In-Depth Characterization of greenflesh Tomato Mutants Obtained by CRISPR/Cas9 Editing: A Case Study With Implications for Breeding and Regulation
Gene editing has already proved itself as an invaluable tool for the generation of mutants for crop breeding, yet its ultimate impact on agriculture will depend on how crops generated by gene editing technologies are regulated, and on our ability to characterize the impact of mutations on plant phen...
| Autores: | , , , , , , , , , , , , , , , , , |
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| Tipo de recurso: | artículo |
| Estado: | Versión publicada |
| Fecha de publicación: | 2022 |
| País: | España |
| Institución: | Consejo Superior de Investigaciones Científicas (CSIC) |
| Repositorio: | DIGITAL.CSIC. Repositorio Institucional del CSIC |
| OAI Identifier: | oai:digital.csic.es:10261/280671 |
| Acceso en línea: | http://hdl.handle.net/10261/280671 https://api.elsevier.com/content/abstract/scopus_id/85134672851 |
| Access Level: | acceso abierto |
| Palabra clave: | CRISPR-Cas9 Breeding Greenflesh Nutritional quality Pathogen resistance Staygreen Tomato |
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oai:digital.csic.es:10261/280671 |
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In-Depth Characterization of greenflesh Tomato Mutants Obtained by CRISPR/Cas9 Editing: A Case Study With Implications for Breeding and Regulation Gianoglio, Silvia Comino, Cinzia Moglia, Andrea Acquadro, Alberto García-Carpintero, Víctor Diretto, Gianfranco Sevi, Filippo Rambla, José Luis Dono, Gabriella Valentino, Danila Moreno Giménez, Elena Fullana Pericàs, Mateu Conesa, Miguel Ángel Galmés, Jeroni Lanteri, Sergio Mazzucato, Andrea Orzáez, Diego Granell, Antonio CRISPR-Cas9 Breeding Greenflesh Nutritional quality Pathogen resistance Staygreen Tomato Gene editing has already proved itself as an invaluable tool for the generation of mutants for crop breeding, yet its ultimate impact on agriculture will depend on how crops generated by gene editing technologies are regulated, and on our ability to characterize the impact of mutations on plant phenotype. A starting operational strategy for evaluating gene editing-based approaches to plant breeding might consist of assessing the effect of the induced mutations in a crop- and locus-specific manner: this involves the analysis of editing efficiency in different cultivars of a crop, the assessment of potential off-target mutations, and a phenotypic evaluation of edited lines carrying different mutated alleles. Here, we targeted the GREENFLESH (GF) locus in two tomato cultivars ('MoneyMaker' and 'San Marzano') and evaluated the efficiency, specificity and mutation patterns associated with CRISPR/Cas9 activity for this gene. The GF locus encodes a Mg-dechelatase responsible for initiating chlorophyll degradation; in gf mutants, ripe fruits accumulate both carotenoids and chlorophylls. Phenotypic evaluations were conducted on two transgene-free T2 'MoneyMaker' gf lines with different mutant alleles (a small insertion of 1 nucleotide and a larger deletion of 123 bp). Both lines, in addition to reduced chlorophyll degradation, showed a notable increase in carotenoid and tocopherol levels during fruit ripening. Infection of gf leaves and fruits with Botrytis cinerea resulted in a significant reduction of infected area and pathogen proliferation compared to the wild type (WT). Our data indicates that the CRISPR/Cas9-mediated mutation of the GF locus in tomato is efficient, specific and reproducible and that the resulting phenotype is robust and consistent with previously characterized greenflesh mutants obtained with different breeding techniques, while also shedding light on novel traits such as vitamin E overaccumulation and pathogen resistance. This makes GF an appealing target for breeding tomato cultivars with improved features for cultivation, as well as consumer appreciation and health. This work was supported in part by the PID2019-108203RB-I00 project of the Spanish Plan Nacional I+D, Spanish Ministry of Science and Innovation and by HARNESSTOM Innovation Action EU-H2020-SFS-2020-1 (Contract number: 101000716). Finally, this work was supported by COST Actions ROXy CA18210 and EUROcarotene CA15136 for networking. Peer reviewed Frontiers Media http://hdl.handle.net/10261/280671 https://api.elsevier.com/content/abstract/scopus_id/85134672851 |
| title |
In-Depth Characterization of greenflesh Tomato Mutants Obtained by CRISPR/Cas9 Editing: A Case Study With Implications for Breeding and Regulation |
| spellingShingle |
In-Depth Characterization of greenflesh Tomato Mutants Obtained by CRISPR/Cas9 Editing: A Case Study With Implications for Breeding and Regulation Gianoglio, Silvia CRISPR-Cas9 Breeding Greenflesh Nutritional quality Pathogen resistance Staygreen Tomato |
| title_short |
In-Depth Characterization of greenflesh Tomato Mutants Obtained by CRISPR/Cas9 Editing: A Case Study With Implications for Breeding and Regulation |
