Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree

Abstract Genotype Nisqually-1 is the first model woody plant with an available well-annotated genome. Nevertheless, a simple and rapid transformation of Nisqually-1 remains to be established. Here, we developed a novel shoot regeneration method for Nisqually-1 using leaf petiole and stem segment exp...

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Autores principales:	Shujuan Li, Cheng Zhen, Wenjing Xu, Chong Wang, Yuxiang Cheng
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2017
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/00ac3fbe7a2043be81d4dae16fe9630f
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spelling	oai:doaj.org-article:00ac3fbe7a2043be81d4dae16fe9630f2021-12-02T11:52:24ZSimple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree10.1038/s41598-017-02651-x2045-2322https://doaj.org/article/00ac3fbe7a2043be81d4dae16fe9630f2017-06-01T00:00:00Zhttps://doi.org/10.1038/s41598-017-02651-xhttps://doaj.org/toc/2045-2322Abstract Genotype Nisqually-1 is the first model woody plant with an available well-annotated genome. Nevertheless, a simple and rapid transformation of Nisqually-1 remains to be established. Here, we developed a novel shoot regeneration method for Nisqually-1 using leaf petiole and stem segment explants. Numerous shoots formed in the incision of explants within two weeks. The optimized shoot regeneration medium (SRM) contained 0.03 mg l−1 6-benzylaminopurine, 0.02 mg l−1 indole-3-butyric acid and 0.0008 mg l−1 thidiazuron. Based on this, Agrobacterium-mediated genetic transformation of stem explants was examined using the vector pBI121 that contains the β-glucuronidase (GUS) as a reporter gene. Consequently, factors affecting transformation frequency of GUS-positive shoots were optimized as follows: Agrobacteria cell suspension with an OD600 of 0.4, 20 min infection time, 2 days of co-cultivation duration and the addition of 80 µM acetosyringone into Agrobacteria infective suspension and co-cultivation SRM. Using this optimized method, transgenic plantlets of Nisqually-1 – with an average transformation frequency of 26.7% – were obtained with 2 months. Southern blot and GUS activity staining confirmed the integration of the foreign GUS gene into Nisqually-1. This novel transformation system for Nisqually-1 was rapid, efficient, and simple to operate and will improve more genetic applications in this model tree.Shujuan LiCheng ZhenWenjing XuChong WangYuxiang ChengNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 7, Iss 1, Pp 1-10 (2017)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Shujuan Li Cheng Zhen Wenjing Xu Chong Wang Yuxiang Cheng Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree
description	Abstract Genotype Nisqually-1 is the first model woody plant with an available well-annotated genome. Nevertheless, a simple and rapid transformation of Nisqually-1 remains to be established. Here, we developed a novel shoot regeneration method for Nisqually-1 using leaf petiole and stem segment explants. Numerous shoots formed in the incision of explants within two weeks. The optimized shoot regeneration medium (SRM) contained 0.03 mg l−1 6-benzylaminopurine, 0.02 mg l−1 indole-3-butyric acid and 0.0008 mg l−1 thidiazuron. Based on this, Agrobacterium-mediated genetic transformation of stem explants was examined using the vector pBI121 that contains the β-glucuronidase (GUS) as a reporter gene. Consequently, factors affecting transformation frequency of GUS-positive shoots were optimized as follows: Agrobacteria cell suspension with an OD600 of 0.4, 20 min infection time, 2 days of co-cultivation duration and the addition of 80 µM acetosyringone into Agrobacteria infective suspension and co-cultivation SRM. Using this optimized method, transgenic plantlets of Nisqually-1 – with an average transformation frequency of 26.7% – were obtained with 2 months. Southern blot and GUS activity staining confirmed the integration of the foreign GUS gene into Nisqually-1. This novel transformation system for Nisqually-1 was rapid, efficient, and simple to operate and will improve more genetic applications in this model tree.
format	article
author	Shujuan Li Cheng Zhen Wenjing Xu Chong Wang Yuxiang Cheng
author_facet	Shujuan Li Cheng Zhen Wenjing Xu Chong Wang Yuxiang Cheng
author_sort	Shujuan Li
title	Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree
title_short	Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree
title_full	Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree
title_fullStr	Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree
title_full_unstemmed	Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree
title_sort	simple, rapid and efficient transformation of genotype nisqually-1: a basic tool for the first sequenced model tree
publisher	Nature Portfolio
publishDate	2017
url	https://doaj.org/article/00ac3fbe7a2043be81d4dae16fe9630f
work_keys_str_mv	AT shujuanli simplerapidandefficienttransformationofgenotypenisqually1abasictoolforthefirstsequencedmodeltree AT chengzhen simplerapidandefficienttransformationofgenotypenisqually1abasictoolforthefirstsequencedmodeltree AT wenjingxu simplerapidandefficienttransformationofgenotypenisqually1abasictoolforthefirstsequencedmodeltree AT chongwang simplerapidandefficienttransformationofgenotypenisqually1abasictoolforthefirstsequencedmodeltree AT yuxiangcheng simplerapidandefficienttransformationofgenotypenisqually1abasictoolforthefirstsequencedmodeltree
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Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree

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