Orchids on Ice: Protecting Imperiled Floridian Orchids through Cryopreservation

Recent advances in cell and tissue culture transformed cryopreservation practices, as the only method of conserving plants’ cellular material viable for long periods of time, from freeze-induced to vitrification and now to encapsulation dehydration, every explant culture can be protected. The methods behind encapsulation dehydration are relatively economical, replacing expensive programmable freezers. This technique has been applied to a wide range of flora species, demonstrating the success, efficiency, and practicality of cryogenically preserving cell suspensions, somatic embryos and meristems. The focus of this study is to evaluate the viability of protocorm-like bodies (PLBs) of critically threatened orchid species (Oncidium ensatum and Bletia purpurea) native to Florida after exposure to long-term cryogenic storage. Optimal encapsulation matrix has been modified previously in the synthetic seed study where 4% sodium alginate with 75 µM of calcium chloride (CACl₂) and PLBs were encapsulated before applying the pre-culture treatments. Pre-culture treatments, using half strength liquid MS media was agitated over the artificial seeds supplemented with 0.5, 0.75, 1.00 and 1.25 moles, prepared by combining 29.4, 44.1, 58.8 and 73.5 g of sucrose, respectively for an interval of 3 days. Viability of these orchids’ germplasm were validated using tetrazolium chloride (TTC) assays in addition to post-growth stage development following two weeks. Establishing a new cryopreservation protocol for these diverse sub-tropical plants will greatly enhance their ecological survival, increase conservation efforts and supply restoration activities in O. ensatum and B. purpurea. In conclusion, the findings of this research can be utilized as an alternative ex-situ methodology with the potential to direct long-term cryogenic storage of critically imperiled Floridian orchid species.