Teves Sheila S, Henikoff Steven
Division of Basic Sciences, Fred Hutchinson Cancer Research Center and Molecular and Cellular Biology Program, University of Washington, Seattle, WA, USA.
Methods Mol Biol. 2012;833:421-32. doi: 10.1007/978-1-61779-477-3_25.
Salt fractionation of nucleosomes, a classical method for defining "active" chromatin based on nucleosome solubility, has recently been adapted for genome-scale profiling. This method has several advantages for profiling chromatin dynamics, including general applicability to cell lines and tissues, quantitative recovery of chromatin, base-pair resolution of nucleosomes, and overall simplicity both in concept and execution. This chapter provides detailed protocols for nuclear isolation, chromatin fragmentation by micrococcal nuclease digestion, successive solubilization of chromatin fractions by addition of increasing concentrations of salt, and genome-wide analyses through microarray hybridization and next-generation sequencing.
核小体的盐分级分离是一种基于核小体溶解性来定义“活性”染色质的经典方法，最近已被应用于基因组规模的分析。该方法在分析染色质动力学方面具有多个优点，包括对细胞系和组织的普遍适用性、染色质的定量回收、核小体的碱基对分辨率，以及在概念和操作上的整体简便性。本章提供了详细的实验方案，包括细胞核分离、通过微球菌核酸酶消化进行染色质片段化、通过添加浓度递增的盐连续溶解染色质组分，以及通过微阵列杂交和新一代测序进行全基因组分析。
