Cryopreservation of Vanda coerulea Griff, ex Lindl. Seeds by vitrification

Abstract

An effective protocol for cryopreservation of seeds of Vanda coerulea Griff, ex Lindl., an endangered Thai epiphytic orchid, in liquid nitrogen (LN) was developed by vitrification. Seeds from 7-month-old pods were sufficiently dehydrated in 2 ml cryotubes filled with highly concentrated plant vitrification solution (PVS2), consisting of 30% (w/v) glycerol, 15% (w/v) ethylene glycol and 15% (w/v) dimethyl sulfoxide, prepared in modified Vacin and Went (1949) (VW) liquid medium at 25 ± 2°C for 70 min. The seeds were then rapidly plunged into LN. After recovering from LN and rapid warming in a waterbath at 40°C for 2 min, the PVS2 solution was replaced with 0.5 ml of 1.2 M sucrose in VW solution and kept at 25 ± 2°C for 20 min prior to transfer on VW agar medium. About 67% of cryopreserved seeds treated with PVS2 solution for 70 min were able to develop into normal seedlings in vitro, while without PVS2 treatment, there was no survival after LN storage.