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Targeted Mutagenesis of the Female-Suppressor SyGI Gene in Tetraploid Kiwifruit by CRISPR/CAS9
Kiwifruit belong to the genus Actinidia with 54 species apparently all functionally dioecious. The sex-determinants of the type XX/XY, with male heterogametic, operate independently of the ploidy level. Recently, the SyGI protein has been described as the suppressor of female development. In the pre...
Autores principales: | , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7823651/ https://www.ncbi.nlm.nih.gov/pubmed/33396671 http://dx.doi.org/10.3390/plants10010062 |
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author | De Mori, Gloria Zaina, Giusi Franco-Orozco, Barbara Testolin, Raffaele De Paoli, Emanuele Cipriani, Guido |
author_facet | De Mori, Gloria Zaina, Giusi Franco-Orozco, Barbara Testolin, Raffaele De Paoli, Emanuele Cipriani, Guido |
author_sort | De Mori, Gloria |
collection | PubMed |
description | Kiwifruit belong to the genus Actinidia with 54 species apparently all functionally dioecious. The sex-determinants of the type XX/XY, with male heterogametic, operate independently of the ploidy level. Recently, the SyGI protein has been described as the suppressor of female development. In the present study, we exploited the CRISPR/Cas9 technology by targeting two different sites in the SyGI gene in order to induce a stable gene knock-out in two tetraploid male accessions of Actinidia chinensis var. chinensis. The two genotypes showed a regenerative efficiency of 58% and 73%, respectively. Despite not yet being able to verify the phenotypic effects on the flower structure, due to the long time required by tissue-cultured kiwifruit plants to flower, we obtained two regenerated lines showing near fixation of a unique modification in their genome, resulting in both cases in the onset of a premature stop codon, which induces the putative gene knock-out. Evaluation of gRNA1 locus for both regenerated plantlets resulted in co-amplification of a minor variant differing from the target region for a single nucleotide. A genomic duplication of the region in proximity of the Y genomic region could be postulated. |
format | Online Article Text |
id | pubmed-7823651 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-78236512021-01-24 Targeted Mutagenesis of the Female-Suppressor SyGI Gene in Tetraploid Kiwifruit by CRISPR/CAS9 De Mori, Gloria Zaina, Giusi Franco-Orozco, Barbara Testolin, Raffaele De Paoli, Emanuele Cipriani, Guido Plants (Basel) Article Kiwifruit belong to the genus Actinidia with 54 species apparently all functionally dioecious. The sex-determinants of the type XX/XY, with male heterogametic, operate independently of the ploidy level. Recently, the SyGI protein has been described as the suppressor of female development. In the present study, we exploited the CRISPR/Cas9 technology by targeting two different sites in the SyGI gene in order to induce a stable gene knock-out in two tetraploid male accessions of Actinidia chinensis var. chinensis. The two genotypes showed a regenerative efficiency of 58% and 73%, respectively. Despite not yet being able to verify the phenotypic effects on the flower structure, due to the long time required by tissue-cultured kiwifruit plants to flower, we obtained two regenerated lines showing near fixation of a unique modification in their genome, resulting in both cases in the onset of a premature stop codon, which induces the putative gene knock-out. Evaluation of gRNA1 locus for both regenerated plantlets resulted in co-amplification of a minor variant differing from the target region for a single nucleotide. A genomic duplication of the region in proximity of the Y genomic region could be postulated. MDPI 2020-12-30 /pmc/articles/PMC7823651/ /pubmed/33396671 http://dx.doi.org/10.3390/plants10010062 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article De Mori, Gloria Zaina, Giusi Franco-Orozco, Barbara Testolin, Raffaele De Paoli, Emanuele Cipriani, Guido Targeted Mutagenesis of the Female-Suppressor SyGI Gene in Tetraploid Kiwifruit by CRISPR/CAS9 |
title | Targeted Mutagenesis of the Female-Suppressor SyGI Gene in Tetraploid Kiwifruit by CRISPR/CAS9 |
title_full | Targeted Mutagenesis of the Female-Suppressor SyGI Gene in Tetraploid Kiwifruit by CRISPR/CAS9 |
title_fullStr | Targeted Mutagenesis of the Female-Suppressor SyGI Gene in Tetraploid Kiwifruit by CRISPR/CAS9 |
title_full_unstemmed | Targeted Mutagenesis of the Female-Suppressor SyGI Gene in Tetraploid Kiwifruit by CRISPR/CAS9 |
title_short | Targeted Mutagenesis of the Female-Suppressor SyGI Gene in Tetraploid Kiwifruit by CRISPR/CAS9 |
title_sort | targeted mutagenesis of the female-suppressor sygi gene in tetraploid kiwifruit by crispr/cas9 |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7823651/ https://www.ncbi.nlm.nih.gov/pubmed/33396671 http://dx.doi.org/10.3390/plants10010062 |
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