Establishing and Validating Secure Germplasm Conservation Protocols of Woody Plants and Banana by Cryopreservation Techniques

Plant cryopreservation is a key strategy for the long-term conservation of genetic resources, particularly for clonally propagated species and taxa that cannot be stored as orthodox seeds. Since its emergence in the mid-twentieth century, cryopreservation has evolved from slow-freezing approaches to advanced vitrification-based techniques that prevent intracellular ice formation and preserve cellular integrity at the temperature of liquid nitrogen (−196 °C). Understanding the physical and biochemical behavior of water, especially vitrification, is central to successful cryostorage. A wide range of plant tissues can be cryopreserved, including shoot tips, embryogenic calli, dormant buds, seeds, and pollen. Among these, shoot tips from in vitro culture and dormant winter buds are especially important for woody fruit crops. Modern methods, such as PVS-based vitrification, encapsulation-dehydration, droplet vitrification, dormant-bud, and cryo-plate techniques, have improved reproducibility, post-thaw regeneration, and large-scale applicability. Dormant-bud cryopreservation is particularly valuable for fruit trees because it allows direct field-to-cryobank conservation and back to field without tissue culture. Cryopreservation protocols must be carefully validated to ensure survival, regrowth capacity, and genetic and epigenetic stability. Post-thaw recovery conditions are critical for minimizing oxidative stress and promoting regeneration. European cryobanks and collaborative initiatives, including the EU COST Action “CopyTree,” play a major role in harmonizing protocols, training researchers, and securing woody plant germplasm for future breeding, research, and agricultural resilience.