Lessons From CABBI: Advancing Vegetative Oil Production in C₄ Crops Through a Vertically Integrated DBTL Framework

The global demand for renewable liquid fuels and biobased chemicals has intensified interest in engineering high-biomass C₄ grasses to accumulate triacylglycerol (TAG) in vegetative tissues. Achieving this goal requires coordinated solutions across molecular biology, crop science, chemical engineering, and life cycle assessment, disciplines that rarely operate within a single research program. The Center for Advanced Bioenergy and Bioproducts Innovation (CABBI) was established to provide this integration through a Design-Build-Test-Learn (DBTL) framework applied to three C₄ feedstocks: saccharum (Saccharum spp. hybrids), sorghum (Sorghum bicolor), and miscanthus (Miscanthus × giganteus). Through iterative DBTL cycles, CABBI has produced transgenic saccharum and sorghum lines with substantially elevated levels of vegetative TAG, with multi-location field trials having confirmed stable trait heritability and lipid yields. We have also leveraged our integration to develop a dual system biorefinery that co-recovers vegetative lipids with fermentable sugars at levels sufficient to achieve industrial ethanol titres. Importantly, this fully integrated cross-disciplinary data integration model has highlighted the key constraints currently preventing TAG-rich vegetative oil from achieving commercial viability as: (i) active lipid turnover, (ii) photosynthetic trade-offs, (iii) spatial misexpression, and (iv) bioprocessing challenges. CABBI uses emerging technologies, including biofoundry-enabled construct screening, precision genome editing, and machine-learning-assisted regulatory design, to further accelerate DBTL cycles. More broadly, this work illustrates how integrative centre-scale research that unites diverse expertise can accelerate discovery and translation, positioning these systems to expand agricultural markets and contribute to a more secure and resilient bioeconomy.