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Salt and osmotic stress can improve the editing efficiency of CRISPR/Cas9-mediated genome editing system in potato

CRISPR/Cas9-mediated genome editing technology has been widely used for the study of gene function in crops, but the differences between species have led to widely varying genome editing efficiencies. The present study utilized a potato hairy root genetic transformation system and incorporated a rap...

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Autores principales: Ye, Mingwang, Yao, Mengfan, Li, Canhui, Gong, Ming
Formato: Online Artículo Texto
Lenguaje:English
Publicado: PeerJ Inc. 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10399558/
https://www.ncbi.nlm.nih.gov/pubmed/37547711
http://dx.doi.org/10.7717/peerj.15771
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author Ye, Mingwang
Yao, Mengfan
Li, Canhui
Gong, Ming
author_facet Ye, Mingwang
Yao, Mengfan
Li, Canhui
Gong, Ming
author_sort Ye, Mingwang
collection PubMed
description CRISPR/Cas9-mediated genome editing technology has been widely used for the study of gene function in crops, but the differences between species have led to widely varying genome editing efficiencies. The present study utilized a potato hairy root genetic transformation system and incorporated a rapid assay with GFP as a screening marker. The results clearly demonstrated that salt and osmotic stress induced by NaCl (10 to 50 mM) and mannitol (50 to 200 mM) treatments significantly increased the positive rates of genetic transformation mediated by A. rhizogenes and the editing efficiency of the CRISPR/Cas9-mediated genome editing system in potato. However, it was observed that the regeneration of potato roots was partially inhibited as a result. The analysis of CRISPR/Cas9-mediated mutation types revealed that chimeras accounted for the largest proportion, ranging from 62.50% to 100%. Moreover, the application of salt and osmotic stress resulted in an increased probability of null mutations in potato. Notably, the highest rate of null mutations, reaching 37.5%, was observed at a NaCl concentration of 10 mM. Three potential off-target sites were sequenced and no off-targeting was found. In conclusion, the application of appropriate salt and osmotic stress significantly improved the editing efficiency of the CRISPR/Cas9-mediated genome editing system in potato, with no observed off-target effects. However, there was a trade-off as the regeneration of potato roots was partially inhibited. Overall, these findings present a new and convenient approach to enhance the genome editing efficiency of the CRISPR/Cas9-mediated gene editing system in potato.
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spelling pubmed-103995582023-08-04 Salt and osmotic stress can improve the editing efficiency of CRISPR/Cas9-mediated genome editing system in potato Ye, Mingwang Yao, Mengfan Li, Canhui Gong, Ming PeerJ Agricultural Science CRISPR/Cas9-mediated genome editing technology has been widely used for the study of gene function in crops, but the differences between species have led to widely varying genome editing efficiencies. The present study utilized a potato hairy root genetic transformation system and incorporated a rapid assay with GFP as a screening marker. The results clearly demonstrated that salt and osmotic stress induced by NaCl (10 to 50 mM) and mannitol (50 to 200 mM) treatments significantly increased the positive rates of genetic transformation mediated by A. rhizogenes and the editing efficiency of the CRISPR/Cas9-mediated genome editing system in potato. However, it was observed that the regeneration of potato roots was partially inhibited as a result. The analysis of CRISPR/Cas9-mediated mutation types revealed that chimeras accounted for the largest proportion, ranging from 62.50% to 100%. Moreover, the application of salt and osmotic stress resulted in an increased probability of null mutations in potato. Notably, the highest rate of null mutations, reaching 37.5%, was observed at a NaCl concentration of 10 mM. Three potential off-target sites were sequenced and no off-targeting was found. In conclusion, the application of appropriate salt and osmotic stress significantly improved the editing efficiency of the CRISPR/Cas9-mediated genome editing system in potato, with no observed off-target effects. However, there was a trade-off as the regeneration of potato roots was partially inhibited. Overall, these findings present a new and convenient approach to enhance the genome editing efficiency of the CRISPR/Cas9-mediated gene editing system in potato. PeerJ Inc. 2023-07-31 /pmc/articles/PMC10399558/ /pubmed/37547711 http://dx.doi.org/10.7717/peerj.15771 Text en © 2023 Ye et al. https://creativecommons.org/licenses/by/4.0/This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ) and either DOI or URL of the article must be cited.
spellingShingle Agricultural Science
Ye, Mingwang
Yao, Mengfan
Li, Canhui
Gong, Ming
Salt and osmotic stress can improve the editing efficiency of CRISPR/Cas9-mediated genome editing system in potato
title Salt and osmotic stress can improve the editing efficiency of CRISPR/Cas9-mediated genome editing system in potato
title_full Salt and osmotic stress can improve the editing efficiency of CRISPR/Cas9-mediated genome editing system in potato
title_fullStr Salt and osmotic stress can improve the editing efficiency of CRISPR/Cas9-mediated genome editing system in potato
title_full_unstemmed Salt and osmotic stress can improve the editing efficiency of CRISPR/Cas9-mediated genome editing system in potato
title_short Salt and osmotic stress can improve the editing efficiency of CRISPR/Cas9-mediated genome editing system in potato
title_sort salt and osmotic stress can improve the editing efficiency of crispr/cas9-mediated genome editing system in potato
topic Agricultural Science
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10399558/
https://www.ncbi.nlm.nih.gov/pubmed/37547711
http://dx.doi.org/10.7717/peerj.15771
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