Location: Vegetable Research
Project Number: 6080-22000-032-022-S
Project Type: Non-Assistance Cooperative Agreement
Start Date: Sep 1, 2025
End Date: Aug 31, 2027
Objective:
1. To develop a field-based detection of tomato brown rugose fruit virus (ToBRFV) using recombinase polymerase amplification;
2. To develop a molecular marker and conduct functional characterization of candidate genes of Solanum pimpinellifolium identified through quantitative trait locus analysis with resistance to ToBRFV;
3. To evaluate vaccines against ToBRFV on tomato using the RNA interference technology; and
4. To develop and characterize genome-edited tomato plants with resistance to viruses or tolerance to whiteflies.
Approach:
- Developing an RPA method for field-based detection of ToBRFV in plant and seed samples. We have identified a conserved sequence region that will be used for primer and probe design. Once the RPA reaction is confirmed to work with fluorescent, we will further simplify the detection method using a lateral flow test (LFT), which could be used for field application on an inexpensive heating block.
- Functional characterization of candidate resistance genes to ToBRFV in Solanum pimpinellifolium. Through quantitative trait locus analysis, we have identified a short genomic region on chromosome 11 with 17 genes that are associated with resistance to ToBRFV. Using virus-induced gene-silencing strategy, the resistance property on S. pimpinellifolium could be silenced. If the silencing would result in making the resistant S. pimpinellifolium plant to be susceptible to ToBRFV, then a resistance gene is identified.
- Evaluating vaccines against ToBRFV on tomato plants using the RNA interference technology. To engineer infectious clones using pepino mosaic virus as a vector to express various parts of ToBRFV genome sequence, we will evaluate these recombinant infectious clones for their ability to protect the vaccinated tomato plants from ToBRFV infection.
Functional characterization of genome-edited tomato plants with resistance to viruses or tolerance to whiteflies. We have developed several lines of genome-edited tomato plants using candidate genes with resistance to viruses (tobamoviruses, potyviruses, begomoviruses and its vector whitefly). T2 plants have been generated, and molecular characterization will be conducted to identify any mutation to the target genes. Biological characterization will be performed to evaluate their properties for the target traits (virus resistance to tolerance to whitefly).
Expected Outcome:
1. Simple and field-deportable RPA detection method to ToBRFV will be developed and validated to be useful for plant and seed health applications.
2. Molecular markers will be developed which could be useful to accelerate the breeding process through marker-assisted selection.
3. The resistance gene to ToBRFV on S. pimpinellifolium will be identified and used for tomato breeding.
4. An effective RNAi vaccine useful for protecting tomato from ToBRFV infection will be developed.
5. Genome edited tomato plants with resistance to viruses or tolerance to whiteflies will be identified and characterized.
6. Six or more manuscripts will be written and published in refereed journals.