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Chromatin tandem affinity purification sequencing.

ABSTRACT: Chromatin immunoprecipitation coupled with ultra-high-throughput sequencing (ChIP-seq) is a widely used method for mapping the interactions of proteins with DNA. However, the requirements for ChIP-grade antibodies impede wider application of this method, and variations in results can be high owing to differences in affinity and cross-reactivity of antibodies. Therefore, we developed chromatin tandem affinity purification (ChTAP) as an effective alternative to ChIP. Through the use of affinity tags and reagents that are identical for all proteins investigated, ChTAP enables one to directly compare the binding between different transcription factors and to directly assess the background in control experiments. Thus, ChTAP-seq can be used to rapidly map the genome-wide binding of multiple DNA-binding proteins in a wide range of cell types. ChTAP can be completed in 3-4 d, starting from cross-linking of chromatin to purification of ChIP DNA.

SUBMITTER: Soleimani VD

PROVIDER: S-EPMC4058862 | biostudies-literature | 2013 Aug

REPOSITORIES: biostudies-literature

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Chromatin tandem affinity purification sequencing.

Soleimani Vahab D VD Palidwor Gareth A GA Ramachandran Parameswaran P Perkins Theodore J TJ Rudnicki Michael A MA

Nature protocols 20130711 8

Chromatin immunoprecipitation coupled with ultra-high-throughput sequencing (ChIP-seq) is a widely used method for mapping the interactions of proteins with DNA. However, the requirements for ChIP-grade antibodies impede wider application of this method, and variations in results can be high owing to differences in affinity and cross-reactivity of antibodies. Therefore, we developed chromatin tandem affinity purification (ChTAP) as an effective alternative to ChIP. Through the use of affinity ta ...[more]

PMID: 23845964

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