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Targeted disruption of exogenous EGFP gene in medaka using zinc‐finger nucleases
Author(s) -
Ansai Satoshi,
Ochiai Hiroshi,
Kanie Yuta,
Kamei Yasuhiro,
Gou Yuki,
Kitano Takeshi,
Yamamoto Takashi,
Kinoshita Masato
Publication year - 2012
Publication title -
development, growth and differentiation
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.864
H-Index - 66
eISSN - 1440-169X
pISSN - 0012-1592
DOI - 10.1111/j.1440-169x.2012.01357.x
Subject(s) - zinc finger nuclease , zinc finger , biology , green fluorescent protein , gene , somatic cell , oryzias , genetics , mutation , gene targeting , genome editing , transcription activator like effector nuclease , microbiology and biotechnology , crispr , transcription factor
Zinc‐finger nucleases ( ZFN s) are artificial enzymes that create site‐specific double‐strand breaks and thereby induce targeted genome editing. Here, we demonstrated successful gene disruption in somatic and germ cells of medaka ( O ryzias latipes ) using ZFN to target exogenous EGFP genes. Embryos that were injected with an RNA sequence pair coding for ZFN s showed mosaic loss of green fluorescent protein fluorescence in skeletal muscle. A number of mutations that included both deletions and insertions were identified within the ZFN target site in each embryo, whereas no mutations were found at the non‐targeted sites. In addition, ZFN ‐induced mutations were introduced in germ cells and efficiently transmitted to the next generation. The mutation frequency varied (6–100%) in the germ cells from each founder, and a founder carried more than two types of mutation in germ cells. Our results have introduced the possibility of targeted gene disruption and reverse genetics in medaka.