Carrot (Daucus carota L.).
Plants are susceptible to infection by a broad range of fungal pathogens. Many horticulturally important crop species lack adequate genetic resistance to disease. Studies on potential mechanisms of disease resistance in plants have revealed the importance of a range of pathogenesis-related (PR) proteins with antifungal activity in reducing colonization of plant tissues by pathogens. We are evaluating a range of PR-proteins, through heterologous expression in transgenic carrot tissues, for their effects on fungal disease development. The protocols for carrot transformation with a thaumatin-like protein are described. In addition, the use of herbicide resistance as a selectable marker in carrot transformation is illustrated. In this protocol, petiole segments from carrot seedlings are exposed to Agrobacterium for 10-30 min and co-cultivated for 3 d, after which herbicide selection is imposed until embryogenic calli are produced after 8-12 wk. The transfer of the embryogenic calli to hormone-free medium yields transgenic plantlets. This genetic transformation protocol has supported the generation of transgenic carrot plants with defined T-DNA inserts at the rate of between 1 and 3 Southern positive independent events out of 100.
Wally
Owen
O
Department of Biological Sciences, Simon Fraser University, Burnaby, British Columbia, Canada.
Jayaraman
Jayaraj
J
Punja
Zamir K
ZK
D016960:Agrobacterium tumefaciens / Q000166:cytology / Q000235:genetics*; D018929:Cell Culture Techniques; D003470:Culture Media; D018552:Daucus carota / Q000235:genetics* / Q000378:metabolism / Q000382:microbiology; D005658:Fungi / Q000502:physiology; D005822:Genetic Vectors; D007113:Immunity, Innate / Q000235:genetics; D010935:Plant Diseases / Q000382:microbiology; D010940:Plant Proteins / Q000235:genetics; D030821:Plants, Genetically Modified / Q000378:metabolism / Q000382:microbiology; D016133:Polymerase Chain Reaction; D040901:Proteomics; D036226:Seedlings / Q000235:genetics / Q000378:metabolism / Q000382:microbiology; D046509:Tissue Culture Techniques; D014170:Transformation, Genetic
D003470:Culture Media; D010940:Plant Proteins