Kenneth Donsky,* Benjamin Ramsell, Britt Eubanks, and Laurent Deluc
*Oregon State University, 1450 NW Division Street, Apartment 32, Corvallis, OR, 97330 (donskyk@oregonstate.edu)

Genome editing can aid breeding programs dedicated to delivering genetically improved material from crops that will not be able to adapt to rapidly evolving climate change. Using the microvine system, we developed a research program to produce high-throughput gene-edited collections of grapevine mutants for easy gene-to-trait interrogations. To achieve this goal, we pursued the following three primary objectives: i) improve plant regeneration from Agrobacterium tumefaciens-mediated transformation events in somatic embryogenic grapevine cells, ii) increase the editing rate using a specific variant of the CRISPR/Cas9 protein, and iii) evaluate the potential of a new nanomaterial, carbon nanodots (CDs), to deliver multiple single guide RNAs for multiplexed editing. The approach to achieve objective 1 involves using the conditional expression of the morphogenic and synthetic gene, miRGRF_GIF4, from Vitis vinifera. For objective 2, we will compare the editing rate of different versions of the CRISPR/Cas9 editing system. In objective 3, we propose to identify the ratio of CD:sgRNA for optimized delivery to embryogenic grapevine cells. Zinc-functionalized CDs have been selected for their ease of synthesis and zinc’s affinity for nucleic acids. If we achieve our goals, we will give the scientific community a new tool for generating KO mutant collections that could expand beyond the grapevine model.

Funding Support: USDA NIFA