Summary of ChIP-Seq Methods and Description of an Optimized ChIP-Seq Protocol.
Methods Mol Biol
; 2842: 419-447, 2024.
Article
in En
| MEDLINE
| ID: mdl-39012609
ABSTRACT
Chromatin immunoprecipitation (ChIP) is an invaluable method to characterize interactions between proteins and genomic DNA, such as the genomic localization of transcription factors and post-translational modification of histones. DNA and proteins are reversibly and covalently crosslinked using formaldehyde. Then the cells are lysed to release the chromatin. The chromatin is fragmented into smaller sizes either by micrococcal nuclease (MN) or sonication and then purified from other cellular components. The protein-DNA complexes are enriched by immunoprecipitation (IP) with antibodies that target the epitope of interest. The DNA is released from the proteins by heat and protease treatment, followed by degradation of contaminating RNAs with RNase. The resulting DNA is analyzed using various methods, including polymerase chain reaction (PCR), quantitative PCR (qPCR), or sequencing. This protocol outlines each of these steps for both yeast and human cells. This chapter includes a contextual discussion of the combination of ChIP with DNA analysis methods such as ChIP-on-Chip, ChIP-qPCR, and ChIP-Seq, recent updates on ChIP-Seq data analysis pipelines, complementary methods for identification of binding sites of DNA binding proteins, and additional protocol information about ChIP-qPCR and ChIP-Seq.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Chromatin Immunoprecipitation Sequencing
Limits:
Humans
Language:
En
Journal:
Methods Mol Biol
/
Methods in molecular biology
/
Methods mol. biol
Journal subject:
BIOLOGIA MOLECULAR
Year:
2024
Document type:
Article
Affiliation country:
United States
Country of publication:
United States