Genetic fidelity of cryopreserved embryogenic cultures of open-pollinated Abies cephalonica

The aim of the present study was to apply cryopreservation for the embryogenic cultures of open-pollinated Abies cephalonica, as well as to test the genetic fidelity of the cryopreserved cell lines using random amplified polymorphic DNAs, i.e. RAPD markers. Dimethylsulfoxide (DMSO) and two mixtures of polyethylene glycol, glucose and DMSO (PGD I and PGD II), were used as cryoprotectants and the cryoprotectant treatments were performed with and without storage in liquid nitrogen. All the tested cryopreservation methods, i.e. DMSO, PGD I or PDG II used as cryoprotectant combined with slow cooling and fast thawing, proved to be applicable for A. cephalonica SE cultures. The best results, measured as survival percentage and mean growth ratio, were achieved using the PGD I treatment. The RAPD assays performed revealed considerable genetic variation in the DMSO treated but non-frozen samples, i.e. 16.8% of the produced RAPD profiles showed intraclonal variation while background variation was seen in 1.7% of the control amplifications. The present results show that cryoprotectants may cause a risk for genetic fidelity of plant material and thus, monitoring it, is important. This is particularly true in the breeding of the long-living conifers, since the effects of occasional mutations or genetic rearrangements may not be readily observed in young plants but expressed substantially later on in mature trees. (C) 1999 Elsevier Science Ireland Ltd. All rights reserved.