Assessing Tn5 and sleeping beauty for transpositional transgenesis by cytoplasmic injection into bovine and ovine zygotes

Autores
Bevacqua, Romina Jimena; Fernández y Martin, Rafael; Canel, Natalia Gabriela; Gibbons, Alejandro Eduardo; Texeira, D.I.A.; Lange, F.; Vans Landschoot, Geraldina; Savy, Virginia; Briski, Olinda; Hiriart, María Inés; Grueso, Esther; Ivics, Zoltán; Taboga, Oscar Alberto; Kues, Wilfried A.; Ferraris, S.R.; Salamone, Daniel Felipe
Año de publicación
2017
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Transgenic domestic animals represent an alternative to bioreactors for large-scale production of biopharmaceuticals and could also provide more accurate biomedical models than rodents. However, their generation remains inefficient. Recently, DNA transposons allowed improved transgenesis efficiencies in mice and pigs. In this work, Tn5 and Sleeping Beauty (SB) transposon systems were evaluated for transgenesis by simple cytoplasmic injection in livestock zygotes. In the case of Tn5, the transposome complex of transposon nucleic acid and Tn5 protein was injected. In the case of SB, the supercoiled plasmids encoding a transposon and the SB transposase were co-injected. In vitro produced bovine zygotes were used to establish the cytoplasmic injection conditions. The in vitro cultured blastocysts were evaluated for reporter gene expression and genotyped. Subsequently, both transposon systems were injected in seasonally available ovine zygotes, employing transposons carrying the recombinant human factor IX driven by the beta-lactoglobulin promoter. The Tn5 approach did not result in transgenic lambs. In contrast, the Sleeping Beauty injection resulted in 2 lambs (29%) carrying the transgene. Both animals exhibited cellular mosaicism of the transgene. The extraembryonic tissues (placenta or umbilical cord) of three additional animals were also transgenic. These results show that transpositional transgenesis by cytoplasmic injection of SB transposon components can be applied for the production of transgenic lambs of pharmaceutical interest.
Instituto de Biotecnología
Fil: Bevacqua, Romina Jimena. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Fernández y Martín, Rafael. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Canel, Natalia Gabriela. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario; Argentina
Fil: Gibbons, Alejandro Eduardo. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Bariloche; Argentina
Fil: Texeira, D.I.A. Universidade Estadual do Ceará. Faculdade de Veterinária; Brasil
Fil: Lange, F. Universidad Maimónides. Laboratorio de Clonación y Transgenesis; Argentina
Fil: Vans Landschoot, Geraldina. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Fil: Lange, F. Universidad Maimónides. Laboratorio de Clonación y Transgenesis; Argentina
Fil: Savy, Virginia. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Briski, Olinda. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Hiriart, María Inés. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Grueso, Esther. Paul-Ehrlich-Institute; Alemania
Fil: Ivics, Zoltán. Paul-Ehrlich-Institute; Alemania
Fil: Taboga, Oscar Alberto. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina.
