Synergistic developmental regulators enable efficient plant regeneration and transformation across species

Plant genetic transformation is a powerful tool for crop improvement and functional genomics, yet many economically important species remain recalcitrant to current methodologies. To address this limitation, we screened nine developmental regulatory genes (DRs) and sixteen DR combinations, identifying the combination of PLETHORA5 (PLT5) and GROWTH-REGULATING FACTOR 4/GRF-INTERACTING FACTOR 1 (GRF4/GIF1) (labeled as PLT5-GRFs combo) as a potent enhancer of regeneration and transformation efficiency. PLT5-GRFs combination was effective across diverse plant species, including petunia (Petunia hybrida), tomato (Solanum lycopersicum), lettuce (Lactuca sativa), ornamental eggplant (Solanum aethiopicum), and recalcitrant species such as chili pepper (Capsicum annuum), begonia (Begonia semperflorens), bahiagrass (Paspalum notatum Flügge), and blueberry (Vaccinium corymbosum). Moreover, it enhanced in planta transformation in Gloxinia (Sinningia speciosa). Transcriptional profiling revealed that PLT5-GRFs synergistically regulates key pathways in development, hormone signaling, epigenetic modifications, and cell fate reprogramming. Our findings establish PLT5-GRFs as a universal tool for plant genetic modification, offering a broadly applicable platform in plant biotechnology with profound implications for basic research, crop improvement, and sustainable agriculture.