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Resumen
Delivery of the CRISPR/Cas9 components to the plant cells is a key step in its application as a genome editing tool. Here, we compared Agrobacterium-mediated transformation and protoplast transfection with CRISPR/Cas9 components for potato genome editing. Two sgRNAs were designed to simultaneously direct Cas9 to the StPPO2 gene encoding for a tuber polyphenol oxidase (PPO). A binary vector (CR-PPO) was utilized for either Agrobacterium-mediated [ver mas...]
dc.contributor.authorGonzalez, Matías Nicolás
dc.contributor.authorMassa, Gabriela Alejandra
dc.contributor.authorAnderson, Mariette
dc.contributor.authorDecima Oneto, Cecilia Andrea
dc.contributor.authorTuresson, Helle
dc.contributor.authorOlsson, Niklas
dc.contributor.authorFält, Ann Sofie
dc.contributor.authorHofvander, Per
dc.contributor.authorFeingold, Sergio Enrique
dc.date.accessioned2021-04-28T10:45:48Z
dc.date.available2021-04-28T10:45:48Z
dc.date.issued2021-01-25
dc.identifier.issn1573-5044 (online)
dc.identifier.issn0167-6857 (print)
dc.identifier.otherhttps://doi.org/10.1007/s11240-020-02008-9
dc.identifier.urihttp://hdl.handle.net/20.500.12123/9204
dc.identifier.urihttps://link.springer.com/article/10.1007%2Fs11240-020-02008-9
dc.description.abstractDelivery of the CRISPR/Cas9 components to the plant cells is a key step in its application as a genome editing tool. Here, we compared Agrobacterium-mediated transformation and protoplast transfection with CRISPR/Cas9 components for potato genome editing. Two sgRNAs were designed to simultaneously direct Cas9 to the StPPO2 gene encoding for a tuber polyphenol oxidase (PPO). A binary vector (CR-PPO) was utilized for either Agrobacterium-mediated transformation or for transient expression in protoplasts, while ribonucleoprotein complexes (RNP-PPO) were additionally assayed in protoplasts. Editing efficiency varied, yielding 9.6%, 18.4% and 31.9% of edited lines from Agrobacterium-mediated transformation, RNP-PPO and CR-PPO transient expression in protoplasts, respectively. Furthermore, only the CR-PPO transient expression resulted in lines edited in all four StPPO2 alleles, observed in 46% of the edited lines and confirmed by tuber PPO activity and enzymatic browning analysis. Lines with on-target DNA insertions were found from all three approaches and characterized by sequencing. The dual-sgRNA strategy resulted in a low incidence of the targeted deletion, likely due to contrasting efficiencies between sgRNAs, that was partially evident in the in silico analysis. Our results demonstrate that gene editing efficiency in potato depends on the CRISPR/Cas9 delivery strategy and provide insights to consider when selecting the appropriate methodology.eng
dc.formatapplication/pdfes_AR
dc.language.isoenges_AR
dc.publisherSpringer Netherlandses_AR
dc.relationinfo:eu-repograntAgreement/INTA/PNBIO-1131024/AR./Desarrollo de sistemas alternativos de generación y utilización de variabilidad genética y su aplicación al mejoramiento de los cultivos.
dc.relationinfo:eu-repograntAgreement/INTA/2019-PE-E6-I115-001/2019-PE-E6-I115-001/AR./Edición génica, transgénesis y mutagénesis como generadores de nueva variabilidad en especies de interés agropecuario
dc.rightsinfo:eu-repo/semantics/restrictedAccesses_AR
dc.sourcePlant Cell, Tissue and Organ Culture 145 : 291-305 (2021)es_AR
dc.subjectPapaes_AR
dc.subjectPotatoeseng
dc.subjectEdición de Geneses_AR
dc.subjectGene Editingeng
dc.subjectProtoplastoses_AR
dc.subjectProtoplastseng
dc.subjectAgrobacterium Tumefacienses_AR
dc.subjectCatecol Oxidasaes_AR
dc.subjectCatechol Oxidaseeng
dc.titleComparative potato genome editing: Agrobacterium tumefaciens-mediated transformation and protoplasts transfection delivery of CRISPR/Cas9 components directed to StPPO2 genees_AR
dc.typeinfo:ar-repo/semantics/artículoes_AR
dc.typeinfo:eu-repo/semantics/articlees_AR
dc.typeinfo:eu-repo/semantics/publishedVersiones_AR
dc.description.origenEEA Balcarcees_AR
dc.description.filFil: González, Matías Nicolás. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Innovación para la Producción Agropecuaria y el Desarrollo Sostenible; Argentina.es_AR
dc.description.filFil: Massa, Gabriela Alejandra. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Innovación para la Producción Agropecuaria y el Desarrollo Sostenible; Argentina.es_AR
dc.description.filFil: Massa, Gabriela Alejandra. Universidad de Buenos Aires. Facultad de Agronomía; Argentina.es_AR
dc.description.filFil: Anderson, Mariette. Swedish University of Agricultural Sciences. Department of Plant Breeding; Suecia.es_AR
dc.description.filFil: Décima Oneto, Cecilia Andrea. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Innovación para la Producción Agropecuaria y el Desarrollo Sostenible; Argentina.es_AR
dc.description.filFil: Décima Oneto, Cecilia Andrea. Universidad de Buenos Aires. Facultad de Agronomía; Argentina.es_AR
dc.description.filFil: Turesson, Helle. Swedish University of Agricultural Sciences. Department of Plant Breeding; Suecia.es_AR
dc.description.filFil: Storani, Leonardo. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Innovación para la Producción Agropecuaria y el Desarrollo Sostenible; Argentina.es_AR
dc.description.filFil: Olsson, Niklas. Swedish University of Agricultural Sciences. Department of Plant Breeding; Suecia.es_AR
dc.description.filFil: Fält , Ann Sofie. Swedish University of Agricultural Sciences. Department of Plant Breeding; Suecia.es_AR
dc.description.filFil: Hofvander, Per. Swedish University of Agricultural Sciences. Department of Plant Breeding; Suecia.es_AR
dc.description.filFil: Feingold, Sergio Enrique. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Innovación para la Producción Agropecuaria y el Desarrollo Sostenible; Argentina.es_AR
dc.subtypecientifico


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