Fil: Kues, Wilfried A. Friedrich-Loeffler-Institut; Alemania
Fil: Ferraris, S.R. Universidad Maimónides. Laboratorio de Clonación y Transgenesis; Argentina
Fil: Salamone, Daniel Felipe. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fuente
PLoS ONE 12 (3) : e0174025
Materia
Ovinos
Bovinae
Cigotos
Transferencia de Genes
Inyección
Brucelosis
Sheep
Zygotes
Gene Transfer
Injection
Tn5
Nivel de accesibilidad
acceso abierto
Condiciones de uso
http://creativecommons.org/licenses/by-nc-sa/4.0/
Repositorio
INTA Digital (INTA)
Institución
Instituto Nacional de Tecnología Agropecuaria
OAI Identificador
oai:localhost:20.500.12123/559
Acceder
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spelling Assessing Tn5 and sleeping beauty for transpositional transgenesis by cytoplasmic injection into bovine and ovine zygotesBevacqua, Romina JimenaFernández y Martin, RafaelCanel, Natalia GabrielaGibbons, Alejandro EduardoTexeira, D.I.A.Lange, F.Vans Landschoot, GeraldinaSavy, VirginiaBriski, OlindaHiriart, María InésGrueso, EstherIvics, ZoltánTaboga, Oscar AlbertoKues, Wilfried A.Ferraris, S.R.Salamone, Daniel FelipeOvinosBovinaeCigotosTransferencia de GenesInyecciónBrucelosisSheepZygotesGene TransferInjectionTn5Transgenic domestic animals represent an alternative to bioreactors for large-scale production of biopharmaceuticals and could also provide more accurate biomedical models than rodents. However, their generation remains inefficient. Recently, DNA transposons allowed improved transgenesis efficiencies in mice and pigs. In this work, Tn5 and Sleeping Beauty (SB) transposon systems were evaluated for transgenesis by simple cytoplasmic injection in livestock zygotes. In the case of Tn5, the transposome complex of transposon nucleic acid and Tn5 protein was injected. In the case of SB, the supercoiled plasmids encoding a transposon and the SB transposase were co-injected. In vitro produced bovine zygotes were used to establish the cytoplasmic injection conditions. The in vitro cultured blastocysts were evaluated for reporter gene expression and genotyped. Subsequently, both transposon systems were injected in seasonally available ovine zygotes, employing transposons carrying the recombinant human factor IX driven by the beta-lactoglobulin promoter. The Tn5 approach did not result in transgenic lambs. In contrast, the Sleeping Beauty injection resulted in 2 lambs (29%) carrying the transgene. Both animals exhibited cellular mosaicism of the transgene. The extraembryonic tissues (placenta or umbilical cord) of three additional animals were also transgenic. These results show that transpositional transgenesis by cytoplasmic injection of SB transposon components can be applied for the production of transgenic lambs of pharmaceutical interest.Instituto de BiotecnologíaFil: Bevacqua, Romina Jimena. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Fernández y Martín, Rafael. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Canel, Natalia Gabriela. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario; ArgentinaFil: Gibbons, Alejandro Eduardo. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Bariloche; ArgentinaFil: Texeira, D.I.A. Universidade Estadual do Ceará. Faculdade de Veterinária; BrasilFil: Lange, F. Universidad Maimónides. Laboratorio de Clonación y Transgenesis; ArgentinaFil: Vans Landschoot, Geraldina. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Fil: Lange, F. Universidad Maimónides. Laboratorio de Clonación y Transgenesis; ArgentinaFil: Savy, Virginia. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Briski, Olinda. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Hiriart, María Inés. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Grueso, Esther. Paul-Ehrlich-Institute; AlemaniaFil: Ivics, Zoltán. Paul-Ehrlich-Institute; AlemaniaFil: Taboga, Oscar Alberto. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina.Fil: Kues, Wilfried A. Friedrich-Loeffler-Institut; AlemaniaFil: Ferraris, S.R. Universidad Maimónides. Laboratorio de Clonación y Transgenesis; ArgentinaFil: Salamone, Daniel Felipe. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina2017-07-05T13:25:14Z2017-07-05T13:25:14Z2017-03-16info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfhttp://hdl.handle.net/20.500.12123/559http://journals.plos.org/plosone/article/related?id=10.1371/journal.pone.0174025https://doi.org/10.1371/journal.pone.0174025PLoS ONE 12 (3) : e0174025reponame:INTA Digital (INTA)instname:Instituto Nacional de Tecnología Agropecuariaenginfo:eu-repo/semantics/openAccesshttp://creativecommons.org/licenses/by-nc-sa/4.0/Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0)2025-10-30T11:22:30Zoai:localhost:20.500.12123/559instacron:INTAInstitucionalhttp://repositorio.inta.gob.ar/Organismo científico-tecnológicoNo correspondehttp://repositorio.inta.gob.ar/oai/requesttripaldi.nicolas@inta.gob.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:l2025-10-30 11:22:30.66INTA Digital (INTA) - Instituto Nacional de Tecnología Agropecuariafalse
dc.title.none.fl_str_mv Assessing Tn5 and sleeping beauty for transpositional transgenesis by cytoplasmic injection into bovine and ovine zygotes
title Assessing Tn5 and sleeping beauty for transpositional transgenesis by cytoplasmic injection into bovine and ovine zygotes
spellingShingle Assessing Tn5 and sleeping beauty for transpositional transgenesis by cytoplasmic injection into bovine and ovine zygotes
Bevacqua, Romina Jimena
Ovinos
Bovinae
Cigotos
Transferencia de Genes
Inyección
Brucelosis
Sheep
Zygotes
Gene Transfer
Injection
Tn5
title_short Assessing Tn5 and sleeping beauty for transpositional transgenesis by cytoplasmic injection into bovine and ovine zygotes
title_full Assessing Tn5 and sleeping beauty for transpositional transgenesis by cytoplasmic injection into bovine and ovine zygotes
title_fullStr Assessing Tn5 and sleeping beauty for transpositional transgenesis by cytoplasmic injection into bovine and ovine zygotes
title_full_unstemmed Assessing Tn5 and sleeping beauty for transpositional transgenesis by cytoplasmic injection into bovine and ovine zygotes
title_sort Assessing Tn5 and sleeping beauty for transpositional transgenesis by cytoplasmic injection into bovine and ovine zygotes
dc.creator.none.fl_str_mv Bevacqua, Romina Jimena
Fernández y Martin, Rafael
Canel, Natalia Gabriela
Gibbons, Alejandro Eduardo
Texeira, D.I.A.
