Chromatin Interaction Analysis with Paired-End Tag Sequencing (ChIA-PET) for mapping chromatin interactions and understanding transcription regulation.

Journal of Visualized Experiments : JoVE
Yufen GohYijun Ruan

Abstract

Genomes are organized into three-dimensional structures, adopting higher-order conformations inside the micron-sized nuclear spaces (7, 2, 12). Such architectures are not random and involve interactions between gene promoters and regulatory elements (13). The binding of transcription factors to specific regulatory sequences brings about a network of transcription regulation and coordination (1, 14). Chromatin Interaction Analysis by Paired-End Tag Sequencing (ChIA-PET) was developed to identify these higher-order chromatin structures (5,6). Cells are fixed and interacting loci are captured by covalent DNA-protein cross-links. To minimize non-specific noise and reduce complexity, as well as to increase the specificity of the chromatin interaction analysis, chromatin immunoprecipitation (ChIP) is used against specific protein factors to enrich chromatin fragments of interest before proximity ligation. Ligation involving half-linkers subsequently forms covalent links between pairs of DNA fragments tethered together within individual chromatin complexes. The flanking MmeI restriction enzyme sites in the half-linkers allow extraction of paired end tag-linker-tag constructs (PETs) upon MmeI digestion. As the half-linkers are biot...Continue Reading

References

Dec 1, 1995·Trends in Biochemical Sciences·K RippeJ Langowski
Sep 14, 2004·Nature Genetics·Cameron S OsbornePeter Fraser
Feb 12, 2008·Current Protocols in Molecular Biology·Patrick NgYijun Ruan
Nov 6, 2009·Nature·Melissa J FullwoodYijun Ruan
Jan 14, 2010·Current Protocols in Molecular Biology·Melissa J FullwoodYijun Ruan
Jul 6, 2010·Current Opinion in Genetics & Development·Wulan Deng, Gerd A Blobel

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Citations

May 18, 2016·Nucleic Acids Research·Przemyslaw SzalajDariusz Plewczynski
Apr 14, 2015·Molecular Endocrinology·Pia D BagamasbadRobert J Denver
Nov 2, 2019·Genes·Enrique Hernández-LemusSoledad Ochoa

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Methods Mentioned

BETA
Immunoprecipitation
ChIP
scraping
by
quantitative
eluted
and
PCR
proximity ligation
DNA Assay

Software Mentioned

ChIA
Illumina Sequencing Control Studio
PET
PET Tool

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