An improved dehydration protocol for cryopreservation of Brassica napus somatic embryos

This study compared three predehydration methods for cryopreservation of so matic embryos of oilseed rape (Brassica napus L.). These methods were (1) f ast-drying in a laminar airflow cabinet at 25 degrees C and 60 to 70% RH, ( 2) slow-drying for two days in closed jars with constant relative humiditie s (85%, 75%, 64% and 51%), and (3) stepwise slow-drying over six days using four humidity conditions (97% --> 92% --> 85% --> 75%). Desiccation tolera nce of oilseed rape somatic embryos increased significantly after precultur e for 7 days in the presence of 10 mg/L abscisic acid (ABA). Under the fast -drying condition, the critical water content of desiccation tolerance (50% survival) for embryos decreased from 1.16 to 0.24 g/g dw after ABA precult ure for 7 days. However, fast-dried somatic embryos with water content rang ing front 2.51 to 0.16 g/g dw did not survive after freezing and storage in liquid nitrogen. Slow-drying improved the survival after cryopreservation. The highest survival rate (55 +/- 3 %) was observed with 75% RH. The best survival rate (81 +/- 3 %) was obtained, using the stepwise slow-drying pro cedure in which RH was progressively lowered from 97 to 75% RH over a perio d of six days. The data suggest that the drying rate during dehydration gre atly affects the survival of somatic embryos after cryopreservation.