Crispr-cas9 editing of TAD1 gene to improve yield and lodging tolerance in tef (eragrostis tef)

Overview

Tef (Eragrostis tef) is an important orphan crop valued for its nutritional profile and drought tolerance, yet it remains one of the lowest-yielding cereals at less than 1.5 tons per hectare. This project applies CRISPR-Cas9 gene editing to mutate the Tillering and Dwarfing1 (TAD1) gene in tef, with the goal of simultaneously increasing grain yield and lodging tolerance. The approach is grounded in validated rice research demonstrating that TAD1 regulates the MOC1 gene—a key determinant of tiller number—and that loss of TAD1 function produces a semidwarf phenotype with enhanced tillering. By translating this mechanism to tef, the project addresses two of the most significant agronomic constraints limiting tef productivity, offering a pathway to meaningful yield gains in a crop that has historically been underserved by breeding and biotechnology investment.

Technical specifications

• Target gene: TAD1 homolog identified in the tef genome, located on chromosomes 1 and 4 (allotetraploid species with two gene copies)
• Editing platform: CRISPR-Cas9 pipeline leveraging "polyvalent" gRNAs designed for simultaneous targeting of multiple gene sequences
• Transformation method: Agrobacterium-mediated (strain LBA-4404-Thy) delivery using a binary vector containing maize Babyboom and Wuschel2 genes to enhance regeneration efficiency
• Transgene excision: Bbm and Wus expression cassettes removed via osmotic stress treatment after callus proliferation to eliminate pleiotropic effects on plant growth and fertility
• Validation approach: Transgenic lines confirmed by whole-genome sequencing to verify absence of integrated transgenes
• Prior validation: Polyvalent gRNA approach previously demonstrated robust suppression of viral genes with at least 70% sequence identity in Nicotiana benthamiana
• Transformation infrastructure: Tef transformation platform under development using technology obtained from Corteva Bioscience

Technology readiness level

The project is at an early-to-mid stage of development. The TAD1 homolog has been identified in the tef genome, and the CRISPR-Cas9 editing pipeline has been validated in a related system (viral gene suppression in N. benthamiana). The tef-specific transformation platform is currently being established using technology licensed from Corteva Bioscience. Future validation steps include editing all TAD1 alleles across both chromosomal copies, generating transgenic lines through Agrobacterium-mediated transformation, excising regeneration cassettes, and confirming transgene-free status by genome sequencing. The work represents a translational application of proven gene-editing methodology to an orphan crop, with proof-of-concept editing expected once the tef transformation pipeline is fully operational.