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Chromatin immunoprecipitation (ChIP) is a powerful method for analyzing the interaction of regulatory proteins with genomic loci, but has been difficult to apply to studies on early embryos due to the limiting amount of genomic material in these samples. Here, we present a comprehensive technique for performing ChIP on blastula and gastrula stage Xenopus embryos. We also describe methods for optimizing crosslinking and chromatin shearing, verifying antibody specificity, maximizing PCR sensitivity, and quantifying PCR results, allowing for the use of as few as 50 early blastula stage embryos (approximately 5x10(4) cells) per experimental condition. Finally, we demonstrate the predicted binding of endogenous beta-catenin to the nodal-related 6 promoter, binding of tagged Fast-1/FoxH1 to the goosecoid promoter, and binding of tagged Tcf3 to the siamois and nodal-related 6 promoters as examples of the potential application of ChIP to embryological investigations. Developmental Dynamics 238:1422-1432, 2009. (c) 2009 Wiley-Liss, Inc.
Acevedo,
Genome-scale ChIP-chip analysis using 10,000 human cells.
2007, Pubmed
Acevedo,
Genome-scale ChIP-chip analysis using 10,000 human cells.
2007,
Pubmed Dahl,
A rapid micro chromatin immunoprecipitation assay (microChIP).
2008,
Pubmed Frank,
Binding of c-Myc to chromatin mediates mitogen-induced acetylation of histone H4 and gene activation.
2001,
Pubmed Heasman,
Beta-catenin signaling activity dissected in the early Xenopus embryo: a novel antisense approach.
2000,
Pubmed
,
Xenbase Jallow,
Specialized and redundant roles of TBP and a vertebrate-specific TBP paralog in embryonic gene regulation in Xenopus.
2004,
Pubmed
,
Xenbase Kim,
Non-canonical Wnt signals are modulated by the Kaiso transcriptional repressor and p120-catenin.
2004,
Pubmed
,
Xenbase Kuo,
In vivo cross-linking and immunoprecipitation for studying dynamic Protein:DNA associations in a chromatin environment.
1999,
Pubmed Luo,
Rb interacts with histone deacetylase to repress transcription.
1998,
Pubmed Messenger,
Functional specificity of the Xenopus T-domain protein Brachyury is conferred by its ability to interact with Smad1.
2005,
Pubmed
,
Xenbase Molenaar,
XTcf-3 transcription factor mediates beta-catenin-induced axis formation in Xenopus embryos.
1996,
Pubmed
,
Xenbase Morgan,
YY1 regulates the neural crest-associated slug gene in Xenopus laevis.
2004,
Pubmed
,
Xenbase Ng,
Epigenetic memory of an active gene state depends on histone H3.3 incorporation into chromatin in the absence of transcription.
2008,
Pubmed
,
Xenbase O'Neill,
Immunoprecipitation of native chromatin: NChIP.
2003,
Pubmed O'Neill,
Epigenetic characterization of the early embryo with a chromatin immunoprecipitation protocol applicable to small cell populations.
2006,
Pubmed Orlando,
Analysis of chromatin structure by in vivo formaldehyde cross-linking.
1997,
Pubmed Park,
Kaiso/p120-catenin and TCF/beta-catenin complexes coordinately regulate canonical Wnt gene targets.
2005,
Pubmed
,
Xenbase Rex,
Multiple interactions between maternally-activated signalling pathways control Xenopus nodal-related genes.
2002,
Pubmed
,
Xenbase Roh,
Active chromatin domains are defined by acetylation islands revealed by genome-wide mapping.
2005,
Pubmed Shen,
Nodal signaling: developmental roles and regulation.
2007,
Pubmed Takahashi,
Two novel nodal-related genes initiate early inductive events in Xenopus Nieuwkoop center.
2000,
Pubmed
,
Xenbase Taneyhill,
Investigating regulatory factors and their DNA binding affinities through real time quantitative PCR (RT-QPCR) and chromatin immunoprecipitation (ChIP) assays.
2008,
Pubmed Watabe,
Molecular mechanisms of Spemann's organizer formation: conserved growth factor synergy between Xenopus and mouse.
1995,
Pubmed
,
Xenbase WICKBOM,
Cytological studies on Dipnoi, Urodela, Anura, and Emys.
1945,
Pubmed Xanthos,
The roles of three signaling pathways in the formation and function of the Spemann Organizer.
2002,
Pubmed
,
Xenbase Yang,
Beta-catenin/Tcf-regulated transcription prior to the midblastula transition.
2002,
Pubmed
,
Xenbase Zeng,
In vivo dual cross-linking for identification of indirect DNA-associated proteins by chromatin immunoprecipitation.
2006,
Pubmed
