Micropropagation of endangered species Daphne cneorum

A new protocol for micropropagation of endangered Daphne cneorum through multiple shoot formation has been developed. Two different types of explants (dormant apical buds and in vitro seed-derived young seedlings) from plants in two different localities were used for the initiation of multiple shoots on agar woody plant medium (WPM) with 0.2 mg dm(-3) benzylaminopurine (BAP), 0.1 mg dm(-3) indolebutyric acid (IBA), 200 mg dm(-3) glutamine, and 200 mg dm(-3) casein hydrolysate. From 10 seeds only one germinated and the multi-apex culture bearing 12 shoots sprouted out from in vitro seed-derived young seedling. After 6-month cultivation 35 multi-apex cultures were achieved from in vitro seed-derived young seedling. On 1/3 strength WPM medium supplemented with 2.83 mg dm(-3) IBA 50% of cultures (clusters of 3-5 shoots) rooted but no rooting occurred in the presence of naphthaleneacetic acid (NAA). The rooted plantlets were acclimatized for 4 weeks in the greenhouse and then transferred into natural conditions. The plants successfully survived the winter and flowered.