CLICK: One-step generation of conditional knockout mice

Yoshiki Miyasaka; Yoshihiro Uno; Kazuto Yoshimi; Yayoi Kunihiro; Takuji Yoshimura; Tomohiro Tanaka; Harumi Ishikubo; Yuichi Hiraoka; Norihiko Takemoto; Takao Tanaka; Yoshihiro Ooguchi; Paul Skehel; Tomomi Aida; Junji Takeda; Tomoji Mashimo

doi:10.1186/s12864-018-4713-y

CLICK: One-step generation of conditional knockout mice

Yoshiki Miyasaka, Yoshihiro Uno, Kazuto Yoshimi, Yayoi Kunihiro, Takuji Yoshimura, Tomohiro Tanaka, Harumi Ishikubo, Yuichi Hiraoka, Norihiko Takemoto, Takao Tanaka, Yoshihiro Ooguchi, Paul Skehel, Tomomi Aida, Junji Takeda, Tomoji Mashimo^*

^*Corresponding author for this work

Deanery of Biomedical Sciences

Research output: Contribution to journal › Article › peer-review

Abstract / Description of output

Background: CRISPR/Cas9 enables the targeting of genes in zygotes; however, efficient approaches to create loxP-flanked (floxed) alleles remain elusive. Results: Here, we show that the electroporation of Cas9, two gRNAs, and long single-stranded DNA (lssDNA) into zygotes, termed CLICK (CRISPR with lssDNA inducing conditional knockout alleles), enables the quick generation of floxed alleles in mice and rats. Conclusions: The high efficiency of CLICK provides homozygous knock-ins in oocytes carrying tissue-specific Cre, which allows the one-step generation of conditional knockouts in founder (F0) mice.

Original language	English
Article number	318
Journal	BMC Genomics
Volume	19
Issue number	1
DOIs	https://doi.org/10.1186/s12864-018-4713-y
Publication status	Published - 2 May 2018

Keywords / Materials (for Non-textual outputs)

CLICK
Cre-loxP system
CRISPR/Cas9
Long single-stranded DNA
Zygote electroporation

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10.1186/s12864-018-4713-y

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Cite this

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title = "CLICK: One-step generation of conditional knockout mice",

abstract = "Background: CRISPR/Cas9 enables the targeting of genes in zygotes; however, efficient approaches to create loxP-flanked (floxed) alleles remain elusive. Results: Here, we show that the electroporation of Cas9, two gRNAs, and long single-stranded DNA (lssDNA) into zygotes, termed CLICK (CRISPR with lssDNA inducing conditional knockout alleles), enables the quick generation of floxed alleles in mice and rats. Conclusions: The high efficiency of CLICK provides homozygous knock-ins in oocytes carrying tissue-specific Cre, which allows the one-step generation of conditional knockouts in founder (F0) mice.",

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doi = "10.1186/s12864-018-4713-y",

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T2 - One-step generation of conditional knockout mice

AU - Miyasaka, Yoshiki

AU - Uno, Yoshihiro

AU - Yoshimi, Kazuto

AU - Kunihiro, Yayoi

AU - Yoshimura, Takuji

AU - Tanaka, Tomohiro

AU - Ishikubo, Harumi

AU - Hiraoka, Yuichi

AU - Takemoto, Norihiko

AU - Tanaka, Takao

AU - Ooguchi, Yoshihiro

AU - Skehel, Paul

AU - Aida, Tomomi

AU - Takeda, Junji

AU - Mashimo, Tomoji

PY - 2018/5/2

Y1 - 2018/5/2

N2 - Background: CRISPR/Cas9 enables the targeting of genes in zygotes; however, efficient approaches to create loxP-flanked (floxed) alleles remain elusive. Results: Here, we show that the electroporation of Cas9, two gRNAs, and long single-stranded DNA (lssDNA) into zygotes, termed CLICK (CRISPR with lssDNA inducing conditional knockout alleles), enables the quick generation of floxed alleles in mice and rats. Conclusions: The high efficiency of CLICK provides homozygous knock-ins in oocytes carrying tissue-specific Cre, which allows the one-step generation of conditional knockouts in founder (F0) mice.

AB - Background: CRISPR/Cas9 enables the targeting of genes in zygotes; however, efficient approaches to create loxP-flanked (floxed) alleles remain elusive. Results: Here, we show that the electroporation of Cas9, two gRNAs, and long single-stranded DNA (lssDNA) into zygotes, termed CLICK (CRISPR with lssDNA inducing conditional knockout alleles), enables the quick generation of floxed alleles in mice and rats. Conclusions: The high efficiency of CLICK provides homozygous knock-ins in oocytes carrying tissue-specific Cre, which allows the one-step generation of conditional knockouts in founder (F0) mice.

KW - CLICK

KW - Cre-loxP system

KW - CRISPR/Cas9

KW - Long single-stranded DNA

KW - Zygote electroporation

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SN - 1471-2164

VL - 19

JO - BMC Genomics

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M1 - 318

ER -

CLICK: One-step generation of conditional knockout mice

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Keywords / Materials (for Non-textual outputs)

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