Development of a system for direct transfer of antifungal candidate genes into European chestnut (Castanea sativa) would provide an alternative approach to conventional breeding for production of chestnut trees that are tolerant to ink disease caused by Phytophthora spp. Overexpression of genes encoding PR proteins (such as thaumatin-like proteins), which display antifungal activity, may represent an important advance in control of the disease. We have used a chestnut thaumatin-like protein gene (CsTL1) isolated from European chestnut cotyledons and have achieved overexpression of the gene in chestnut somatic embryogenic lines used as target material. We have also acclimatized the transgenic plants and grown them on in the greenhouse. Here, we describe the various steps of the process, from the induction of somatic embryogenesis to the production of transgenic plants.