CRISPR/Cas9 mediated genome editing in ES cells and its application for chimeric analysis in mice

Oji, Asami; Noda, Taichi; Fujihara, Yoshitaka; Miyata, Haruhiko; Kim, Yeon Joo; Muto, Masanaga; Nozawa, Kaori; Matsumura, Takafumi; Isotani, Ayako; Ikawa, Masahito

doi:https://doi.org/10.1038/srep31666

CRISPR/Cas9 mediated genome editing in ES cells and its application for chimeric analysis in mice

Oji, Asami; Noda, Taichi; Fujihara, Yoshitaka et al.

Scientific Reports, 2016, 6, 31666

Number of Access:987（2025-08-22 01:06 Counts）

Identifier to cite or link to this item: https://hdl.handle.net/11094/78585

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Title	CRISPR/Cas9 mediated genome editing in ES cells and its application for chimeric analysis in mice
Creator	NRID 1000050803417 1000050803417 CreatorName Oji, Asami
	NRID 1000050712551 1000050712551 CreatorName Noda, Taichi
	NRID 1000070578848 1000070578848 CreatorName Fujihara, Yoshitaka
	NRID 1000050604732 1000050604732 researchmap permalink hmiyata hmiyata Scopus Author ID 37161837000 37161837000 ORCID 0000-0003-4758-5803 0000-0003-4758-5803 CreatorName Miyata, Haruhiko
	CreatorName Kim, Yeon Joo
	CreatorName Muto, Masanaga
	CreatorName Nozawa, Kaori
	CreatorName Matsumura, Takafumi
	NRID 1000020444523 1000020444523 CreatorName Isotani, Ayako
	NRID 1000020304066 1000020304066 ORCID 0000-0001-9859-6217 0000-0001-9859-6217 researchmap permalink read0054910 read0054910 Scopus Author ID 16689054500 16689054500 CreatorName Ikawa, Masahito
Description	Oji, A., Noda, T., Fujihara, Y. et al. CRISPR/Cas9 mediated genome editing in ES cells and its application for chimeric analysis in mice. Sci Rep 6, 31666 (2016). https://doi.org/10.1038/srep31666
Abstract	Targeted gene disrupted mice can be efficiently generated by expressing a single guide RNA (sgRNA)/CAS9 complex in the zygote. However, the limited success of complicated genome editing, such as large deletions, point mutations, and knockins, remains to be improved. Further, the mosaicism in founder generations complicates the genotypic and phenotypic analyses in these animals. Here we show that large deletions with two sgRNAs as well as dsDNA-mediated point mutations are efficient in mouse embryonic stem cells (ESCs). The dsDNA-mediated gene knockins are also feasible in ESCs. Finally, we generated chimeric mice with biallelic mutant ESCs for a lethal gene, Dnajb13, and analyzed their phenotypes. Not only was the lethal phenotype of hydrocephalus suppressed, but we also found that Dnajb13 is required for sperm cilia formation. The combination of biallelic genome editing in ESCs and subsequent chimeric analysis provides a useful tool for rapid gene function analysis in the whole organism.
Publisher	Nature Research
Source Title	Scientific Reports
Volume (Issue)	6
Page	31666
Date of Issued	2016-08-17
Language	English
Handle URL	https://hdl.handle.net/11094/78585
eISSN	20452322
Relation.isIdenticalTo	DOI (Publisher Version) https://doi.org/10.1038/srep31666
Relation.isIdenticalTo	PMID 27530713
Access Rights	open access
CopyRight	© 2016 The Author(s). This article is licensed under a Creative Commons Attribution 4.0 International License.
oaire:version	Version of Record
Category	学術雑誌論文 Journal Article

Resource Type	journal article
Local Type	Journal Article
DCTERMS.bibliographicCitation	Scientific Reports. 2016, 6, p. 31666
DC.title	CRISPR/Cas9 mediated genome editing in ES cells and its application for chimeric analysis in mice
DC.creator	Oji, Asami
	Noda, Taichi
	Fujihara, Yoshitaka
	Miyata, Haruhiko
	Kim, Yeon Joo
	Muto, Masanaga
	Nozawa, Kaori
	Matsumura, Takafumi
	Isotani, Ayako
	Ikawa, Masahito
DC.publisher	Nature Research
DC.language' scheme='DCTERMS.RFC1766	English
DCTERMS.issued' scheme='DCTERMS.W3CDTF	2016-08-17
DC.identifier	https://doi.org/10.1038/srep31666
DC.description	Oji, A., Noda, T., Fujihara, Y. et al. CRISPR/Cas9 mediated genome editing in ES cells and its application for chimeric analysis in mice. Sci Rep 6, 31666 (2016). https://doi.org/10.1038/srep31666
DCTERMS.abstract	Targeted gene disrupted mice can be efficiently generated by expressing a single guide RNA (sgRNA)/CAS9 complex in the zygote. However, the limited success of complicated genome editing, such as large deletions, point mutations, and knockins, remains to be improved. Further, the mosaicism in founder generations complicates the genotypic and phenotypic analyses in these animals. Here we show that large deletions with two sgRNAs as well as dsDNA-mediated point mutations are efficient in mouse embryonic stem cells (ESCs). The dsDNA-mediated gene knockins are also feasible in ESCs. Finally, we generated chimeric mice with biallelic mutant ESCs for a lethal gene, Dnajb13, and analyzed their phenotypes. Not only was the lethal phenotype of hydrocephalus suppressed, but we also found that Dnajb13 is required for sperm cilia formation. The combination of biallelic genome editing in ESCs and subsequent chimeric analysis provides a useful tool for rapid gene function analysis in the whole organism.
DC.rights	© 2016 The Author(s). This article is licensed under a Creative Commons Attribution 4.0 International License.
DC.format	78585
citation_title	CRISPR/Cas9 mediated genome editing in ES cells and its application for chimeric analysis in mice
citation_author	Oji, Asami
	Noda, Taichi
	Fujihara, Yoshitaka
	Miyata, Haruhiko
	Kim, Yeon Joo
	Muto, Masanaga
	Nozawa, Kaori
	Matsumura, Takafumi
	Isotani, Ayako
	Ikawa, Masahito
citation_publisher	Nature Research
citation_language	English
citation_date	2016-08-17
citation_journal_title	Scientific Reports
citation_volume	6
citation_firstpage	31666
citation_doi	https://doi.org/10.1038/srep31666
citation_pmid	27530713
citation_public_url	https://hdl.handle.net/11094/78585

CRISPR/Cas9 mediated genome editing in ES cells and its application for chimeric analysis in mice

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