Protocols and quality controls are given for the following: 1) extraction, purification, and NeuN neuronal marker immunotagging of nuclei from adult human cerebral cortex; 2) fluorescence-activated nuclei sorting; 3) preparation of chromatin by micrococcal nuclease digest; 4) ChIP for open chromatin-associated histone methylation and acetylation; and 5) generation and sequencing of ChIP-seq libraries.
We present a ChIP-seq pipeline for epigenome mapping in the neuronal and nonneuronal nuclei from the postmortem brain. This includes a stepwise system of quality controls and user-friendly data presentation platforms.
Our practical guidelines will be useful for projects aimed at histone posttranslational modification mapping in chromatin extracted from hundreds of postmortem brain samples in cell-type-specific manner.
给出了以下方案和质量控制:1)提取,净化,和NeuN神经元标记对成年人大脑皮层的核进行免疫标记;2)荧光激活的核分选;3)通过微球菌核酸酶消化制备染色质;4)ChIP用于开放染色质相关组蛋白甲基化和乙酰化;5)ChIP-seq文库的生成和测序。
我们提出了一个ChIP-seq管道,用于死后大脑的神经元和非神经元核中的表观基因组作图。这包括逐步的质量控制系统和用户友好的数据呈现平台。
我们的实用指南对于以细胞类型特异性方式从数百个死后大脑样本中提取的染色质中的组蛋白翻译后修饰作图的项目将是有用的。