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Genes Cells. 2018 Apr;23(4):318-325. doi: 10.1111/gtc.12573. Epub 2018 Feb 26.

Transgenic mouse lines expressing the 3xFLAG-dCas9 protein for enChIP analysis.

Author information

1
Department of Biochemistry and Genome Biology, Hirosaki University Graduate School of Medicine, Hirosaki, Japan.
2
Chromatin Biochemistry Research Group, Combined Program on Microbiology and Immunology, Research Institute for Microbial Diseases, Osaka University, Suita, Japan.
3
Department of Experimental Genome Research, Genome Information Research Center, Research Institute for Microbial Diseases, Osaka University, Suita, Japan.
4
Laboratory for Transcriptional Regulation, RIKEN Center for Integrative Medical Sciences, Yokohama City, Japan.

Abstract

We developed the engineered DNA-binding molecule-mediated chromatin immunoprecipitation (enChIP) technology to isolate specific genomic regions while retaining their molecular interactions. In enChIP, the locus of interest is tagged with an engineered DNA-binding molecule, such as a modified form of the clustered regularly interspaced short palindromic repeats (CRISPR) system containing a guide RNA (gRNA) and a catalytically inactive form of Cas9 (dCas9). The locus is then affinity-purified to enable identification of associated molecules. In this study, we generated transgenic mice expressing 3xFLAG-tagged Streptococcus pyogenes dCas9 (3xFLAG-dCas9) and retrovirally transduced gRNA into primary CD4+ T cells from these mice for enChIP. Using this approach, we achieved high yields of enChIP at the targeted genomic region. Our novel transgenic mouse lines provide a valuable tool for enChIP analysis in primary mouse cells.

KEYWORDS:

ChIP; chromatin immunoprecipitation; dCas9; enChIP; transgenic mouse

PMID:
29480524
DOI:
10.1111/gtc.12573
[Indexed for MEDLINE]
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