Overcoming Challenges for Shoot Tip Cryopreservation of Root and Tuber Crops

Abstract

Root and tuber crops (RTCs) are the second-most important carbohydrate commodity after cereals. Many species of the RTCs are vegetatively propagated, making their shoot tips the preferred material to be conserved for future uses. Shoot tip cryopreservation provides an important tool to support the long-term conservation of plant genetic resources. Over the past four decades, significant efforts have been undertaken to move shoot tip cryopreservation of RTCs from research projects to full-scale implementation in cryobanks. This comprehensive review focuses on the history of cryopreservation protocols developed in RTCs. The encapsulation and vitrification solution-based cryopreservation techniques followed by ultra-rapid freezing and thawing have been highly successful. Additionally, different strategies for improving the cryotolerance of shoot tips have been introduced to further increase post-cryopreservation recovery. Finally, the research conducted to explain the mechanism underlying cryoprotection and differential cryotolerance including the use of histological studies are highlighted.

Critical Role of Regrowth Conditions in Post-Cryopreservation of In Vitro Plant Germplasm

“…Aluminum foils and cryoplates are primarily used in droplet-vitrification and cryoplate methods, respectively, due to fast heat transfer between the specimens and cooling or rewarming environment [14,19]. Cryo-tubes are employed in programmed freezing and vitrification methods when the samples are cryopreserved in a CPA mixture, and to cryopreserved explants embedded in alginate beads (encapsulation methods) [18,24,30]. Preparation of donor (stock) plant material (Stage 1 in Figure 1) is vital to the success of cryopreservation.…”

“…Cryopreservation has been applied to diverse plant materials: seeds, pollen, dormant buds, shoot tips or axillary buds of in vitro plants, hairy or adventitious roots, embryonic axes, embryogenic and non-embryogenic cell cultures, protoplasts, microtubers, rhizome buds, etc. [ 1 , 12 , 13 , 14 , 15 , 16 , 17 , 18 ]. Establishing an in vitro culture system is often inevitable for the regeneration of cryopreserved propagules and the multiplication of plant material for study, conservation, and restoration purposes [ 14 , 19 ].…”

“…Preparation of donor (stock) plant material (Stage 1 in Figure 1 ) is vital to the success of cryopreservation. Multiple studies have investigated the effects of cold-hardening, donor plant age, and the selection of appropriate explants on the recovery after cryopreservation, as previously reviewed [ 11 , 13 , 18 , 19 , 24 ].…”

“…However, some materials require special culture conditions (altered medium composition, specific growth regulators) and changes to the surrounding physical environment (darkness, gradual change in medium osmotic potential, etc.) to accomplish the recovery process [ 18 , 24 ]. The recovery stage becomes critical when materials are cryopreserved through non-optimized protocols and therefore are weakened from multiple injuries.…”

Cinchona officinalis L. in vitro conservation by means of slow-growth storage and cryopreservation techniques

“…Maintaining a water content between 10-20% is crucial for subsequent recovery (Gonzalez-Arnao et al, 2008). Sucrose acts as an osmoprotector; its concentration enhances tolerance to severe dehydration conditions in tissues exposed to cold (Zamecnik et al, 2021;Kim and Popova, 2023;Zhang et al, 2023).…”