Generation of genetically modified mice using SpCas9-NG engineered nuclease

Although genetically modified mice can be generated with high efficiency by using CRISPR/Cas9-mediated genome editing in mouse zygotes, only the loci with a protospacer-adjacent motif (PAM) sequence are targetable. The present study investigated the usability of engineered Streptococcus pyogenes Cas...

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Autores principales: Fujii, Wataru, Ito, Haruka, Kanke, Takuya, Ikeda, Arisa, Sugiura, Koji, Naito, Kunihiko
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group UK 2019
Materias:
Article
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6733909/
https://www.ncbi.nlm.nih.gov/pubmed/31501500
http://dx.doi.org/10.1038/s41598-019-49394-5
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author Fujii, Wataru
Ito, Haruka
Kanke, Takuya
Ikeda, Arisa
Sugiura, Koji
Naito, Kunihiko
author_facet Fujii, Wataru
Ito, Haruka
Kanke, Takuya
Ikeda, Arisa
Sugiura, Koji
Naito, Kunihiko
author_sort Fujii, Wataru
collection PubMed
description Although genetically modified mice can be generated with high efficiency by using CRISPR/Cas9-mediated genome editing in mouse zygotes, only the loci with a protospacer-adjacent motif (PAM) sequence are targetable. The present study investigated the usability of engineered Streptococcus pyogenes Cas9 (SpCas9-NG) in mouse zygotes. In addition to the 5′-NGG sequence, SpCas9-NG recognized the 5′-NGA, 5′-NGC and 5′-NGT sequences in mouse zygotes as PAMs that were appropriate for the generation of knockout mice. Moreover, SpCas9-NG-mediated genome editing enabled the generation of knock-in mice untargetable by the conventional SpCas9 in mouse zygotes. These results suggest that SpCas9-NG-mediated genome editing in zygotes is available for the generation of knockout and knock-in mice at the locus corresponding to NGN-PAM.
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spelling pubmed-67339092019-09-20 Generation of genetically modified mice using SpCas9-NG engineered nuclease Fujii, Wataru Ito, Haruka Kanke, Takuya Ikeda, Arisa Sugiura, Koji Naito, Kunihiko Sci Rep Article Although genetically modified mice can be generated with high efficiency by using CRISPR/Cas9-mediated genome editing in mouse zygotes, only the loci with a protospacer-adjacent motif (PAM) sequence are targetable. The present study investigated the usability of engineered Streptococcus pyogenes Cas9 (SpCas9-NG) in mouse zygotes. In addition to the 5′-NGG sequence, SpCas9-NG recognized the 5′-NGA, 5′-NGC and 5′-NGT sequences in mouse zygotes as PAMs that were appropriate for the generation of knockout mice. Moreover, SpCas9-NG-mediated genome editing enabled the generation of knock-in mice untargetable by the conventional SpCas9 in mouse zygotes. These results suggest that SpCas9-NG-mediated genome editing in zygotes is available for the generation of knockout and knock-in mice at the locus corresponding to NGN-PAM. Nature Publishing Group UK 2019-09-09 /pmc/articles/PMC6733909/ /pubmed/31501500 http://dx.doi.org/10.1038/s41598-019-49394-5 Text en © The Author(s) 2019 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.
spellingShingle Article
Fujii, Wataru
Ito, Haruka
Kanke, Takuya
Ikeda, Arisa
Sugiura, Koji
Naito, Kunihiko
Generation of genetically modified mice using SpCas9-NG engineered nuclease
title Generation of genetically modified mice using SpCas9-NG engineered nuclease
title_full Generation of genetically modified mice using SpCas9-NG engineered nuclease
title_fullStr Generation of genetically modified mice using SpCas9-NG engineered nuclease
title_full_unstemmed Generation of genetically modified mice using SpCas9-NG engineered nuclease
title_short Generation of genetically modified mice using SpCas9-NG engineered nuclease
title_sort generation of genetically modified mice using spcas9-ng engineered nuclease
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6733909/
https://www.ncbi.nlm.nih.gov/pubmed/31501500
http://dx.doi.org/10.1038/s41598-019-49394-5
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