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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Autor*in:	Shujuan Li [verfasserIn] Cheng Zhen [verfasserIn] Wenjing Xu [verfasserIn] Chong Wang [verfasserIn] Yuxiang Cheng [verfasserIn]

Format:	E-Artikel
Sprache:	Englisch

Erschienen:	2017

Übergeordnetes Werk:	In: Scientific Reports - Nature Portfolio, 2011, 7(2017), 1, Seite 10
Übergeordnetes Werk:	volume:7 ; year:2017 ; number:1 ; pages:10

Links:	Link aufrufen Link aufrufen Link aufrufen Journal toc

DOI / URN:	10.1038/s41598-017-02651-x

Katalog-ID:	DOAJ067873944

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520			\|a 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.
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allfields	10.1038/s41598-017-02651-x doi (DE-627)DOAJ067873944 (DE-599)DOAJ00ac3fbe7a2043be81d4dae16fe9630f DE-627 ger DE-627 rakwb eng Shujuan Li verfasserin aut Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree 2017 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier 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. Medicine R Science Q Cheng Zhen verfasserin aut Wenjing Xu verfasserin aut Chong Wang verfasserin aut Yuxiang Cheng verfasserin aut In Scientific Reports Nature Portfolio, 2011 7(2017), 1, Seite 10 (DE-627)663366712 (DE-600)2615211-3 20452322 nnns volume:7 year:2017 number:1 pages:10 https://doi.org/10.1038/s41598-017-02651-x kostenfrei https://doaj.org/article/00ac3fbe7a2043be81d4dae16fe9630f kostenfrei https://doi.org/10.1038/s41598-017-02651-x kostenfrei https://doaj.org/toc/2045-2322 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_171 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_370 GBV_ILN_381 GBV_ILN_602 GBV_ILN_2005 GBV_ILN_2009 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2055 GBV_ILN_2111 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4335 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 7 2017 1 10
spelling	10.1038/s41598-017-02651-x doi (DE-627)DOAJ067873944 (DE-599)DOAJ00ac3fbe7a2043be81d4dae16fe9630f DE-627 ger DE-627 rakwb eng Shujuan Li verfasserin aut Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree 2017 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier 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. Medicine R Science Q Cheng Zhen verfasserin aut Wenjing Xu verfasserin aut Chong Wang verfasserin aut Yuxiang Cheng verfasserin aut In Scientific Reports Nature Portfolio, 2011 7(2017), 1, Seite 10 (DE-627)663366712 (DE-600)2615211-3 20452322 nnns volume:7 year:2017 number:1 pages:10 https://doi.org/10.1038/s41598-017-02651-x kostenfrei https://doaj.org/article/00ac3fbe7a2043be81d4dae16fe9630f kostenfrei https://doi.org/10.1038/s41598-017-02651-x kostenfrei https://doaj.org/toc/2045-2322 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_171 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_370 GBV_ILN_381 GBV_ILN_602 GBV_ILN_2005 GBV_ILN_2009 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2055 GBV_ILN_2111 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4335 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 7 2017 1 10
allfields_unstemmed	10.1038/s41598-017-02651-x doi (DE-627)DOAJ067873944 (DE-599)DOAJ00ac3fbe7a2043be81d4dae16fe9630f DE-627 ger DE-627 rakwb eng Shujuan Li verfasserin aut Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree 2017 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier 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. Medicine R Science Q Cheng Zhen verfasserin aut Wenjing Xu verfasserin aut Chong Wang verfasserin aut Yuxiang Cheng verfasserin aut In Scientific Reports Nature Portfolio, 2011 7(2017), 1, Seite 10 (DE-627)663366712 (DE-600)2615211-3 20452322 nnns volume:7 year:2017 number:1 pages:10 https://doi.org/10.1038/s41598-017-02651-x kostenfrei https://doaj.org/article/00ac3fbe7a2043be81d4dae16fe9630f kostenfrei https://doi.org/10.1038/s41598-017-02651-x kostenfrei https://doaj.org/toc/2045-2322 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_171 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_370 GBV_ILN_381 GBV_ILN_602 GBV_ILN_2005 GBV_ILN_2009 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2055 GBV_ILN_2111 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4335 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 7 2017 1 10
allfieldsGer	10.1038/s41598-017-02651-x doi (DE-627)DOAJ067873944 (DE-599)DOAJ00ac3fbe7a2043be81d4dae16fe9630f DE-627 ger DE-627 rakwb eng Shujuan Li verfasserin aut Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree 2017 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier 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. Medicine R Science Q Cheng Zhen verfasserin aut Wenjing Xu verfasserin aut Chong Wang verfasserin aut Yuxiang Cheng verfasserin aut In Scientific Reports Nature Portfolio, 2011 7(2017), 1, Seite 10 (DE-627)663366712 (DE-600)2615211-3 20452322 nnns volume:7 year:2017 number:1 pages:10 https://doi.org/10.1038/s41598-017-02651-x kostenfrei https://doaj.org/article/00ac3fbe7a2043be81d4dae16fe9630f kostenfrei https://doi.org/10.1038/s41598-017-02651-x kostenfrei https://doaj.org/toc/2045-2322 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_171 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_370 GBV_ILN_381 GBV_ILN_602 GBV_ILN_2005 GBV_ILN_2009 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2055 GBV_ILN_2111 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4335 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 7 2017 1 10
allfieldsSound	10.1038/s41598-017-02651-x doi (DE-627)DOAJ067873944 (DE-599)DOAJ00ac3fbe7a2043be81d4dae16fe9630f DE-627 ger DE-627 rakwb eng Shujuan Li verfasserin aut Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree 2017 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier 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. Medicine R Science Q Cheng Zhen verfasserin aut Wenjing Xu verfasserin aut Chong Wang verfasserin aut Yuxiang Cheng verfasserin aut In Scientific Reports Nature Portfolio, 2011 7(2017), 1, Seite 10 (DE-627)663366712 (DE-600)2615211-3 20452322 nnns volume:7 year:2017 number:1 pages:10 https://doi.org/10.1038/s41598-017-02651-x kostenfrei https://doaj.org/article/00ac3fbe7a2043be81d4dae16fe9630f kostenfrei https://doi.org/10.1038/s41598-017-02651-x kostenfrei https://doaj.org/toc/2045-2322 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_171 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_370 GBV_ILN_381 GBV_ILN_602 GBV_ILN_2005 GBV_ILN_2009 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2055 GBV_ILN_2111 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4335 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 7 2017 1 10
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author	Shujuan Li
spellingShingle	Shujuan Li misc Medicine misc R misc Science misc Q Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree
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topic_title	Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree
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title	Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree
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title_sort	simple, rapid and efficient transformation of genotype nisqually-1: a basic tool for the first sequenced model tree
title_auth	Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree
abstract	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.
abstractGer	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.
abstract_unstemmed	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.
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title_short	Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree
url	https://doi.org/10.1038/s41598-017-02651-x https://doaj.org/article/00ac3fbe7a2043be81d4dae16fe9630f https://doaj.org/toc/2045-2322
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up_date	2024-07-03T14:34:39.522Z
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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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