Pre- and post-agroinfection strategies for efficient leaf disk transformation and regeneration of transgenic strawberry plants.
Following previously described Agrobacterium tumefaciens-mediated transformation procedures for Fragaria x ananassa Duch. 'Chandler', we undertook several experiments to establish the importance of some parameters affecting transformation. The most important factor that increased the percent recovery of transformants was the introduction of a pre-selection phase, in-between co-cultivation and selection, in which leaf disks were cultured on pre-selection regeneration medium containing validamycin A, timentin, and cefotaxime. The average percentage of leaf disks forming shoots on selection medium containing cefotaxime (250 mg l(-1)) + timentin (250 mg l(-1)) was 5.4% and about three shoots per regenerating leaf disk. Maximum transformation percentage, based on polymerase chain reaction, was 31.25%. Transgene integration and copy number were assessed by Southern hybridization confirming single copy as well as multiple copies of transgene integration in shoots as well as roots separately. This confirmed the non-chimeric nature of these transgenic plants. The system is very promising for the regeneration of genetically transformed cells and obtaining transgenic strawberry plants at high efficiency.
Husaini
Amjad Masood
AM
Department of Biotechnology, Faculty of Science, Jamia Hamdard, New Delhi, India. amjadhusaini@yahoo.com
D000098:Acetophenones / Q000494:pharmacology; D016960:Agrobacterium tumefaciens / Q000235:genetics; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D003470:Culture Media; D018744:DNA, Plant / Q000235:genetics; D031985:Fragaria / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D005818:Genetic Engineering / Q000379:methods*; D007612:Kanamycin / Q000494:pharmacology; D010671:Phenylurea Compounds / Q000494:pharmacology; D018515:Plant Leaves / Q000235:genetics* / Q000378:metabolism; D018520:Plant Shoots / Q000254:growth & development; D030821:Plants, Genetically Modified / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D012038:Regeneration; D013830:Thiadiazoles / Q000494:pharmacology; D046509:Tissue Culture Techniques / Q000379:methods*; D019076:Transgenes
D000098:Acetophenones; D000900:Anti-Bacterial Agents; D003470:Culture Media; D018744:DNA, Plant; D010671:Phenylurea Compounds; D013830:Thiadiazoles; C016785:thidiazuron; D007612:Kanamycin; C051667:acetosyringone