Cytoplasmic injection of murine zygotes with Sleeping Beauty transposon plasmids and minicircles results in the efficient generation of germline transgenic mice

Autores
Garrels, Wiebke; Talluri, Thirumala R.; Ziegler, Maren; Most, Ilka; Forcato, Diego Oscar; Schmeer, Marco; Schleef, Martin; Ivics, Zoltán; Kues, Wilfried A.
Año de publicación
2016
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Transgenesis in the mouse is an essential tool for the understanding of gene function and genome organization. Here, we describe a simplified microinjection protocol for efficient germline transgenesis and sustained transgene expression in the mouse model employing binary Sleeping Beauty transposon constructs of different topology. The protocol is based on co‐injection of supercoiled plasmids or minicircles, encoding the Sleeping Beauty transposase and a transposon construct, into the cytoplasm of murine zygotes. Importantly, this simplified injection avoids the mechanical penetration of the vulnerable pronuclear membrane, resulting in higher survival rates of treated embryos and a more rapid pace of injections. Upon translation of the transposase, transposase‐catalyzed transposition into the genome results in stable transgenic animals carrying monomeric transgenes. In summary, cytoplasmic injection of binary transposon constructs is a feasible, plasmid‐based, and simplified microinjection method to generate genetically modified mice. The modular design of the components allows the multiplexing of different transposons, and the generation of multi‐transposon transgenic mice in a single step.
Fil: Garrels, Wiebke. Friedrich Loeffler Institut; Alemania
Fil: Talluri, Thirumala R.. Friedrich Loeffler Institut; Alemania
Fil: Ziegler, Maren. Friedrich Loeffler Institut; Alemania
Fil: Most, Ilka. Friedrich Loeffler Institut; Alemania
Fil: Forcato, Diego Oscar. Friedrich Loeffler Institut; Alemania. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Biología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Schmeer, Marco. PlasmidFactory; Alemania
Fil: Schleef, Martin. PlasmidFactory; Alemania
Fil: Ivics, Zoltán. Paul Ehrlich Institute; Alemania
Fil: Kues, Wilfried A.. Friedrich Loeffler Institut; Alemania
Materia
Active Transgenesis
Genetic Engineering
Simplified Microinjection
Synthetic Biology
Transposition
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/69856
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/69856
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Cytoplasmic injection of murine zygotes with Sleeping Beauty transposon plasmids and minicircles results in the efficient generation of germline transgenic miceGarrels, WiebkeTalluri, Thirumala R.Ziegler, MarenMost, IlkaForcato, Diego OscarSchmeer, MarcoSchleef, MartinIvics, ZoltánKues, Wilfried A.Active TransgenesisGenetic EngineeringSimplified MicroinjectionSynthetic BiologyTranspositionhttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1Transgenesis in the mouse is an essential tool for the understanding of gene function and genome organization. Here, we describe a simplified microinjection protocol for efficient germline transgenesis and sustained transgene expression in the mouse model employing binary Sleeping Beauty transposon constructs of different topology. The protocol is based on co‐injection of supercoiled plasmids or minicircles, encoding the Sleeping Beauty transposase and a transposon construct, into the cytoplasm of murine zygotes. Importantly, this simplified injection avoids the mechanical penetration of the vulnerable pronuclear membrane, resulting in higher survival rates of treated embryos and a more rapid pace of injections. Upon translation of the transposase, transposase‐catalyzed transposition into the genome results in stable transgenic animals carrying monomeric transgenes. In summary, cytoplasmic injection of binary transposon constructs is a feasible, plasmid‐based, and simplified microinjection method to generate genetically modified mice. The modular design of the components allows the multiplexing of different transposons, and the generation of multi‐transposon transgenic mice in a single step.Fil: Garrels, Wiebke. Friedrich Loeffler Institut; AlemaniaFil: Talluri, Thirumala R.. Friedrich Loeffler Institut; AlemaniaFil: Ziegler, Maren. Friedrich Loeffler Institut; AlemaniaFil: Most, Ilka. Friedrich Loeffler Institut; AlemaniaFil: Forcato, Diego Oscar. Friedrich Loeffler Institut; Alemania. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Biología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Schmeer, Marco. PlasmidFactory; AlemaniaFil: Schleef, Martin. PlasmidFactory; AlemaniaFil: Ivics, Zoltán. Paul Ehrlich Institute; AlemaniaFil: Kues, Wilfried A.. Friedrich Loeffler Institut; AlemaniaWiley VCH Verlag2016-01info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/69856Garrels, Wiebke; Talluri, Thirumala R.; Ziegler, Maren; Most, Ilka; Forcato, Diego Oscar; et al.; Cytoplasmic injection of murine zygotes with Sleeping Beauty transposon plasmids and minicircles results in the efficient generation of germline transgenic mice; Wiley VCH Verlag; Biotechnology Journal; 11; 1; 1-2016; 178-1841860-7314CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/doi/10.1002/biot.201500218info:eu-repo/semantics/altIdentifier/url/https://onlinelibrary.wiley.com/doi/abs/10.1002/biot.201500218info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T09:44:56Zoai:ri.conicet.gov.ar:11336/69856instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-09-29 09:44:56.699CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Cytoplasmic injection of murine zygotes with Sleeping Beauty transposon plasmids and minicircles results in the efficient generation of germline transgenic mice
title Cytoplasmic injection of murine zygotes with Sleeping Beauty transposon plasmids and minicircles results in the efficient generation of germline transgenic mice
spellingShingle Cytoplasmic injection of murine zygotes with Sleeping Beauty transposon plasmids and minicircles results in the efficient generation of germline transgenic mice
Garrels, Wiebke
Active Transgenesis
Genetic Engineering
Simplified Microinjection
Synthetic Biology
Transposition
title_short Cytoplasmic injection of murine zygotes with Sleeping Beauty transposon plasmids and minicircles results in the efficient generation of germline transgenic mice
title_full Cytoplasmic injection of murine zygotes with Sleeping Beauty transposon plasmids and minicircles results in the efficient generation of germline transgenic mice
title_fullStr Cytoplasmic injection of murine zygotes with Sleeping Beauty transposon plasmids and minicircles results in the efficient generation of germline transgenic mice
title_full_unstemmed Cytoplasmic injection of murine zygotes with Sleeping Beauty transposon plasmids and minicircles results in the efficient generation of germline transgenic mice
title_sort Cytoplasmic injection of murine zygotes with Sleeping Beauty transposon plasmids and minicircles results in the efficient generation of germline transgenic mice
dc.creator.none.fl_str_mv Garrels, Wiebke
Talluri, Thirumala R.
Ziegler, Maren
Most, Ilka
Forcato, Diego Oscar
Schmeer, Marco
Schleef, Martin
Ivics, Zoltán
Kues, Wilfried A.
author Garrels, Wiebke
author_facet Garrels, Wiebke
Talluri, Thirumala R.
Ziegler, Maren
Most, Ilka
Forcato, Diego Oscar
Schmeer, Marco
Schleef, Martin
Ivics, Zoltán
Kues, Wilfried A.
author_role author
author2 Talluri, Thirumala R.
Ziegler, Maren
Most, Ilka
Forcato, Diego Oscar
Schmeer, Marco
Schleef, Martin
Ivics, Zoltán
Kues, Wilfried A.
author2_role author
author
author
author
author
author
author
author
dc.subject.none.fl_str_mv Active Transgenesis
Genetic Engineering
Simplified Microinjection
Synthetic Biology
Transposition
topic Active Transgenesis
Genetic Engineering
Simplified Microinjection
Synthetic Biology
Transposition
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv Transgenesis in the mouse is an essential tool for the understanding of gene function and genome organization. Here, we describe a simplified microinjection protocol for efficient germline transgenesis and sustained transgene expression in the mouse model employing binary Sleeping Beauty transposon constructs of different topology. The protocol is based on co‐injection of supercoiled plasmids or minicircles, encoding the Sleeping Beauty transposase and a transposon construct, into the cytoplasm of murine zygotes. Importantly, this simplified injection avoids the mechanical penetration of the vulnerable pronuclear membrane, resulting in higher survival rates of treated embryos and a more rapid pace of injections. Upon translation of the transposase, transposase‐catalyzed transposition into the genome results in stable transgenic animals carrying monomeric transgenes. In summary, cytoplasmic injection of binary transposon constructs is a feasible, plasmid‐based, and simplified microinjection method to generate genetically modified mice. The modular design of the components allows the multiplexing of different transposons, and the generation of multi‐transposon transgenic mice in a single step.
