Location: Floral and Nursery Plants Research2012 Annual Report
1a. Objectives (from AD-416):
Utilize genetic engineering to improve resistance of bulb crops to fungal diseases and plant parasitic nematodes. (a) Develop transformation technology for Easter lily; (b) test candidate genes to confer resistance to Fusarium in Gladiolus; and (c) determine whether dsRNA expressed in plants can target RNAi to control plant parasitic nematodes.
1b. Approach (from AD-416):
Transform a commercially important Gladiolus cultivar, Easter lilies, and Ornithogalum with genes for resistance to either cucumber mosaic virus or ornithogalum mosaic virus. Determine if the genetically engineered plants are resistant to these viruses. Complete experiments involving cucumber mosaic virus resistance in transgenic Gladiolus plants that contain either the CMV coat protein subgroup 1 or subgroup 2, CMV replicase, or single chain antibodies to the CMV coat protein. Optimize the transformation system for Easter lilies and Ornithogalum using either the gene gun or Agrobacterium. Develop a transformation system for roots of Easter lilies using Agrobacterium rhizogenes. Transgenic Gladiolus plants with D4E1 and cpo have been developed using the gene gun and are being grown for challenging with Fusarium in the greenhouse. Gladiolus will also be transformed with various chitinase genes for possible resistance to Fusarium. Evaluate effectiveness of genetically engineered resistance to Fusarium oxysporum in Gladiolus by comparison to biological control technologies. Develop Easter lilies that are resistant to the nematode Pratylenchus penetrans, the root lesion nematode. Genes involved in nematode development will be isolated and tested for their ability to kill Pratylenchus penetrans for affect its reproduction using RNAi.
3. Progress Report:
Transgenic lilies with the gus gene under control of either the rolD or mas2 promoters have been developed. Transgenic Gladiolus lines containing the synthetic antimicrobial peptide D4E1 were challenged with Fusarium oxysporum in vitro, and two lines were selected for further study based upon having the most resistance in vitro. Conditions have been optimized for transformation of lilies using the gene gun. Direct bombardment of bulb scales resulted in a 6% transformation frequency. Conditions for Agrobacterium-mediated transformation were optimized based upon transient GUS expression, but stable transformants were not recovered using the optimized conditions.