A chimera including aGROWTH-REGULATING FACTOR(GRF) and its cofactorGRF-INTERACTING FACTOR(GIF) increases transgenic plant regeneration efficiency

Genome editing allows precise DNA manipulation, but its potential is limited in many crops by low regeneration efficiencies and few transformable genotypes. Here, we show that expression of a chimeric protein including wheat GROWTH-REGULATING FACTOR 4 (GRF4) and its cofactor GRF-INTERACTING FACTOR 1 (GIF1) dramatically increases the efficiency and speed of regeneration in wheat, triticale and rice and expands the number of transformable wheat genotypes. Moreover, GRF4-GIF1 induces efficient wheat regeneration in the absence of exogenous cytokinins, which facilitates selection of transgenic plants without selectable markers. By combining GRF4-GIF1 and CRISPR-Cas9 technologies, we were able to generate large numbers of edited wheat plants. The GRF4-GIF1 transgenic plants were fertile and without obvious developmental defects, likely due to post-transcriptional regulatory mechanisms operating on GRF4 in adult tissues. Finally, we show that a dicot GRF-GIF chimera improves regeneration efficiency in citrus suggesting that this strategy can be expanded to dicot crops.