Endogenous zebrafish proneural Cre drivers generated by CRISPR/Cas9 short homology directed targeted integration

Abstract We previously reported efficient precision targeted integration of reporter DNA in zebrafish and human cells using CRISPR/Cas9 and short regions of homology. Here, we apply this strategy to isolate zebrafish Cre recombinase drivers whose spatial and temporal restricted expression mimics end...

Descripción completa

Guardado en:

Detalles Bibliográficos
Autores principales:	Maira P. Almeida, Jordan M. Welker, Sahiba Siddiqui, Jon Luiken, Stephen C. Ekker, Karl J. Clark, Jeffrey J. Essner, Maura McGrail
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/b65caa7fc4ef4e09a6d1cd7e35dfe5c2
Etiquetas:	Agregar Etiqueta Sin Etiquetas, Sea el primero en etiquetar este registro!

id	oai:doaj.org-article:b65caa7fc4ef4e09a6d1cd7e35dfe5c2
record_format	dspace
spelling	oai:doaj.org-article:b65caa7fc4ef4e09a6d1cd7e35dfe5c22021-12-02T15:23:38ZEndogenous zebrafish proneural Cre drivers generated by CRISPR/Cas9 short homology directed targeted integration10.1038/s41598-021-81239-y2045-2322https://doaj.org/article/b65caa7fc4ef4e09a6d1cd7e35dfe5c22021-01-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-81239-yhttps://doaj.org/toc/2045-2322Abstract We previously reported efficient precision targeted integration of reporter DNA in zebrafish and human cells using CRISPR/Cas9 and short regions of homology. Here, we apply this strategy to isolate zebrafish Cre recombinase drivers whose spatial and temporal restricted expression mimics endogenous genes. A 2A-Cre recombinase transgene with 48 bp homology arms was targeted into proneural genes ascl1b, olig2 and neurod1. We observed high rates of germline transmission ranging from 10 to 100% (2/20 olig2; 1/5 neurod1; 3/3 ascl1b). The transgenic lines Tg(ascl1b-2A-Cre) is75 , Tg(olig2-2A-Cre) is76 , and Tg(neurod1-2A-Cre) is77 expressed functional Cre recombinase in the expected proneural cell populations. Somatic targeting of 2A-CreERT2 into neurod1 resulted in tamoxifen responsive recombination in the nervous system. The results demonstrate Cre recombinase expression is driven by the native promoter and regulatory elements of the targeted genes. This approach provides a straightforward, efficient, and cost-effective method to generate cell type specific zebrafish Cre and CreERT2 drivers, overcoming challenges associated with promoter-BAC and transposon mediated transgenics.Maira P. AlmeidaJordan M. WelkerSahiba SiddiquiJon LuikenStephen C. EkkerKarl J. ClarkJeffrey J. EssnerMaura McGrailNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-12 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Maira P. Almeida Jordan M. Welker Sahiba Siddiqui Jon Luiken Stephen C. Ekker Karl J. Clark Jeffrey J. Essner Maura McGrail Endogenous zebrafish proneural Cre drivers generated by CRISPR/Cas9 short homology directed targeted integration
description	Abstract We previously reported efficient precision targeted integration of reporter DNA in zebrafish and human cells using CRISPR/Cas9 and short regions of homology. Here, we apply this strategy to isolate zebrafish Cre recombinase drivers whose spatial and temporal restricted expression mimics endogenous genes. A 2A-Cre recombinase transgene with 48 bp homology arms was targeted into proneural genes ascl1b, olig2 and neurod1. We observed high rates of germline transmission ranging from 10 to 100% (2/20 olig2; 1/5 neurod1; 3/3 ascl1b). The transgenic lines Tg(ascl1b-2A-Cre) is75 , Tg(olig2-2A-Cre) is76 , and Tg(neurod1-2A-Cre) is77 expressed functional Cre recombinase in the expected proneural cell populations. Somatic targeting of 2A-CreERT2 into neurod1 resulted in tamoxifen responsive recombination in the nervous system. The results demonstrate Cre recombinase expression is driven by the native promoter and regulatory elements of the targeted genes. This approach provides a straightforward, efficient, and cost-effective method to generate cell type specific zebrafish Cre and CreERT2 drivers, overcoming challenges associated with promoter-BAC and transposon mediated transgenics.
format	article
author	Maira P. Almeida Jordan M. Welker Sahiba Siddiqui Jon Luiken Stephen C. Ekker Karl J. Clark Jeffrey J. Essner Maura McGrail
author_facet	Maira P. Almeida Jordan M. Welker Sahiba Siddiqui Jon Luiken Stephen C. Ekker Karl J. Clark Jeffrey J. Essner Maura McGrail
author_sort	Maira P. Almeida
title	Endogenous zebrafish proneural Cre drivers generated by CRISPR/Cas9 short homology directed targeted integration
title_short	Endogenous zebrafish proneural Cre drivers generated by CRISPR/Cas9 short homology directed targeted integration
title_full	Endogenous zebrafish proneural Cre drivers generated by CRISPR/Cas9 short homology directed targeted integration
title_fullStr	Endogenous zebrafish proneural Cre drivers generated by CRISPR/Cas9 short homology directed targeted integration
title_full_unstemmed	Endogenous zebrafish proneural Cre drivers generated by CRISPR/Cas9 short homology directed targeted integration
title_sort	endogenous zebrafish proneural cre drivers generated by crispr/cas9 short homology directed targeted integration
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/b65caa7fc4ef4e09a6d1cd7e35dfe5c2
work_keys_str_mv	AT mairapalmeida endogenouszebrafishproneuralcredriversgeneratedbycrisprcas9shorthomologydirectedtargetedintegration AT jordanmwelker endogenouszebrafishproneuralcredriversgeneratedbycrisprcas9shorthomologydirectedtargetedintegration AT sahibasiddiqui endogenouszebrafishproneuralcredriversgeneratedbycrisprcas9shorthomologydirectedtargetedintegration AT jonluiken endogenouszebrafishproneuralcredriversgeneratedbycrisprcas9shorthomologydirectedtargetedintegration AT stephencekker endogenouszebrafishproneuralcredriversgeneratedbycrisprcas9shorthomologydirectedtargetedintegration AT karljclark endogenouszebrafishproneuralcredriversgeneratedbycrisprcas9shorthomologydirectedtargetedintegration AT jeffreyjessner endogenouszebrafishproneuralcredriversgeneratedbycrisprcas9shorthomologydirectedtargetedintegration AT mauramcgrail endogenouszebrafishproneuralcredriversgeneratedbycrisprcas9shorthomologydirectedtargetedintegration
_version_	1718387287238115328

Endogenous zebrafish proneural Cre drivers generated by CRISPR/Cas9 short homology directed targeted integration

Ejemplares similares