Agrobacterium tumefaciens-Mediated Stable Transformation of Malus domestica (Apple) cv Fuji Raku Raku and Royal Gala

The domesticated Malus x domestica Borkh plays a significant role in global horticulture and includes the majority of commercially produced species. Malus domestica Borkh cultivation, estimated at 82.93 million tons annually, is essential due to its nutritional value and health benefits. Traditional breeding methods, albeit effective, are time-consuming, requiring 15 to 25 years for new cultivar development. Considering this, genetic engineering offers a promising alternative, facilitated by the sequencing of the apple genome in 2010. Transformation via Agrobacterium tumefaciens is the most prevalent method used elsewhere. However, a lack of a universal apple transformation protocol underscores the need for tailored approaches. Here, we present a detailed protocol for generating transgenic apple trees via in vitro transformation of leaf explants, focusing on cultivars Fuji Raku Raku and Royal Gala.