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用于染色质免疫沉淀(ChIP)分析的冷冻组织染色质制备

Chromatin Preparation from Frozen Tissues for Chromatin Immunoprecipitation (ChIP) Assays.

作者信息

Martínez Virginia López, Fraga Mario Fernández

机构信息

Cancer Epigenetics and Nanomedicine Laboratory, Centro de Investigación en Nanomateriales y Nanotecnología-Consejo Superior de Investigaciones Científicas (CINN-CSIC), Instituto de Investigación Sanitaria del Principado de Asturias (ISPA-FINBA), Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER), Instituto de Salud Carlos III (ISCIII), Instituto Universitario de Oncología del Principado de Asturias (IUOPA), Universidad de Oviedo, Oviedo, Spain.

出版信息

Methods Mol Biol. 2025;2930:203-217. doi: 10.1007/978-1-0716-4558-1_14.

Abstract

Chromatin immunoprecipitation coupled with next-generation sequencing (ChIP-seq) is a crucial method for examining transcription factor binding and histone modifications across the entire genome. This is a key step in deciphering the complex mechanisms that control cancer immunosurveillance. Here, we provide a comprehensive protocol covering all the steps needed to obtain DNA for ChIP-seq library preparation, starting from tissue dissection, followed by fixation, chromatin preparation, immunoprecipitation, and finally DNA purification. The protocol is optimized for frozen mice tissues, but can be easily adapted for use with any model organism. The resulting immunoprecipitated chromatin is suitable for library preparation and sequencing on an Illumina platform.

摘要

染色质免疫沉淀结合下一代测序(ChIP-seq)是一种用于检测全基因组中转录因子结合和组蛋白修饰的关键方法。这是破译控制癌症免疫监视的复杂机制的关键一步。在这里,我们提供了一个全面的方案,涵盖了从组织解剖开始,经过固定、染色质制备、免疫沉淀,最后到DNA纯化,获取用于ChIP-seq文库制备的DNA所需的所有步骤。该方案针对冷冻小鼠组织进行了优化,但可以很容易地适用于任何模式生物。所得的免疫沉淀染色质适用于在Illumina平台上进行文库制备和测序。

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