Increased efficiency using the encapsulation-dehydration cryopreservation technique for Arabidopsis thaliana

Arabidopsis thaliana shoot tips were successfully cryopreserved using encapsulation-dehydration cryopreservation methods. Between one and seven shoot tips were encapsulated within 4 mm calcium-alginate beads. Beads were formed in the presence of 2 M glycerol + 0.4 M sucrose. The time required to make 10 beads, each containing five shoot tips (4 min), was less than the time required to make 50 beads containing one shoot tip (12 min). Shoot tip regrowth after cryoexposure was between 60 and 68%, with one to seven shoot tips per bead. Using five Arabidopsis shoot tips per bead, alginate beads were formed either in the presence of 2 M glycerol + 0.4 M sucrose or 0.5 M sucrose. Beads formed in the presence of glycerol were immediately air-dried to moisture contents between 0.21 to 0.26 g H2O/g FW (0.27 to 0.38 g H2O/g DW). Alginate beads formed in 0.5 M sucrose were incubated in solutions of 0.5, 0.75, and 1 M sucrose for one day each prior to air-dehydration, achieving moisture contents of 0.19 to 0.21 g H2O/g FW (0.23 to 0.27 g H2O/g DW). Shoot tip regrowth after cryoexposure was between 42 and 65%, with no significant differences among treatments. We successfully reduced the amount of time needed for shoot tip processing for Arabidopsis by encapsulating five shoot tips per alginate bead and by using a glycerol-encapsulation method, without lowering shoot tip regrowth levels after cryopreservation.