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Methods Mol Biol. 2018;1675:111-130. doi: 10.1007/978-1-4939-7318-7_8.

Cell Type-Specific Profiling of Chromatin Modifications and Associated Proteins.

Author information

1
Centre National de la Recherche Scientifique (CNRS) UMR8197, Institut de Biologie de l'Ecole Normale Supérieure, Institut National de la Santé et de la Recherche Médicale (INSERM) U1024, Ecole Normale Supérieure, 46 rue d'Ulm, 75230, Paris Cedex 05, France.
2
Centre National de la Recherche Scientifique (CNRS) UMR8197, Institut de Biologie de l'Ecole Normale Supérieure, Institut National de la Santé et de la Recherche Médicale (INSERM) U1024, Ecole Normale Supérieure, 46 rue d'Ulm, 75230, Paris Cedex 05, France. francois.roudier@ens-lyon.fr.
3
Laboratoire Reproduction et Développement des Plantes, Univ Lyon, ENS de Lyon, UCB Lyon 1, CNRS, INRA, F-69342, Lyon, France. francois.roudier@ens-lyon.fr.

Abstract

Progression of a cell along a differentiation path is characterized by changes in gene expression profiles. Alterations of these transcriptional programs result from cell type-specific transcription factors that act in a dynamic chromatin environment. Understanding the precise contribution of these molecular factors during the differentiation process requires accessing specific cell types within a developing organ. This chapter describes a streamlined and alternative version of INTACT, a method enabling the isolation of specific cell populations by affinity-purification of tagged nuclei and the subsequent analysis of gene expression, transcription factor binding profiles, as well as chromatin state at a genome-wide scale. In particular, modifications of the nuclei isolation, capture, and purification procedures are proposed that improve time scale, yield, and purity. In addition, the combination of different tags enables the analysis of distinct cell populations from a single transgenic line and the subtractive purification of subpopulations of cells, including those for which no specific promoter is available. Finally, we describe a chromatin immunoprecipitation protocol that has been successfully used to profile histone modifications and other chromatin-associated proteins such as RNA Polymerase II in different cell populations of the Arabidopsis root, including the quiescent center of the stem cell niche.

KEYWORDS:

Cell type; Chromatin; Epigenome; Histone modification; INTACT; Nucleus; RNA polymerase II; Transcriptome

PMID:
29052189
DOI:
10.1007/978-1-4939-7318-7_8
[Indexed for MEDLINE]

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