Fil: Garrels, Wiebke. Friedrich Loeffler Institut; Alemania
Fil: Talluri, Thirumala R.. Friedrich Loeffler Institut; Alemania
Fil: Ziegler, Maren. Friedrich Loeffler Institut; Alemania
Fil: Most, Ilka. Friedrich Loeffler Institut; Alemania
Fil: Forcato, Diego Oscar. Friedrich Loeffler Institut; Alemania. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Biología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Schmeer, Marco. PlasmidFactory; Alemania
Fil: Schleef, Martin. PlasmidFactory; Alemania
Fil: Ivics, Zoltán. Paul Ehrlich Institute; Alemania
Fil: Kues, Wilfried A.. Friedrich Loeffler Institut; Alemania
description Transgenesis in the mouse is an essential tool for the understanding of gene function and genome organization. Here, we describe a simplified microinjection protocol for efficient germline transgenesis and sustained transgene expression in the mouse model employing binary Sleeping Beauty transposon constructs of different topology. The protocol is based on co‐injection of supercoiled plasmids or minicircles, encoding the Sleeping Beauty transposase and a transposon construct, into the cytoplasm of murine zygotes. Importantly, this simplified injection avoids the mechanical penetration of the vulnerable pronuclear membrane, resulting in higher survival rates of treated embryos and a more rapid pace of injections. Upon translation of the transposase, transposase‐catalyzed transposition into the genome results in stable transgenic animals carrying monomeric transgenes. In summary, cytoplasmic injection of binary transposon constructs is a feasible, plasmid‐based, and simplified microinjection method to generate genetically modified mice. The modular design of the components allows the multiplexing of different transposons, and the generation of multi‐transposon transgenic mice in a single step.
publishDate 2016
dc.date.none.fl_str_mv 2016-01
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
http://purl.org/coar/resource_type/c_6501
info:ar-repo/semantics/articulo
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/69856
Garrels, Wiebke; Talluri, Thirumala R.; Ziegler, Maren; Most, Ilka; Forcato, Diego Oscar; et al.; Cytoplasmic injection of murine zygotes with Sleeping Beauty transposon plasmids and minicircles results in the efficient generation of germline transgenic mice; Wiley VCH Verlag; Biotechnology Journal; 11; 1; 1-2016; 178-184
1860-7314
CONICET Digital
CONICET
url http://hdl.handle.net/11336/69856
identifier_str_mv Garrels, Wiebke; Talluri, Thirumala R.; Ziegler, Maren; Most, Ilka; Forcato, Diego Oscar; et al.; Cytoplasmic injection of murine zygotes with Sleeping Beauty transposon plasmids and minicircles results in the efficient generation of germline transgenic mice; Wiley VCH Verlag; Biotechnology Journal; 11; 1; 1-2016; 178-184
1860-7314
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/doi/10.1002/biot.201500218
info:eu-repo/semantics/altIdentifier/url/https://onlinelibrary.wiley.com/doi/abs/10.1002/biot.201500218
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
dc.publisher.none.fl_str_mv Wiley VCH Verlag
publisher.none.fl_str_mv Wiley VCH Verlag
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
instname:Consejo Nacional de Investigaciones Científicas y Técnicas
reponame_str CONICET Digital (CONICET)
collection CONICET Digital (CONICET)
instname_str Consejo Nacional de Investigaciones Científicas y Técnicas
repository.name.fl_str_mv CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicas
repository.mail.fl_str_mv dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar
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score 13.070432

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