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Transgenic mouse lines expressing the 3x FLAG ‐ dC as9 protein for enCh IP analysis
Author(s) -
Fujita Toshitsugu,
Kitaura Fusako,
Oji Asami,
Tanigawa Naoki,
Yuno Miyuki,
Ikawa Masahito,
Taniuchi Ichiro,
Fujii Hodaka
Publication year - 2018
Publication title -
genes to cells
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.912
H-Index - 115
eISSN - 1365-2443
pISSN - 1356-9597
DOI - 10.1111/gtc.12573
Subject(s) - crispr , transgene , genetically modified mouse , biology , microbiology and biotechnology , gene , biochemistry
We developed the engineered DNA ‐binding molecule‐mediated chromatin immunoprecipitation (enCh IP ) technology to isolate specific genomic regions while retaining their molecular interactions. In enCh IP , 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 ( dC as9). The locus is then affinity‐purified to enable identification of associated molecules. In this study, we generated transgenic mice expressing 3x FLAG ‐tagged Streptococcus pyogenes dC as9 (3x FLAG ‐ dC as9) and retrovirally transduced gRNA into primary CD 4 + T cells from these mice for enCh IP . Using this approach, we achieved high yields of enCh IP at the targeted genomic region. Our novel transgenic mouse lines provide a valuable tool for enCh IP analysis in primary mouse cells.