Genome-wide snapshot of chromatin regulators and states in Xenopus embryos by ChIP-Seq

Journal of Visualized Experiments : JoVE
George E GentschJames C Smith

Abstract

The recruitment of chromatin regulators and the assignment of chromatin states to specific genomic loci are pivotal to cell fate decisions and tissue and organ formation during development. Determining the locations and levels of such chromatin features in vivo will provide valuable information about the spatio-temporal regulation of genomic elements, and will support aspirations to mimic embryonic tissue development in vitro. The most commonly used method for genome-wide and high-resolution profiling is chromatin immunoprecipitation followed by next-generation sequencing (ChIP-Seq). This protocol outlines how yolk-rich embryos such as those of the frog Xenopus can be processed for ChIP-Seq experiments, and it offers simple command lines for post-sequencing analysis. Because of the high efficiency with which the protocol extracts nuclei from formaldehyde-fixed tissue, the method allows easy upscaling to obtain enough ChIP material for genome-wide profiling. Our protocol has been used successfully to map various DNA-binding proteins such as transcription factors, signaling mediators, components of the transcription machinery, chromatin modifiers and post-translational histone modifications, and for this to be done at various sta...Continue Reading

Citations

Jan 5, 2019·Cold Spring Harbor Protocols·George E Gentsch, James C Smith

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Methods Mentioned

BETA
immunoprecipitation
ChIP-Seq
ChIP
PCR
electrophoresis
RNA-Seq

Software Mentioned

homerTools
Java TreeView
Primer3
MEME
b2g4pipe
Java Web Start
SAMtools
cisFinder
UCSC Genome Browser
findMotifsGenome

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