Cryopreservation of embryogenic suspension cultures of Cyclamen persicum mill.
We have developed a simple protocol for the cryopreservation of embryogenic suspension cultures of Cyclamen persicum. Embryogenic suspension cultures in the linear growth phase 7-10 days after subculture were used for cryopreservation. Of the different cryoprotectants tested during a 2-day pre-culture, 0.6 M sucrose resulted in the highest re-growth rates of 75%. An additional pre-treatment with 0.6 M sucrose and 10% DMSO (dimethylsulfoxide) for 1 h also positively affected re-growth. Microscopic studies on viability revealed that only few small embryogenic cells survived cryopreservation, while vacuolated single cells died. Experiments in which the duration of the pre-culture period--i.e. the length of time the embryogenic suspension cells were exposed to 0.6 M sucrose--was varied showed that 2-4 days was the most optimal exposure time to 0.6 M sucrose. Callus re-grown after cryopreservation showed growth rates similar to that of unfrozen callus and regenerated even higher numbers of somatic embryos than unfrozen callus.
Winkelmann
Traud
T
Section of Floriculture, Institute of Floriculture, Tree Nursery Science and Plant Breeding, University of Hannover, Herrenhaeuser Str. 2, 30419, Hannover, Germany. traud.winkelmann@zier.uni-hannover.de
Mussmann
Viola
V
Serek
Margrethe
M
D018929:Cell Culture Techniques / Q000379:methods*; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured / Q000166:cytology* / Q000187:drug effects / Q000502:physiology; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents / Q000494:pharmacology*; D031824:Cyclamen / Q000187:drug effects / Q000196:embryology* / Q000254:growth & development*; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D013395:Sucrose / Q000494:pharmacology; D013997:Time Factors
D003451:Cryoprotective Agents; D013395:Sucrose; D004121:Dimethyl Sulfoxide