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An Improved Procedure for Agrobacterium-Mediated Transformation of ‘Carrizo’ Citrange

Although several protocols for genetic transformation of citrus have been published, it is highly desirable to further improve its efficiency. Here we report treatments of Agrobacterium cells and citrus explants prior to and during co-cultivation process to enhance transformation efficiency using a...

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Detalles Bibliográficos
Autores principales: Li, Yanjun, Tang, Dan, Liu, Zongrang, Chen, Jianjun, Cheng, Baoping, Kumar, Rahul, Yer, Huseyin, Li, Yi
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9183180/
https://www.ncbi.nlm.nih.gov/pubmed/35684233
http://dx.doi.org/10.3390/plants11111457
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author Li, Yanjun
Tang, Dan
Liu, Zongrang
Chen, Jianjun
Cheng, Baoping
Kumar, Rahul
Yer, Huseyin
Li, Yi
author_facet Li, Yanjun
Tang, Dan
Liu, Zongrang
Chen, Jianjun
Cheng, Baoping
Kumar, Rahul
Yer, Huseyin
Li, Yi
author_sort Li, Yanjun
collection PubMed
description Although several protocols for genetic transformation of citrus have been published, it is highly desirable to further improve its efficiency. Here we report treatments of Agrobacterium cells and citrus explants prior to and during co-cultivation process to enhance transformation efficiency using a commercially used rootstock ‘Carrizo’ citrange [Citrus sinensis (L.) Osb. × Poncirius trifoliata (L.) Raf.] as a model plant. We found explants from light-grown seedlings exhibited higher transformation efficiency than those from etiolated seedlings. We pre-cultured Agrobacterium cells in a 1/10 MS, 0.5 g/L 2-(N-morpholino) ethanesulfonic acid (MES) and 100 µM acetosyringone liquid medium for 6 h at 25 °C before used to infect citrus explants. We incubated epicotyl segments in an MS liquid medium containing 13.2 µM 6-BA, 4.5 µM 2,4-D, 0.5 µM NAA for 3 h at 25 °C prior to Agrobacterium infection. In the co-cultivation medium, we added 30 µM paclobutrazol and 10 µM lipoic acid. Each of these treatments significantly increased the efficiencies of transformation up to 30.4% (treating Agrobacterium with acetosyringone), 31.8% (treating explants with cytokinin and auxin), 34.9% (paclobutrazol) and 38.6% (lipoic acid), respectively. When the three treatments were combined, we observed that the transformation efficiency was enhanced from 11.5% to 52.3%. The improvement of genetic transformation efficiency mediated by these three simple treatments may facilitate more efficient applications of transgenic and gene editing technologies for functional characterization of citrus genes and for genetic improvement of citrus cultivars.
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spelling pubmed-91831802022-06-10 An Improved Procedure for Agrobacterium-Mediated Transformation of ‘Carrizo’ Citrange Li, Yanjun Tang, Dan Liu, Zongrang Chen, Jianjun Cheng, Baoping Kumar, Rahul Yer, Huseyin Li, Yi Plants (Basel) Article Although several protocols for genetic transformation of citrus have been published, it is highly desirable to further improve its efficiency. Here we report treatments of Agrobacterium cells and citrus explants prior to and during co-cultivation process to enhance transformation efficiency using a commercially used rootstock ‘Carrizo’ citrange [Citrus sinensis (L.) Osb. × Poncirius trifoliata (L.) Raf.] as a model plant. We found explants from light-grown seedlings exhibited higher transformation efficiency than those from etiolated seedlings. We pre-cultured Agrobacterium cells in a 1/10 MS, 0.5 g/L 2-(N-morpholino) ethanesulfonic acid (MES) and 100 µM acetosyringone liquid medium for 6 h at 25 °C before used to infect citrus explants. We incubated epicotyl segments in an MS liquid medium containing 13.2 µM 6-BA, 4.5 µM 2,4-D, 0.5 µM NAA for 3 h at 25 °C prior to Agrobacterium infection. In the co-cultivation medium, we added 30 µM paclobutrazol and 10 µM lipoic acid. Each of these treatments significantly increased the efficiencies of transformation up to 30.4% (treating Agrobacterium with acetosyringone), 31.8% (treating explants with cytokinin and auxin), 34.9% (paclobutrazol) and 38.6% (lipoic acid), respectively. When the three treatments were combined, we observed that the transformation efficiency was enhanced from 11.5% to 52.3%. The improvement of genetic transformation efficiency mediated by these three simple treatments may facilitate more efficient applications of transgenic and gene editing technologies for functional characterization of citrus genes and for genetic improvement of citrus cultivars. MDPI 2022-05-30 /pmc/articles/PMC9183180/ /pubmed/35684233 http://dx.doi.org/10.3390/plants11111457 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/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 (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Li, Yanjun
Tang, Dan
Liu, Zongrang
Chen, Jianjun
Cheng, Baoping
Kumar, Rahul
Yer, Huseyin
Li, Yi
An Improved Procedure for Agrobacterium-Mediated Transformation of ‘Carrizo’ Citrange
title An Improved Procedure for Agrobacterium-Mediated Transformation of ‘Carrizo’ Citrange
title_full An Improved Procedure for Agrobacterium-Mediated Transformation of ‘Carrizo’ Citrange
title_fullStr An Improved Procedure for Agrobacterium-Mediated Transformation of ‘Carrizo’ Citrange
title_full_unstemmed An Improved Procedure for Agrobacterium-Mediated Transformation of ‘Carrizo’ Citrange
title_short An Improved Procedure for Agrobacterium-Mediated Transformation of ‘Carrizo’ Citrange
title_sort improved procedure for agrobacterium-mediated transformation of ‘carrizo’ citrange
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9183180/
https://www.ncbi.nlm.nih.gov/pubmed/35684233
http://dx.doi.org/10.3390/plants11111457
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