Agrobacterium-mediated transformation of recalcitrant hexaploid wheat cultivars using morphogenic regulators and/or expressing effector AvrPto with the type III secretion system

Wheat (Triticum aestivum L.) is a key crop for the global food supply. Its complex polyploid genome and recalcitrance to tissue culture pose challenges for genetic transformation, especially for elite cultivars. This study aimed to improve transformation efficiency in elite Korean wheat cultivars by integrating morphogenic regulator genes (GRF4–GIF1), using an engineered Agrobacterium tumefaciens strain. By integrating speed breeding to accelerate donor plant growth, we also reduced the time required to generate transgenic plants. Modifying the resting period and incorporating GRF4–GIF1 significantly improved the outcomes, leading to a transformation efficiency of 4.46% ± 0.93% in Bobwhite cultivar. Application of these methods to Korean elite cultivars also successfully yielded transformations with Keumkang and Baekkang cultivars. Furthermore, using a T3SS AGL-1 strain expressing the type III effector AvrPto enhanced the transformation efficiency in Baekkang cultivar by 2.27 times (17.7%) compared to the standard AGL-1 strain. Using the protocol as described here, we successfully developed stable transgenic lines of the Korean wheat cultivar, marking a pioneering achievement in Korean wheat molecular breeding. The modified protocol can contribute to wheat genetic research and breeding programs in Korea.