BIOTECHNOLOGY OF PRESERVATION OF THE RARE WILD ORCHID OF UKRAINIAN FLORA Ophrys sphegodes subsp. mammosa (Desf.) Soó ex E. Nelson

Abstract

The aim of the work was to determine the optimal conditions for callusogenesis in (in vitro) cultures of ovules, ovaries and anthers from vanishing wild orchid Ophrys sphegodes subsp. mammosa (Desf.) Soó ex E. Nelson followed by using of the obtained callus tissue to develop micropropagation techniques and conservation ex situ. In this study we used biotechnological (cultivation of explants in vitro) and microscopic (making temporary preparations, light microscopy, cytomorphological callus tissue analysis) methods. Nutrient media with optimal concentrations of growth regulators were selected and calluses derived from the generative organs were obtained (we used for ovaries Murashige–Skoog medium that contains 2.5 mg/l 6 of BAP and 1.5 mg/l of 2,4-D; for ovules — nutrient medium Nitsch and Nitsch containing 2.0 mg/l of 6-benzyladenine and 2.5 mg/l of 2,4-D, for the explants from anther — nutrient medium Nitsch and Nitsch containing 3 mg/l 6 of 6- benzyladenine and 2.5 mg/l of IBA). Cytomorphological analysis reveals the presence of meristematic foci in calluses. It indicates the beginning of the secondary processes of differentiation in the callus tissue. The results may be used in further investigations to obtain O. sphegodes regenerated plants from callus.

Keywords

• Ophrys sphegodes
• in vitro culture
• callus
• phytohormones