Chromatin immunoprecipitation of transcription factors and histone modifications in Comma-Dβ mammary epithelial cells

Holly Holliday; Amanda Khoury; Alexander Swarbrick

doi:10.1016/j.xpro.2021.100514

Chromatin immunoprecipitation of transcription factors and histone modifications in Comma-Dβ mammary epithelial cells

STAR Protoc. 2021 May 3;2(2):100514. doi: 10.1016/j.xpro.2021.100514. eCollection 2021 Jun 18.

Authors

Holly Holliday^{1

2

3}, Amanda Khoury^{1

2}, Alexander Swarbrick^{1

2}

Affiliations

¹ The Kinghorn Cancer Centre, Garvan Institute of Medical Research, Sydney, NSW 2010, Australia.
² St Vincent's Clinical School, Faculty of Medicine, UNSW Sydney, Sydney, NSW 2010, Australia.
³ Children's Cancer Institute, Lowy Cancer Research Centre, UNSW Sydney, Kensington, NSW 2052, Australia.

Abstract

Chromatin immunoprecipitation (ChIP) is used to study interactions between proteins and DNA. Nuclear lysates are prepared, and chromatin is fragmented by sonication. Antibodies are used to purify a protein of interest (e.g., a transcription factor or histone mark) along with any bound DNA. The genomic binding sites can then be mapped by sequencing the bound DNA (ChIP-seq) or by qPCR if binding sites are already known. ChIP requires optimization for each cell type, and success is highly antibody dependent. This protocol can be adapted to other cell lines with careful optimization. For complete details on the use and execution of this protocol, please refer to Holliday et al. (2021).

Keywords: ChIP-seq; Chromatin immunoprecipitation (ChIP); Molecular Biology.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Breast Neoplasms / metabolism*
Cell Line, Tumor
Chromatin Immunoprecipitation*
Female
Histone Code*
Humans
Mammary Glands, Human / metabolism*
Neoplasm Proteins / metabolism*
Transcription Factors / metabolism*

Substances

Neoplasm Proteins
Transcription Factors