Masters in Forestry North Carolina State University Raleigh, North Carolina
Body of Abstract: CRISPR-Cas mediated genome editing is a powerful technology that could address some of the limitations in conventional tree breeding, such as the specificity of target traits improvement and the long generation time of trees. Wood and its bioproducts are a key source of renewable materials for construction, biofuels, textiles, and other applications. The increasing demand for this sustainable material is driving the forest industry to improve yield and different tree properties in a sustainable and efficient way. One of the major components of wood and an important trait to modify in content and composition is lignin, due to its recalcitrance and cost of removal (water, chemicals, and energy) during bioprocessing. In previous studies, our group has identified gene targets for mediating the reduction and improvement in composition of lignin in Populus trichocarpa, the model tree species using transgene and transgene-free CRISPR approaches. In the present project, we established a novel protoplast isolation and purification method for Eucalyptus spp., and achieved single gene editing for 9 genes related to lignin biosynthesis (up to 9% editing efficiency), multiplex gene editing of up to 4 genes, and tested a new chimeric system for genome editing, achieving up to 17.7% editing efficiency in single gene targets. These results are comparable to those in well studied agriculture model species, and further optimization will be done to improve the single and multiplex gene editing of Eucalyptus spp. In the near future, we will regenerate plants from edited Eucalyptus protoplasts and validate their genotypes and phenotypes.