Cryopreservation of embryogenic cultures from mature Quercus suber trees using vitrification

Recent progress in somatic embryogenesis from selected mature trees of Quercus suber, has led to a demand for maintenance of a large number of selected embryogenic lines. To facilitate the management of this material a protocol for the long-term storage of this germplasm should be defined. This study reports on the use of a simple vitrification procedure for the successful cryopreservation of three cork oak embryogenic lines. High embryo recovery levels (88-93 percent) were obtained by first preculturing 2-4 mg clumps of two or three globular embryos on semisolid medium containing 0.3 M sucrose for three days, followed by incubation in PVS2 vitrification solution at 0 degree C for 60 min before direct immersion in liquid nitrogen. The mean number of embryos produced per explant was significantly greater for cryostored embryos than for untreated stock cultures, but the productivity of the latter was recovered in subsequent subcultures of the material produced by cryostored embryos. The germination and plant regeneration rates achieved by cultures derived from cryostored embryos, around 60 percent, were similar to those of non-cryopreserved stock cultures.