted histones can be used in Western blot, and maybe other assays (not fully tested). However, immunoprecipitation is not recommended. In this protocol, we will present a detailed histone acid extraction assay, and describe how to release histones from chromatin, how to precipitate histones, and how to wash and dissolve histones in ddH2O.
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