Development of an efficient regeneration protocol for four Cyclamen species endemic to Turkey

In this study, embryo-like structures (ELSs) were induced in four endemic Turkish Cyclamen species (C. cilicium Boiss. et Heldr., C. parviflorum Pobed., C. mirabile Hildebr. and C. pseudibericum Hildebr.) in the presence of 13 combinations of two plant growth regulators (PGRs) (2,4-dichlorophenoxyacetic acid and 6-(gamma,gamma-dimethylallylamino)purine) and four explant types (ovules, ovaries, leaves and petioles). The ratio of callus induction, different stages of ELS formation and the conversion of ELSs to plantlets were quantified. The most effective explant types for callus induction were leaves (56 % for C. cilicium and 59 % for C. parviflorum) and petioles (80 % for C. mirabile and 100 % for C. pseudibericum). Callus growth from the leaves and petioles of C. cilicium was 30 days earlier than that of C. mirabile and C. pseudibericum. In contrast, most callus formed from the petioles of C. pseudibericum (100 %) in medium with 2.5 mg l(-1) 2,4-D and 1 mg l(-1) 2iP. The highest number of ELSs was obtained succesfully from petioles (2.5 mg l(-1) 2,4-D and 1 mg l(-1) 2iP) and ovaries (2.5 mg l(-1) 2,4-D and 0.5 mg l(-1) 2iP) of C. pseudibericum, in 39 % and as 32 % of explants, respectively. The percentage conversion of ELSs to plantlets was 38, 31, 16 and 15 % for C. mirabile, C. cilicium, C. pseudibericum and C. parviflorum, respectively. The plantlets were successfully acclimatized in the greenhouse with 54, 70, 63 and 25 % of C. cilicium, C. mirabile, C. pseudibericum and C. parviflorum plantlets, respectively surviving after transfer to ex vitro conditions. This paper describes a unique, reliable and consistent protocol for the induction of ELSs from four endangered endemic Turkish Cyclamen species, opening up the possibility of preserving these valuable genetic resources in vitro and also other applied biotechnologies that rely on a stable embryogenic or callus-based protocol.