Genome Editing of Grape Cytidine Diphosphate Diacylglycerol Synthases to Confer Broad-Spectrum Disease Resistance. AVF project #2024-2873.

Summary of Progress:

Full-length cDNAs of two grapevine cytidine diphosphate diacylglycerol (CDP‐DAG) synthase (CDS) genes VvRBL1-1 (Vitvi10g00174) and VvRBL1-2 (Vitvi19g00570) were cloned from Thompson Seedless leaves. Both VvRBL1-1 and VvRBL1-2 were proposed as genome editing targets for enhancing broad-spectrum disease resistance in grapevine.

For rapid and rough testing of their impact on plant response to different pathogens, we transiently expressed VvRBL1-1/2 in Nicotiana benthamiana and then inoculated grapevine red blotch virus (GRBV), Botrytis cinerea (grey mold), or Pseudomonas syringae DC3000 (bacterial speck).

Transient expression of VvRBL1-2 seems to make plant susceptible to GRBV. Results of B. cinerea and DC3000 inoculations did not generate consistent results, which may be caused by the interference of Agrobacterium and/or high variation of transient VvRBL1-1/2 expression.

We generated stable VvRBL1-1/2 overexpression transformants using N. benthamiana and Arabidopsis, and currently in the process of selecting homozygous single-locus T-DNA insertional lines. Those transgenic lines would again be challenged by different pathogens, without the interference of Agrobacterium or the huge variation of transient gene expression.

I am currently in the process of setting up two research support agreements with UC Davis plant transformation facility to generate VvRBL1-1/2 editing plants and with Fresno State University to hire a part-time technician.