Generation of knock-in lampreys by CRISPR-Cas9-mediated genome engineering

Abstract The lamprey represents the oldest group of living vertebrates and has been a key organism in various research fields such as evolutionary developmental biology and neuroscience. However, no knock-in technique for this animal has been established yet, preventing application of advanced genet...

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Autores principales:	Daichi G. Suzuki, Hiroshi Wada, Shin-ichi Higashijima
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/30e6116cc3fb44749827c167ab5db9d5
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spelling	oai:doaj.org-article:30e6116cc3fb44749827c167ab5db9d52021-12-02T17:13:18ZGeneration of knock-in lampreys by CRISPR-Cas9-mediated genome engineering10.1038/s41598-021-99338-12045-2322https://doaj.org/article/30e6116cc3fb44749827c167ab5db9d52021-10-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-99338-1https://doaj.org/toc/2045-2322Abstract The lamprey represents the oldest group of living vertebrates and has been a key organism in various research fields such as evolutionary developmental biology and neuroscience. However, no knock-in technique for this animal has been established yet, preventing application of advanced genetic techniques. Here, we report efficient generation of F0 knock-in lampreys by CRISPR-Cas9-mediated genome editing. A donor plasmid containing a heat-shock promoter was co-injected with a short guide RNA (sgRNA) for genome digestion, a sgRNA for donor plasmid digestion, and Cas9 mRNA. Targeting different genetic loci, we succeeded in generating knock-in lampreys expressing photoconvertible protein Dendra2 as well as those expressing EGFP. With its simplicity, design flexibility, and high efficiency, we propose that the present method has great versatility for various experimental uses in lamprey research and that it can also be applied to other “non-model” organisms.Daichi G. SuzukiHiroshi WadaShin-ichi HigashijimaNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-8 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Daichi G. Suzuki Hiroshi Wada Shin-ichi Higashijima Generation of knock-in lampreys by CRISPR-Cas9-mediated genome engineering
description	Abstract The lamprey represents the oldest group of living vertebrates and has been a key organism in various research fields such as evolutionary developmental biology and neuroscience. However, no knock-in technique for this animal has been established yet, preventing application of advanced genetic techniques. Here, we report efficient generation of F0 knock-in lampreys by CRISPR-Cas9-mediated genome editing. A donor plasmid containing a heat-shock promoter was co-injected with a short guide RNA (sgRNA) for genome digestion, a sgRNA for donor plasmid digestion, and Cas9 mRNA. Targeting different genetic loci, we succeeded in generating knock-in lampreys expressing photoconvertible protein Dendra2 as well as those expressing EGFP. With its simplicity, design flexibility, and high efficiency, we propose that the present method has great versatility for various experimental uses in lamprey research and that it can also be applied to other “non-model” organisms.
format	article
author	Daichi G. Suzuki Hiroshi Wada Shin-ichi Higashijima
author_facet	Daichi G. Suzuki Hiroshi Wada Shin-ichi Higashijima
author_sort	Daichi G. Suzuki
title	Generation of knock-in lampreys by CRISPR-Cas9-mediated genome engineering
title_short	Generation of knock-in lampreys by CRISPR-Cas9-mediated genome engineering
title_full	Generation of knock-in lampreys by CRISPR-Cas9-mediated genome engineering
title_fullStr	Generation of knock-in lampreys by CRISPR-Cas9-mediated genome engineering
title_full_unstemmed	Generation of knock-in lampreys by CRISPR-Cas9-mediated genome engineering
title_sort	generation of knock-in lampreys by crispr-cas9-mediated genome engineering
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/30e6116cc3fb44749827c167ab5db9d5
work_keys_str_mv	AT daichigsuzuki generationofknockinlampreysbycrisprcas9mediatedgenomeengineering AT hiroshiwada generationofknockinlampreysbycrisprcas9mediatedgenomeengineering AT shinichihigashijima generationofknockinlampreysbycrisprcas9mediatedgenomeengineering
_version_	1718381311154978816

Generation of knock-in lampreys by CRISPR-Cas9-mediated genome engineering

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