Factors affecting in vitro adventitious bud induction from excised embryos of Swiss stone pine (Pinus cembra L.)

Several culture and preculture factors affecting in vitro adventitious bud induction on excised embryos of Swiss stone pine (Pinus cembra) were investigated. Embryos gave rise to adventitious buds within 4 weeks of culture in a cytokinin-containing medium. Among the tested cytokinins, benzyladenine (BA) was the most effective, especially when 2.5 or 5 µsmallcap˜M were added to the von Arnold and Eriksson's (AE) medium. All the tested pre-induction treatments (4, 8, or 12 week seed chilling, embryo preculture in a hormone-free medium prior to culturing on bud induction medium) negligibly affected adventitious budding. Whole embryos were the most suitable explants for bud induction; poor regeneration was achieved from isolated hypocotyls and epicotyls. A 21-day exposure of the unchilled explants to BA induced 100% regenerating embryos, and more than 16 buds per explant. Fewer adventitious shoots were obtained with a 7-day treatment, but shoot growth was moderately accelerated. Bud development was achieved on a hormone-free AE medium (3 weeks), while shoot elongation was optimal on activated charcoal-containing (0.1%) media (4 weeks). Some 15% of elongated shoots produced adventitious roots, after they were pulsed in liquid 1-msmallcap˜M IBA for 5 hours, prior to being transferred to a hormone-free medium.