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Nat Protoc. 2008;3(4):698-709. doi: 10.1038/nprot.2008.38.

Chromatin immunoprecipitation (ChIP) coupled to detection by quantitative real-time PCR to study transcription factor binding to DNA in Caenorhabditis elegans.

Author information

  • 1Program in Gene Function and Expression, Program in Molecular Medicine, Aaron Lazare Research Building, University of Massachusetts Medical School, 364 Plantation Street, Worcester, Massachusetts 01605, USA.

Abstract

In order to determine how signaling pathways differentially regulate gene expression, it is necessary to identify the interactions between transcription factors (TFs) and their cognate cis-regulatory DNA elements. Here, we have outlined a chromatin immunoprecipitation (ChIP) protocol for use in whole Caenorhabditis elegans extracts. We discuss optimization of the procedure, including growth and harvesting of the worms, formaldehyde fixation, TF immunoprecipitation and analysis of bound sequences through real-time PCR. It takes approximately 10-12 d to obtain the worm culture for ChIP; the ChIP procedure is spaced out over a period of 2.5 d with two overnight incubations.

PMID:
18388953
[PubMed - indexed for MEDLINE]
PMCID:
PMC2681100
Free PMC Article

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