Agrobacterium-mediated transformation is widely used in plant genetic engineering. This method involves both Agrobacterium infection and plant regeneration. In this context, establishing an efficient plant regeneration system is a critical prerequisite for genetic engineering in plants. This study aims to identify highbush blueberry (Vaccinium corymbosum L.) cultivars suitable for genetic transformation. Furthermore, we seek to elucidate the molecular and genetic factors that determine genotype-dependent shoot regeneration capacity by utilizing the diverse genetic background in highbush blueberries. Additionally, this study explores cultivar-specific differences in Agrobacterium susceptibility, which remain unexplored in highbush blueberries. Regeneration from leaf explants of 15 highbush blueberry cultivars was investigated on media containing 1.0 mg/L TDZ or 1.0 mg/L TDZ and 0.5 mg/L NAA. There was considerable variation in callus formation and regeneration rates, the number of regenerated shoots, the time required for regeneration, indicating that regeneration in highbush blueberry is highly genotype-dependent. The regeneration rate was high (>75%) in ‘Blue Muffin’, ‘Legacy’, ‘Gulfcoast’ and ‘Georgiagem’. However, ‘Georgiagem’ required three additional weeks for shoot regeneration from the time of meristem formation compared to the other three high regeneration cultivars. All four northern highbush cultivars exhibited low regeneration rates (
