In vitro asymbiotic germination, seedling formation and tuberization of Orchis sancta

Plant tissue culture techniques have recently become attractive tools for propagating rare and endangered species and producing their economically important natural products. Here, we introduce a protocol for an effective, fast and reliable production route of plantlets and tubers from salep orchid, Orchis sancta, a native orchid species growing wild in Turkey. In the course of this study, the efficacy of basal media and plant growth regulators on asymbiotic germination of mature seed, seedling formation, tuberization and glucomannan contents of tubers were assessed. Amongst all basal media evaluated, Orchimax including activated charcoal gave the best germination rate (62.46%). Supplementing this medium with 1.0 mg/L zeatin resulted in the highest germination rate (81.18%). Lowest thidiazuron concentration (0.25 mg/L) was very effective on shoot elongation while kinetin (0.5 and 1.0 mg/L) promoted leaf formation. The aforesaid basal medium with IBA favored root extension (31.16 mm) and root formation (4.5 roots per plantlet) at 1.0 mg/L and 2.0 mg/L IBA, respectively. Cytokinins, zeatin (2.0 mg/L) in particular, promoted tuber formation and glucomannan accumulation. Plantlets were successfully adapted to soil step by step.