An efficient and simple plant regeneration system via organogenesis from leaf segments of persimmon (Diospyros kaki Thunb.) cultivars`Fuyu' and`Nishimurawase' has been developed. The regeneration capacity was influenced by the culture vessels, gelling agents, plant growth regulators, and light conditions. Leaf explants taken from in vitro shoots were cultured on a modified Murashige and Skoog medium (MS 1 2 N) for 16 wk without transfer to fresh medium. Adventitious shoots appeared after 4 and 8 wk in culture of`Nishimurawase' and`Fuyu' tissues, respectively. The culture of leaf explants in Erlenmeyer flasks with medium containing 4 g l 21 agar enhanced shoot formation in comparison to media with increased agar concentrations. Optimal shoot regeneration was obtained with 5 mg l 21 (22.8 mM) zeatin and 0.1 mg l 21 (0.05 mM) indole-3-butyric acid (IBA) for`Nishimurawase', and 10 mg l 21 (45.6 mM) zeatin and 0.1 mg l 21 (0.05 mM) IBA for`Fuyu'. Shoot regeneration frequencies in both cultivars were 100%, and shoot numbers per explant reached up to 9.2 for`Nishimurawase' and 2.2 for`Fuyu'. Dark incubation during the first 4±5 wk was the most effective condition to successfully influence shoot regeneration in both cultivars. While dark incubation was essential for adventitious shoot formation by`Fuyu', it was only slightly beneficial to`Nishimurawase'. More than 80% of the regenerated shoots rooted within 4 wk on hormone-free MS 1 2 N medium after having been dipped for 30 s in 250 mg l 21 (1.1 mM) IBA solution.