| title_full |
In-Depth Characterization of greenflesh Tomato Mutants Obtained by CRISPR/Cas9 Editing: A Case Study With Implications for Breeding and Regulation |
| title_fullStr |
In-Depth Characterization of greenflesh Tomato Mutants Obtained by CRISPR/Cas9 Editing: A Case Study With Implications for Breeding and Regulation |
| title_full_unstemmed |
In-Depth Characterization of greenflesh Tomato Mutants Obtained by CRISPR/Cas9 Editing: A Case Study With Implications for Breeding and Regulation |
| title_sort |
In-Depth Characterization of greenflesh Tomato Mutants Obtained by CRISPR/Cas9 Editing: A Case Study With Implications for Breeding and Regulation |
| author |
Gianoglio, Silvia |
| author_facet |
Gianoglio, Silvia Comino, Cinzia Moglia, Andrea Acquadro, Alberto García-Carpintero, Víctor Diretto, Gianfranco Sevi, Filippo Rambla, José Luis Dono, Gabriella Valentino, Danila Moreno Giménez, Elena Fullana Pericàs, Mateu Conesa, Miguel Ángel Galmés, Jeroni Lanteri, Sergio Mazzucato, Andrea Orzáez, Diego Granell, Antonio |
| author_role |
author |
| author2 |
Comino, Cinzia Moglia, Andrea Acquadro, Alberto García-Carpintero, Víctor Diretto, Gianfranco Sevi, Filippo Rambla, José Luis Dono, Gabriella Valentino, Danila Moreno Giménez, Elena Fullana Pericàs, Mateu Conesa, Miguel Ángel Galmés, Jeroni Lanteri, Sergio Mazzucato, Andrea Orzáez, Diego Granell, Antonio |
| author2_role |
author author author author author author author author author author author author author author author author author |
| topic |
CRISPR-Cas9 Breeding Greenflesh Nutritional quality Pathogen resistance Staygreen Tomato |
| topic_facet |
CRISPR-Cas9 Breeding Greenflesh Nutritional quality Pathogen resistance Staygreen Tomato |
| description |
Gene editing has already proved itself as an invaluable tool for the generation of mutants for crop breeding, yet its ultimate impact on agriculture will depend on how crops generated by gene editing technologies are regulated, and on our ability to characterize the impact of mutations on plant phenotype. A starting operational strategy for evaluating gene editing-based approaches to plant breeding might consist of assessing the effect of the induced mutations in a crop- and locus-specific manner: this involves the analysis of editing efficiency in different cultivars of a crop, the assessment of potential off-target mutations, and a phenotypic evaluation of edited lines carrying different mutated alleles. Here, we targeted the GREENFLESH (GF) locus in two tomato cultivars ('MoneyMaker' and 'San Marzano') and evaluated the efficiency, specificity and mutation patterns associated with CRISPR/Cas9 activity for this gene. The GF locus encodes a Mg-dechelatase responsible for initiating chlorophyll degradation; in gf mutants, ripe fruits accumulate both carotenoids and chlorophylls. Phenotypic evaluations were conducted on two transgene-free T2 'MoneyMaker' gf lines with different mutant alleles (a small insertion of 1 nucleotide and a larger deletion of 123 bp). Both lines, in addition to reduced chlorophyll degradation, showed a notable increase in carotenoid and tocopherol levels during fruit ripening. Infection of gf leaves and fruits with Botrytis cinerea resulted in a significant reduction of infected area and pathogen proliferation compared to the wild type (WT). Our data indicates that the CRISPR/Cas9-mediated mutation of the GF locus in tomato is efficient, specific and reproducible and that the resulting phenotype is robust and consistent with previously characterized greenflesh mutants obtained with different breeding techniques, while also shedding light on novel traits such as vitamin E overaccumulation and pathogen resistance. This makes GF an appealing target for breeding tomato cultivars with improved features for cultivation, as well as consumer appreciation and health. |
| publishDate |
2022 |
| format |
article |
| status_str |
publishedVersion |
| url |
http://hdl.handle.net/10261/280671 https://api.elsevier.com/content/abstract/scopus_id/85134672851 |
| eu_rights_str_mv |
openAccess |
| publisher |
Frontiers Media |
| institution |
Consejo Superior de Investigaciones Científicas (CSIC) |
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DIGITAL.CSIC. Repositorio Institucional del CSIC |
| reponame_str |
DIGITAL.CSIC. Repositorio Institucional del CSIC |
| instname_str |
Consejo Superior de Investigaciones Científicas (CSIC) |
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1878432365705953280 |
| publishDateSort |
2022 |
| author_browse |
Acquadro, Alberto Comino, Cinzia Conesa, Miguel Ángel Diretto, Gianfranco Dono, Gabriella Fullana Pericàs, Mateu Galmés, Jeroni García-Carpintero, Víctor Gianoglio, Silvia Granell, Antonio Lanteri, Sergio Mazzucato, Andrea Moglia, Andrea Moreno Giménez, Elena Orzáez, Diego Rambla, José Luis Sevi, Filippo Valentino, Danila |
| publisherStr |
Frontiers Media |
| score |
6.9303427 |