The influence of disinfection methods and liquid phase media on Paphiopedilum insigne seeds germination and media supplements on morphological features of protocorms in tissue culture

Abstract

Paphiopedilum is a very original orchid, which labellum resembles a slipper. It is cultivated as a cut flower or a pot plant. The aim of the presented work was to estimate the influence of a disinfection method and addition of a liquid media on germination of Paphiopedilum insigne seeds in vitro. The whole seed capsules were disinfected with the use of 0.5% of AgNO3 for 20 min, 0.1% of HgCl2 for 5 s, 1% NaOCl for 30 min or immersed in a 96% ethanol and burned in a direct flame. The disinfected seeds were placed on a 1/4 MS (Murashige and Skoog) solidified media on top of which a liquid phase containing 1/4 MS elements, GA3 in concentration of 400 mg·dm–3 or distilled sterile water were added. The media without the liquid phase was also tested. It was noted that all methods of disinfection used effectively reduced contaminations of Paphiopedilum insigne seeds. The highest germination rate was observed when capsules were direct flamed and the liquid phase was added on top of the solidified media. The influence of growth regulators (BA, TDZ, KIN and 2,4-D) and casein hydrolysate added to the media on the morphological features of the obtained plants was also tested. It was observed that the most regenerated protocorms of the best quality were obtained when the media was supplemented with 1 mg·dm–3 of BA + 2 mg·dm–3 of TDZ or 5 mg·dm–3 of KIN + 1 mg·dm–3 of BA.