Transcriptional Regulation pp 157-173

Part of the Methods in Molecular Biology book series (MIMB, volume 809)

Mapping Protein–DNA Interactions Using ChIP-Sequencing

Protocol

Abstract

Chromatin immunoprecipitation (ChIP) allows enrichment of genomic regions which are associated with specific transcription factors, histone modifications, and indeed any other epitopes which are present on chromatin. The original ChIP methods used site-specific PCR and Southern blotting to confirm which regions of the genome were enriched, on a candidate basis. The combination of ChIP with genomic tiling arrays (ChIP-chip) allowed a more unbiased approach to map ChIP-enriched sites. However, limitations of microarray probe design and probe number have a detrimental impact on the coverage, resolution, sensitivity, and cost of whole-genome tiling microarray sets for higher eukaryotes with large genomes. The combination of ChIP with high-throughput sequencing technology has allowed more comprehensive surveys of genome occupancy, greater resolution, and lower cost for whole genome coverage. Herein, we provide a comparison of high-throughput sequencing platforms and a survey of ChIP-seq analysis tools, discuss experimental design, and describe a detailed ChIP-seq method.

Chromatin immunoprecipitation (ChIP) allows enrichment of genomic regions which are associated with specific transcription factors, histone modifications, and indeed any other epitopes which are present on chromatin. The original ChIP methods used site-specific PCR and Southern blotting to confirm which regions of the genome were enriched, on a candidate basis. The combination of ChIP with genomic tiling arrays (ChIP-chip) allowed a more unbiased approach to map ChIP-enriched sites. However, limitations of microarray probe design and probe number have a detrimental impact on the coverage, resolution, sensitivity, and cost of whole-genome tiling microarray sets for higher eukaryotes with large genomes. The combination of ChIP with high-throughput sequencing technology has allowed more comprehensive surveys of genome occupancy, greater resolution, and lower cost for whole genome coverage. Herein, we provide a comparison of high-throughput sequencing platforms and a survey of ChIP-seq analysis tools, discuss experimental design, and describe a detailed ChIP-seq method.

Key words

Chromatin immunoprecipitation High-throughput sequencing “Next-generation” sequencing Illumina (Solexa) sequencing Transcription Cancer Nuclear hormone receptor Androgen receptor 

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Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  1. 1.CRUK Cambridge Research InstituteCambridgeUK
  2. 2.Department of HaematologyCambridge Institute for Medical ResearchCambridgeUK
  3. 3.Centre for Molecular Medicine Norway, Nordic European Molecular Biology Laboratory PartnershipUniversity of OsloOsloNorway

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