Lange, F.
Vans Landschoot, Geraldina
Savy, Virginia
Briski, Olinda
Hiriart, María Inés
Grueso, Esther
Ivics, Zoltán
Taboga, Oscar Alberto
Kues, Wilfried A.
Ferraris, S.R.
Salamone, Daniel Felipe
author Bevacqua, Romina Jimena
author_facet Bevacqua, Romina Jimena
Fernández y Martin, Rafael
Canel, Natalia Gabriela
Gibbons, Alejandro Eduardo
Texeira, D.I.A.
Lange, F.
Vans Landschoot, Geraldina
Savy, Virginia
Briski, Olinda
Hiriart, María Inés
Grueso, Esther
Ivics, Zoltán
Taboga, Oscar Alberto
Kues, Wilfried A.
Ferraris, S.R.
Salamone, Daniel Felipe
author_role author
author2 Fernández y Martin, Rafael
Canel, Natalia Gabriela
Gibbons, Alejandro Eduardo
Texeira, D.I.A.
Lange, F.
Vans Landschoot, Geraldina
Savy, Virginia
Briski, Olinda
Hiriart, María Inés
Grueso, Esther
Ivics, Zoltán
Taboga, Oscar Alberto
Kues, Wilfried A.
Ferraris, S.R.
Salamone, Daniel Felipe
author2_role author
author
author
author
author
author
author
author
author
author
author
author
author
author
author
dc.subject.none.fl_str_mv Ovinos
Bovinae
Cigotos
Transferencia de Genes
Inyección
Brucelosis
Sheep
Zygotes
Gene Transfer
Injection
Tn5
topic Ovinos
Bovinae
Cigotos
Transferencia de Genes
Inyección
Brucelosis
Sheep
Zygotes
Gene Transfer
Injection
Tn5
dc.description.none.fl_txt_mv Transgenic domestic animals represent an alternative to bioreactors for large-scale production of biopharmaceuticals and could also provide more accurate biomedical models than rodents. However, their generation remains inefficient. Recently, DNA transposons allowed improved transgenesis efficiencies in mice and pigs. In this work, Tn5 and Sleeping Beauty (SB) transposon systems were evaluated for transgenesis by simple cytoplasmic injection in livestock zygotes. In the case of Tn5, the transposome complex of transposon nucleic acid and Tn5 protein was injected. In the case of SB, the supercoiled plasmids encoding a transposon and the SB transposase were co-injected. In vitro produced bovine zygotes were used to establish the cytoplasmic injection conditions. The in vitro cultured blastocysts were evaluated for reporter gene expression and genotyped. Subsequently, both transposon systems were injected in seasonally available ovine zygotes, employing transposons carrying the recombinant human factor IX driven by the beta-lactoglobulin promoter. The Tn5 approach did not result in transgenic lambs. In contrast, the Sleeping Beauty injection resulted in 2 lambs (29%) carrying the transgene. Both animals exhibited cellular mosaicism of the transgene. The extraembryonic tissues (placenta or umbilical cord) of three additional animals were also transgenic. These results show that transpositional transgenesis by cytoplasmic injection of SB transposon components can be applied for the production of transgenic lambs of pharmaceutical interest.
