Optimization of Agrobacterium tumefaciens-Mediated Transformation of Xylaria grammica EL000614, an Endolichenic Fungus Producing Grammicin

An endolichenic fungus Xylaria grammica EL000614 produces grammicin, a potent nematicidal pyrone derivative that can serve as a new control option for root-knot nematodes. We optimized an Agrobacterium tumefaciens-mediated transformation (ATMT) protocol for X. grammica to support genetic studies. Tr...

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Autores principales:	Min-Hye Jeong, Jung A. Kim, Seogchan Kang, Eu Ddeum Choi, Youngmin Kim, Yerim Lee, Mi Jin Jeon, Nan Hee Yu, Ae Ran Park, Jin-Cheol Kim, Soonok Kim, Sook-Young Park
Formato:	article
Lenguaje:	EN
Publicado:	Taylor & Francis Group 2021
Materias:	atmt fungal transformation gene manipulation xylaria grammica Botany QK1-989
Acceso en línea:	https://doaj.org/article/45daa552bc4841db8c03f2a385e3b884
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spelling	oai:doaj.org-article:45daa552bc4841db8c03f2a385e3b8842021-11-11T14:23:41ZOptimization of Agrobacterium tumefaciens-Mediated Transformation of Xylaria grammica EL000614, an Endolichenic Fungus Producing Grammicin1229-80932092-932310.1080/12298093.2021.1961431https://doaj.org/article/45daa552bc4841db8c03f2a385e3b8842021-09-01T00:00:00Zhttp://dx.doi.org/10.1080/12298093.2021.1961431https://doaj.org/toc/1229-8093https://doaj.org/toc/2092-9323An endolichenic fungus Xylaria grammica EL000614 produces grammicin, a potent nematicidal pyrone derivative that can serve as a new control option for root-knot nematodes. We optimized an Agrobacterium tumefaciens-mediated transformation (ATMT) protocol for X. grammica to support genetic studies. Transformants were successfully generated after co-cultivation of homogenized young mycelia of X. grammica with A. tumefaciens strain AGL-1 carrying a binary vector that contains the bacterial hygromycin B phosphotransferase (hph) gene and the eGFP gene in T-DNA. The resulting transformants were mitotically stable, and PCR analysis showed the integratin of both genes in the genome of transformants. Expression of eGFP was confirmed via fluorescence microscopy. Southern analysis showed that 131 (78.9%) out of 166 transformants contained a single T-DNA insertion. Crucial factors for producing predominantly single T-DNA transformants include 48 h of co-cultivation, pre-treatment of A. tumefaciens cells with acetosyringone before co-cultivation, and using freshly prepared mycelia. The established ATMT protocol offers an efficient tool for random insertional mutagenesis and gene transfer in studying the biology and ecology of X. grammica.Min-Hye JeongJung A. KimSeogchan KangEu Ddeum ChoiYoungmin KimYerim LeeMi Jin JeonNan Hee YuAe Ran ParkJin-Cheol KimSoonok KimSook-Young ParkTaylor & Francis Grouparticleatmtfungal transformationgene manipulationxylaria grammicaBotanyQK1-989ENMycobiology, Vol 49, Iss 5, Pp 491-497 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	atmt fungal transformation gene manipulation xylaria grammica Botany QK1-989
spellingShingle	atmt fungal transformation gene manipulation xylaria grammica Botany QK1-989 Min-Hye Jeong Jung A. Kim Seogchan Kang Eu Ddeum Choi Youngmin Kim Yerim Lee Mi Jin Jeon Nan Hee Yu Ae Ran Park Jin-Cheol Kim Soonok Kim Sook-Young Park Optimization of Agrobacterium tumefaciens-Mediated Transformation of Xylaria grammica EL000614, an Endolichenic Fungus Producing Grammicin
description	An endolichenic fungus Xylaria grammica EL000614 produces grammicin, a potent nematicidal pyrone derivative that can serve as a new control option for root-knot nematodes. We optimized an Agrobacterium tumefaciens-mediated transformation (ATMT) protocol for X. grammica to support genetic studies. Transformants were successfully generated after co-cultivation of homogenized young mycelia of X. grammica with A. tumefaciens strain AGL-1 carrying a binary vector that contains the bacterial hygromycin B phosphotransferase (hph) gene and the eGFP gene in T-DNA. The resulting transformants were mitotically stable, and PCR analysis showed the integratin of both genes in the genome of transformants. Expression of eGFP was confirmed via fluorescence microscopy. Southern analysis showed that 131 (78.9%) out of 166 transformants contained a single T-DNA insertion. Crucial factors for producing predominantly single T-DNA transformants include 48 h of co-cultivation, pre-treatment of A. tumefaciens cells with acetosyringone before co-cultivation, and using freshly prepared mycelia. The established ATMT protocol offers an efficient tool for random insertional mutagenesis and gene transfer in studying the biology and ecology of X. grammica.
format	article
author	Min-Hye Jeong Jung A. Kim Seogchan Kang Eu Ddeum Choi Youngmin Kim Yerim Lee Mi Jin Jeon Nan Hee Yu Ae Ran Park Jin-Cheol Kim Soonok Kim Sook-Young Park
author_facet	Min-Hye Jeong Jung A. Kim Seogchan Kang Eu Ddeum Choi Youngmin Kim Yerim Lee Mi Jin Jeon Nan Hee Yu Ae Ran Park Jin-Cheol Kim Soonok Kim Sook-Young Park
author_sort	Min-Hye Jeong
title	Optimization of Agrobacterium tumefaciens-Mediated Transformation of Xylaria grammica EL000614, an Endolichenic Fungus Producing Grammicin
title_short	Optimization of Agrobacterium tumefaciens-Mediated Transformation of Xylaria grammica EL000614, an Endolichenic Fungus Producing Grammicin
title_full	Optimization of Agrobacterium tumefaciens-Mediated Transformation of Xylaria grammica EL000614, an Endolichenic Fungus Producing Grammicin
title_fullStr	Optimization of Agrobacterium tumefaciens-Mediated Transformation of Xylaria grammica EL000614, an Endolichenic Fungus Producing Grammicin
title_full_unstemmed	Optimization of Agrobacterium tumefaciens-Mediated Transformation of Xylaria grammica EL000614, an Endolichenic Fungus Producing Grammicin
title_sort	optimization of agrobacterium tumefaciens-mediated transformation of xylaria grammica el000614, an endolichenic fungus producing grammicin
publisher	Taylor & Francis Group
publishDate	2021
url	https://doaj.org/article/45daa552bc4841db8c03f2a385e3b884
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Optimization of Agrobacterium tumefaciens-Mediated Transformation of Xylaria grammica EL000614, an Endolichenic Fungus Producing Grammicin

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