Abstract
Isolation of specific genomic regions retaining molecular interactions is necessary for their biochemical analysis. Here, we describe engineered DNA-binding molecule-mediated chromatin immunoprecipitation (enChIP) using the CRISPR system, for purification of specific genomic regions retaining molecular interactions. In this form of enChIP, specific genomic regions are immunoprecipitated with antibody against a tag(s), which is fused to a catalytically inactive form of Cas9 (dCas9), which is co-expressed with a guide RNA (gRNA) and recognizes endogenous DNA sequence in the genomic regions of interest. enChIP combined with mass spectrometry (enChIP-MS), next-generation sequencing (enChIP-Seq), and RNA-Seq (enChIP-RNA-Seq) can identify proteins, other genomic regions, and RNA, respectively, that interact with the target genomic region.
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Acknowledgement
This work was supported by Research Institute for Microbial Diseases (T.F. and H.F.), Program for Combined Research Fields from Immunology Frontier Research Center, Osaka University (H.F.), the Takeda Science Foundation (H.F. and T.F.), the Asahi Glass Foundation (H.F.), the Kurata Memorial Hitachi Science and Technology Foundation (T.F. and H.F.), Grant-in-Aid for Young Scientists (B) (#25830131) (T.F.), and Grant-in-Aid for Scientific Research on Innovative Areas "Cell Fate" (#23118516) (T.F.), "The Genofield" (#23114707), "Transcription Cycle" (#25118512) (H.F.) from the Ministry of Education, Culture, Sports, Science and Technology of Japan.
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Fujita, T., Fujii, H. (2015). Isolation of Specific Genomic Regions and Identification of Associated Molecules by Engineered DNA-Binding Molecule-Mediated Chromatin Immunoprecipitation (enChIP) Using CRISPR. In: Chellappan, S. (eds) Chromatin Protocols. Methods in Molecular Biology, vol 1288. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2474-5_4
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DOI: https://doi.org/10.1007/978-1-4939-2474-5_4
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-2473-8
Online ISBN: 978-1-4939-2474-5
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