Dried Cryopreserved Somatic Embryos of Two Picea Species Provide Suitable Material for Direct Plantlet Regeneration and Germplasm Storage

Abstract

The present study aimed to develop a cryopreservation method for long-term storage of mature somatic embryos of Picea spp. The effects of drying rate, embryo water content prior to cryopreservation, thawing rate, and rehydration on survival of cryopreserved somatic embryos were investigated. Emphasis was placed on the capacity of cryopreserved somatic embryos to germinate and regenerate plantlets directly, or to reinduce embryogenic tissue for new embryo production. Firstly, a slow drying rate at 97 or 88% relative humidity (RH) was needed to achieve high germination (96.7–100%) and high plantlet conversion rates (26.7–46.7%) (not different from controls). Secondly, somatic embryos had to reach a water content of 0.23g H2Og−1d.wt (48h of desiccation at 97% RH) before immersion in liquid nitrogen to germinate at high frequency (93.8%). Thawing techniques had no effect on embryo survival. Dried and cryopreserved somatic embryos of Picea can also be used to reinduce embryogenic tissue and start new embryo production. Best reinduction frequency (66.7%) was obtained from cryopreserved embryos dried at 97% RH and rehydrated at 100% RH for 12h prior to reinduction. No difference in embryo production was noticed between the parent line (1st embryogenic cycle) and the sub-lines (2nd embryogenic cycle). Second generation embryos germinated and regenerated into plantlets at rates similar to controls. The optimal cryopreservation method was successfully applied to severalP. mariana and P. glauca genotypes.