Search

Cryopreservation (also called as cryobanking) of gel-embedded shoot tips on the cryo-plate dehydrated by vitrification solution (V cryo-plate method) was examined for saving plant resources of hybrid aspen. Cold-hardened shoot tips of in vitro grown hybrid aspen were precultured for 1 day at 25℃ on solidified 1/2 Murashige and Skoog (MS) medium containing 0.3 M sucrose. Pretreated shoot tips were placed in elliptical wells on an aluminium cryo-plate and embedded in calcium alginate gel. Osmoprotection treatment was performed in a loading solution containing 2.0 M glycerol and 1.0 M sucrose for 30 min at 25℃. After the shoot tip samples on the plates were treated with the plant vitrification solution 2 for 20 min at 25℃, the samples on the plates were directly plunged into liquid nitrogen. For rewarming, shoot tip samples on the cryo-plates were immersed in 1.0 M sucrose solution at 25℃. Then, shoot tips in the gels were regrown at 25℃ on solidified 1/2 MS medium. The regrowth rate of cryopreserved shoot tips was almost 100%, and normal shoot growth was observed without callus formation. This protocol appears to be a promising technique for cryobanking of hybrid aspen genetic resources. [ABSTRACT FROM AUTHOR]