Cryopreservation of cold-acclimated shoot tips of pear in vitro cultures after encapsulation-dehydration

Cryopreservation of axillary shoot-tips of pear in vitro cultures ( Pyrus communis L. cv Beurré Hardy) was performed after encapsulation in alginate beads. Encapsulated shoot-tips were first precultured in medium enriched with sucrose and then dried in a sterile air flow and cooled in liquid nitrogen. After slow rewarming in air at room temperature, alginate beads were transferred to solid culture medium for 1 week before removal of shoot-tips from beads and subculture onto fresh medium. Shoot recovery from cryopreserved shoot-tips was greatly improved by 8–12 weeks of cold acclimation at 0 °C of donor in vitro cultures. The best results (80% shoot recovery) were obtained using 0.75 M sucrose for preculture and 4-h dehydration (giving 20% residual water). The resistance of encapsulated and dehydrated shoot-tips to liquid nitrogen did not depend on cooling rate. Apical shoot-tips about 3 mm in length with several axillary buds were also cryopreserved successfully (47% shoot recovery).