Biolistic transformation of Carrizo citrange (Citrus sinensis Osb. × Poncirus trifoliata L. Raf.).
Identifieur interne : 000310 ( Main/Exploration ); précédent : 000309; suivant : 000311Biolistic transformation of Carrizo citrange (Citrus sinensis Osb. × Poncirus trifoliata L. Raf.).
Auteurs : Hao Wu [États-Unis] ; Yosvanis Acanda [États-Unis] ; Hongge Jia [États-Unis] ; Nian Wang [États-Unis] ; Janice Zale [États-Unis]Source :
- Plant cell reports [ 1432-203X ] ; 2016.
Abstract
The development of transgenic citrus plants by the biolistic method. A protocol for the biolistic transformation of epicotyl explants and transgenic shoot regeneration of immature citrange rootstock, cv. Carrizo (Citrus sinensis Osb. × Poncirus trifoliata L. Raf.) and plant regeneration is described. Immature epicotyl explants were bombarded with a vector containing the nptII selectable marker and the gfp reporter. The number of independent, stably transformed tissues/total number of explants, recorded by monitoring GFP fluorescence 4 weeks after bombardment was substantial at 18.4 %, and some fluorescing tissues regenerated into shoots. Fluorescing GFP, putative transgenic shoots were micro-grafted onto immature Carrizo rootstocks in vitro, confirmed by PCR amplification of nptII and gfp coding regions, followed by secondary grafting onto older rootstocks grown in soil. Southern blot analysis indicated that all the fluorescing shoots were transgenic. Multiple and single copies of nptII integrations were confirmed in five regenerated transgenic lines. There is potential to develop a higher throughput biolistics transformation system by optimizing the tissue culture medium to improve shoot regeneration and narrowing the window for plant sampling. This system will be appropriate for transformation with minimal cassettes.
DOI: 10.1007/s00299-016-2010-2
PubMed: 27277128
Affiliations:
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Raf.).</title><author><name sortKey="Wu, Hao" sort="Wu, Hao" uniqKey="Wu H" first="Hao" last="Wu">Hao Wu</name><affiliation wicri:level="1"><nlm:affiliation>Plant Pathology Department, Institute of Food and Agricultural Sciences, Citrus Research and Education Center, University of Florida, Lake Alfred, FL, 33850, USA. whao@ufl.edu.</nlm:affiliation><country xml:lang="fr">États-Unis</country><wicri:regionArea>Plant Pathology Department, Institute of Food and Agricultural Sciences, Citrus Research and Education Center, University of Florida, Lake Alfred, FL, 33850</wicri:regionArea><wicri:noRegion>33850</wicri:noRegion></affiliation></author><author><name sortKey="Acanda, Yosvanis" sort="Acanda, Yosvanis" uniqKey="Acanda Y" first="Yosvanis" last="Acanda">Yosvanis Acanda</name><affiliation wicri:level="1"><nlm:affiliation>Plant Pathology Department, Institute of Food and Agricultural Sciences, Citrus Research and Education Center, University of Florida, Lake Alfred, FL, 33850, USA.</nlm:affiliation><country xml:lang="fr">États-Unis</country><wicri:regionArea>Plant Pathology Department, Institute of Food and Agricultural Sciences, Citrus Research and Education Center, University of Florida, Lake Alfred, FL, 33850</wicri:regionArea><wicri:noRegion>33850</wicri:noRegion></affiliation></author><author><name sortKey="Jia, Hongge" sort="Jia, Hongge" uniqKey="Jia H" first="Hongge" last="Jia">Hongge Jia</name><affiliation wicri:level="1"><nlm:affiliation>Plant Pathology Department, Institute of Food and Agricultural Sciences, Citrus Research and Education Center, University of Florida, Lake Alfred, FL, 33850, USA.</nlm:affiliation><country xml:lang="fr">États-Unis</country><wicri:regionArea>Plant Pathology Department, Institute of Food and Agricultural Sciences, Citrus Research and Education Center, University of Florida, Lake Alfred, FL, 33850</wicri:regionArea><wicri:noRegion>33850</wicri:noRegion></affiliation></author><author><name sortKey="Wang, Nian" sort="Wang, Nian" uniqKey="Wang N" first="Nian" last="Wang">Nian Wang</name><affiliation wicri:level="1"><nlm:affiliation>Plant Pathology Department, Institute of Food and Agricultural Sciences, Citrus Research and Education Center, University of Florida, Lake Alfred, FL, 33850, USA.</nlm:affiliation><country xml:lang="fr">États-Unis</country><wicri:regionArea>Plant Pathology Department, Institute of Food and Agricultural Sciences, Citrus Research and Education Center, University of Florida, Lake Alfred, FL, 33850</wicri:regionArea><wicri:noRegion>33850</wicri:noRegion></affiliation></author><author><name sortKey="Zale, Janice" sort="Zale, Janice" uniqKey="Zale J" first="Janice" last="Zale">Janice Zale</name><affiliation wicri:level="1"><nlm:affiliation>Plant Pathology Department, Institute of Food and Agricultural Sciences, Citrus Research and Education Center, University of Florida, Lake Alfred, FL, 33850, USA. jzale@ufl.edu.</nlm:affiliation><country xml:lang="fr">États-Unis</country><wicri:regionArea>Plant Pathology Department, Institute of Food and Agricultural Sciences, Citrus Research and Education Center, University of Florida, Lake Alfred, FL, 33850</wicri:regionArea><wicri:noRegion>33850</wicri:noRegion></affiliation></author></analytic><series><title level="j">Plant cell reports</title><idno type="eISSN">1432-203X</idno><imprint><date when="2016" type="published">2016</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">The development of transgenic citrus plants by the biolistic method. A protocol for the biolistic transformation of epicotyl explants and transgenic shoot regeneration of immature citrange rootstock, cv. Carrizo (Citrus sinensis Osb. × Poncirus trifoliata L. Raf.) and plant regeneration is described. Immature epicotyl explants were bombarded with a vector containing the nptII selectable marker and the gfp reporter. The number of independent, stably transformed tissues/total number of explants, recorded by monitoring GFP fluorescence 4 weeks after bombardment was substantial at 18.4 %, and some fluorescing tissues regenerated into shoots. Fluorescing GFP, putative transgenic shoots were micro-grafted onto immature Carrizo rootstocks in vitro, confirmed by PCR amplification of nptII and gfp coding regions, followed by secondary grafting onto older rootstocks grown in soil. Southern blot analysis indicated that all the fluorescing shoots were transgenic. Multiple and single copies of nptII integrations were confirmed in five regenerated transgenic lines. There is potential to develop a higher throughput biolistics transformation system by optimizing the tissue culture medium to improve shoot regeneration and narrowing the window for plant sampling. 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