Adventitious shoot regeneration from in vitro juvenile explants of black alder (Alnus glutinosa [L.] Gaertn.)

We studied the in vitro regeneration ability of black alder (Alnus glutinosa L.) using juvenile explants and a modified woody plant medium (mWPM). In our first experiment (Exp. 1), mature seeds were cultured in the presence of different concentrations of thidiazuron (TDZ: 0, 1, 5 and 10 mu M) for 2 or 4 weeks under light conditions (16 h photoperiod). In the second experiment (Exp. 2), hypocotyl segments (Hyp), cotyledonary nodes (CN) and cotyledons (Cot) were excised from 2-week-old in vitro seedlings and cultured on mWPM supplemented with different concentrations of 6-(gamma,gamma-dimethylallylamino)-purine (2iP: 0, 5, 10, 25 and 50 mu M) and 6-benzylaminopurine (BA: 1, 2, 3, 4 and 5 mu M) for 2, 4 or 6 weeks in the dark. After the induction periods, the explants were transferred to the expression medium consisting of mWPM containing 1 mu M BA under light conditions. The regeneration frequencies and the number of buds/shoots per regenerated explant were determined after 10 weeks for Exp. 1 and 8 weeks for Exp. 2. For Exp. 1, using mature seeds, 1 mu M TDZ for 2 weeks under light conditions proved to be the best induction treatment, with 91% of explants producing an average of 11 buds/shoots per explant, including more than four long shoots (> 1 cm). In Exp. 2, there were low shoot regeneration frequencies, 3% for the Hyp and 10% for the Cot. In contrast, the CN was the most responsive explant, with 100% explants producing 12-14 buds/shoots after the most effective induction treatment, based on 1-3 mu M BA for 2 weeks in the dark.