Tendrilless powdery mildew-resistant cucumber for vertical production systems

Overview

Cucumber production in controlled environment agriculture (CEA) and vertical farming systems faces two major challenges: the daily labor required to remove tendrils and the persistent threat of powdery mildew, the most damaging phytopathogen in indoor-grown cucumbers. This project applies CRISPR/Cpf1-mediated multi-gene editing to stack loss-of-function mutations in tendril-development genes (CsGCN5 and TEN) with mutations in powdery mildew susceptibility genes (CsaMLO8, CsaMLO1, and CsaMLO11) in elite cucumber genotypes. The result is a tendrilless cucumber line with pre- and post-invasion resistance to powdery mildew, purpose-built for vertical production environments where labor efficiency and disease management are critical.

Technical specifications

Key features:

• Tendrilless trait achieved by targeting the CsGCN5 histone acetyltransferase and TEN TCP transcription factor genes, both validated to produce the tendrilless phenotype
• Broad-spectrum powdery mildew resistance through simultaneous editing of three susceptibility genes (CsaMLO8, CsaMLO1, CsaMLO11), conferring both pre- and post-invasion resistance
• Cpf1-mediated multi-gene editing system enabling efficient stacking of five gene edits in a single breeding cycle, replacing lengthy conventional breeding approaches
• Virus-mediated delivery system using a disarmed TYLCV-based construct to deliver Cpf1 and guide RNA expression cassettes, demonstrated in peanut and peach and applicable to cucumber
• Agrobacterium-mediated transfection and vacuum infiltration delivery methods for introducing editing constructs into cucumber plants
• NGS-based mutation screening for high-throughput identification of edited plants
• Greenhouse bioassays for evaluating powdery mildew resistance, with field validation at the Coastal Research and Education Center

Technology readiness level

The project is at TRL 3–4. The genome editing construct for the CsaMLO genes was previously developed under a USDA-FAS project and will be modified in year one to incorporate guide RNAs targeting GCN5 and TEN. The Cpf1 editing system and virus-mediated delivery have been validated in other crops (peanut, wheat, peach) by the same research group, and TYLCV-based delivery has been demonstrated to infect cucumber. In year two, edited plants will be screened via next-generation sequencing and tested for powdery mildew resistance in greenhouse trials, with subsequent evaluation at the Coastal Research and Education Center. All necessary facilities and resources are available. The team seeks funding to support key personnel, services, and supplies to advance the technology toward a commercially viable, CEA-optimized cucumber cultivar.