Cryopreservation of woody-plant, dormant buds may provide cost-effective, long-term, back-up conservation of germplasm for vegetatively propagated crops that are presently maintained as trees in field gene banks. Dormant buds can be recovered quickly by grafting to dwarfing rootstocks, thus producing flowers for breeding purposes, with minimum potential for inducing somaclonal variants. These attributes are essential to preserve the clonal integrity of unique gene combinations such as those found in tree fruit crops. Previous research has shown that dormant buds from cold-hardy apples can be recovered from storage in liquid nitrogen (LN) with high survival rates (80% to 100%) using controlled desiccation and slow freezing before immersing in LN. On the other hand, dormant buds from cold-tender taxa and buds collected at less than optimal stages for desiccation and freezing have much lower (0% to 50%) survival rates. We increased survival of cold-tender taxa by using a modified vitrification procedure. Dormant apple buds from tender and hardy cultivars were perfused with modified PVS [15% (w/v) ethylene glycol, 15% (w/v) propylene glycol, 7% (w/v) DMSO, and 15% (w/v) glycerol in 0.5 m sorbitol]. Toxicity from the PVS was reduced by dilution soaking in 1 m sorbitol, 0.2 m raffinose, and 15 mm CaCl2 before and after quench-freezing and slow-freezing cryopreservation. Up to 100% of some cold-tender taxa survived. In addition, xylem ray parenchyma tissues that supercool and are normally killed at about -40C with the desiccation protocol survived this vitrification procedure.
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