Transgenic plants from shoot apical meristems of Vitis vinifera L. "Thompson Seedless" via Agrobacterium-mediated transformation.
Shoot apical meristem explants of Vitis vinifera "Thompson Seedless" were used for Agrobacterium-mediated genetic transformation. It was determined that the meristems had to be subjected to a dark growth phase then wounded to obtain transgenic plants. Morphological and histological studies illustrated the role of wounding to expose apical meristem cells for transformation. A bifunctional egfp/nptII fusion gene was used to select kanamycin resistant plants that expressed green fluorescent protein (GFP). Kanamycin at a concentration of 16 mg L(-1) in selection medium resulted in recovery of non-chimeric transgenic plants that uniformly expressed GFP, whereas 8 mg L(-1) kanamycin allowed non-transgenic and/or chimeric plants to develop. Polymerase chain reaction (PCR) and Southern blot analyses confirmed the presence of transgenes and their stable integration into the genome of regenerated plants. Up to 1% of shoot tips produced stable transgenic cultures within 6 weeks of treatment, resulting in a total of 18 independent lines.
Dutt
M
M
Citrus Research and Education Center, University of Florida/IFAS, Lake Alfred, FL 33850, USA.
Li
Z T
ZT
Dhekney
S A
SA
Gray
D J
DJ
D015139:Blotting, Southern; D004351:Drug Resistance / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D007612:Kanamycin / Q000494:pharmacology; D018519:Meristem / Q000187:drug effects / Q000235:genetics* / Q000648:ultrastructure; D008855:Microscopy, Electron, Scanning; D018520:Plant Shoots / Q000187:drug effects / Q000235:genetics* / Q000648:ultrastructure; D030821:Plants, Genetically Modified / Q000187:drug effects / Q000235:genetics* / Q000648:ultrastructure; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012231:Rhizobium / Q000235:genetics; D046509:Tissue Culture Techniques; D014170:Transformation, Genetic; D019076:Transgenes / Q000235:genetics; D027843:Vitis / Q000187:drug effects / Q000235:genetics* / Q000648:ultrastructure
D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D007612:Kanamycin
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