Cryopreservation of Centaurea ultreiae (Compositae) a critically endangered species from Galicia (Spain)

Ex situ conservation of endangered plants is an important aim in order to preserve biodiversity of flora in threatened ecosystems. Among the biotechnological techniques which can be used, cryopreservation is emerging as a preferred option in many instances. This study describes a cryopreservation technique developed for shoot tips of the endangered species Centaurea ultreiae (Compositae) using a vitrification procedure. Basal medium (BM) for preculture and loading phases consisted of 1/2 MS basal salts with modified vitamins (3 μM thiamine). For preculturing shoot tips, BM with five osmotic treatments were investigated: 0.3 M sucrose ± 20 μM ABA, 0.6 M glycerol ± 20 μM ABA and 0.25 M sucrose + 0.25 M glycerol + 10 μM ABA. A loading solution treatment (BM with 2 M glycerol and 0.4 M sucrose) was applied prior to exposure of shoot tips to PVS2 and found to be indispensable to obtaining successful post-LN recovery. Highest (95.5%) regrowth of LN immersed shoot tips was obtained following incubation on BM + 0.3 M sucrose + 20 μM ABA or 0.25 M sucrose + 0.25 M glycerol + 10 μM ABA, with loading treatment and PVS2 exposure for 20 minutes at 0ºC