Instituto de Biotecnología
Fil: Bevacqua, Romina Jimena. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Fernández y Martín, Rafael. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Canel, Natalia Gabriela. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario; Argentina
Fil: Gibbons, Alejandro Eduardo. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Bariloche; Argentina
Fil: Texeira, D.I.A. Universidade Estadual do Ceará. Faculdade de Veterinária; Brasil
Fil: Lange, F. Universidad Maimónides. Laboratorio de Clonación y Transgenesis; Argentina
Fil: Vans Landschoot, Geraldina. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Fil: Lange, F. Universidad Maimónides. Laboratorio de Clonación y Transgenesis; Argentina
Fil: Savy, Virginia. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Briski, Olinda. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Hiriart, María Inés. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Grueso, Esther. Paul-Ehrlich-Institute; Alemania
Fil: Ivics, Zoltán. Paul-Ehrlich-Institute; Alemania
Fil: Taboga, Oscar Alberto. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina.
Fil: Kues, Wilfried A. Friedrich-Loeffler-Institut; Alemania
Fil: Ferraris, S.R. Universidad Maimónides. Laboratorio de Clonación y Transgenesis; Argentina
Fil: Salamone, Daniel Felipe. Universidad de Buenos Aires. Facultad de Agronomía. Pabellón de Zootecnica. Laboratorio de Biotecnología Animal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
description Transgenic domestic animals represent an alternative to bioreactors for large-scale production of biopharmaceuticals and could also provide more accurate biomedical models than rodents. However, their generation remains inefficient. Recently, DNA transposons allowed improved transgenesis efficiencies in mice and pigs. In this work, Tn5 and Sleeping Beauty (SB) transposon systems were evaluated for transgenesis by simple cytoplasmic injection in livestock zygotes. In the case of Tn5, the transposome complex of transposon nucleic acid and Tn5 protein was injected. In the case of SB, the supercoiled plasmids encoding a transposon and the SB transposase were co-injected. In vitro produced bovine zygotes were used to establish the cytoplasmic injection conditions. The in vitro cultured blastocysts were evaluated for reporter gene expression and genotyped. Subsequently, both transposon systems were injected in seasonally available ovine zygotes, employing transposons carrying the recombinant human factor IX driven by the beta-lactoglobulin promoter. The Tn5 approach did not result in transgenic lambs. In contrast, the Sleeping Beauty injection resulted in 2 lambs (29%) carrying the transgene. Both animals exhibited cellular mosaicism of the transgene. The extraembryonic tissues (placenta or umbilical cord) of three additional animals were also transgenic. These results show that transpositional transgenesis by cytoplasmic injection of SB transposon components can be applied for the production of transgenic lambs of pharmaceutical interest.
publishDate 2017
dc.date.none.fl_str_mv 2017-07-05T13:25:14Z
2017-07-05T13:25:14Z
2017-03-16
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/20.500.12123/559
http://journals.plos.org/plosone/article/related?id=10.1371/journal.pone.0174025
https://doi.org/10.1371/journal.pone.0174025
url http://hdl.handle.net/20.500.12123/559
http://journals.plos.org/plosone/article/related?id=10.1371/journal.pone.0174025
https://doi.org/10.1371/journal.pone.0174025
dc.language.none.fl_str_mv eng
language eng
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
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Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0)
eu_rights_str_mv openAccess
rights_invalid_str_mv http://creativecommons.org/licenses/by-nc-sa/4.0/
Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0)
dc.format.none.fl_str_mv application/pdf
dc.source.none.fl_str_mv PLoS ONE 12 (3) : e0174025
reponame:INTA Digital (INTA)
instname:Instituto Nacional de Tecnología Agropecuaria
reponame_str INTA Digital (INTA)
collection INTA Digital (INTA)
instname_str Instituto Nacional de Tecnología Agropecuaria
repository.name.fl_str_mv INTA Digital (INTA) - Instituto Nacional de Tecnología Agropecuaria
repository.mail.fl_str_mv tripaldi.nicolas@inta.gob.ar
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