Fish'n ChIPs: chromatin immunoprecipitation in the zebrafish embryo.
Chromatin immunoprecipitation (ChIP) is arguably the assay of choice to determine the genomic localization of DNA- or chromatin-binding proteins, including post-translationally modified histones, in cells. The increasing importance of the zebrafish, Danio rerio, as a model organism in functional genomics has recently sparked investigations of ChIP-based genome-scale mapping of modified histones on promoters, and studies on the role of specific transcription factors in developmental processes. ChIP assays used in these studies are cumbersome and conventionally require relatively large number of embryos. To simplify the procedure and to be able to apply the ChIP assay to reduced number of embryos, we re-evaluated the protocol for preparation of embryonic chromatin destined to ChIP. We found that manual homogenization of embryos rather than protease treatment to remove the chorion enhances ChIP efficiency and quickens the assay. We also incorporated key steps from a recently published ChIP assay for small cell numbers. We report here a protocol for immunoprecipitation of modified histones from mid-term blastula zebrafish embryos.
Lindeman
Leif C
LC
Department of Biochemistry, Institute of Basic Medical Sciences, University of Oslo, Oslo, Norway.
Vogt-Kielland
Linn T
LT
Aleström
Peter
P
Collas
Philippe
P
D000818:Animals; D047369:Chromatin Immunoprecipitation / Q000379:methods*; D003432:Cross-Linking Reagents / Q000494:pharmacology; D004247:DNA / Q000032:analysis / Q000302:isolation & purification / Q000378:metabolism; D004625:Embryo, Nonmammalian / Q000166:cytology* / Q000378:metabolism; D016133:Polymerase Chain Reaction / Q000379:methods; D015027:Zebrafish / Q000196:embryology* / Q000235:genetics* / Q000378:metabolism / Q000502:physiology
D003432:Cross-Linking Reagents; D004247:DNA