UC Davis

Gene delivery into plant cells has been studied for multiple decades and Agrobacterium-mediated transformation is still the most commonly used method. Mostly, his requires in vitro regeneration of whole plants; however, in vitro regeneration is genotype dependent and multiple elite crops, such as cotton, pepper, and sunflower, remain recalcitrant to regeneration and transformation. The molecular mechanisms of in vitro regeneration have been thoroughly investigated in model species such as Arabidopsis, and ectopic expression of characterized molecular players has been shown to increase regeneration rates in other species. Lettuce (Lactuca sativa L.) follows the de novo organogenesis pathway and little effort has been invested in identifying molecular mechanisms important to this mode of regeneration in lettuce. Previously, I have reviewed the molecular determinants of in vitro regeneration in model species as well as their potential use to enhance regeneration rates in lettuce. Genetic analysis of regeneration using a recombinant inbred line (RIL) mapping population identified eight quantitative trait loci (QTLs) associated with de novo shoot organogenesis in lettuce. Multiple candidate genes with known functions in de novo organogenesis and somatic embryogenesis were at or near the peaks of each QTL. Furthermore, in vitro regeneration rates in multiple diverse lettuce genotypes were boosted by the introduction of a GROWTH REGULATING FACTOR (GRF) and GRF-INTERACTING FACTOR (GIF) gene fusion (GRF-GIF). Co-transformation of a GRF-GIF construct with a gene of interest construct increased recovery of transgenic plants harboring the gene of interest. The use of magnetic nanoparticles as a DNA delivery agent was explored. Results from this work will be used to validate candidate genes, increase regeneration rates, and develop genotype-independent regeneration for gene delivery in lettuce. This work can be extended to other recalcitrant crops, particularly in the Compositae, such as sunflower.