In vitro conservation of two plant species (Prunus cerasifera Ehrh. and Rubus fruticosus L.) shoot tips by encapsulation dehydration

Abstract

In vitro grown shoot tips of cherry plum (Prunus cerasifera Ehrh.) and blackberry ?Cacanska Bestrna? (Rubus fruticosus L.) were tested for regrowth after cryopreservation using encapsulation dehydration method. Apical, 2-3 mm long shoot tips, were encapsulated in alginate beads composed of 3, 5 and 10% (w/v) alginic acid sodium salt in calcium-free liquid Murashige and Skoog (MS) medium containing 1.0 mg l-1 benzyladenine, 0.1 mg l-1 indole-3-butyric acid and 0.1 mg l-1 gibberellic acid. Polymerization was done in liquid MS medium with 100 mM CaCl2 for 30 min at room temperature. Encapsulated shoot tips were pre-treated in liquid MS medium with 0.75 or 1 M sucrose for 24 h in growth room and dehydrated for 4 and 8 h (29% and 20% moisture content respectively) before rapid immersion in liquid nitrogen. Upon thawing which involved placing the cryovials in the air current of the laminar airflow cabinet for 2 min, the beads were directly transferred to regrowth medium. In cherry plum, osmotic dehydration in 0.75 M sucrose followed by 8-hour desiccation gave the highest regrowth (60%) of explants encapsulated in 3% and 5% alginate beads. However, in comparison with cherry plum, blackberry displayed significantly lower capacity for regrowth after cryopreservation under described experimental conditions. In this genotype, osmotic dehydration in 1 M sucrose followed by 8-hour desiccation resulted in the highest regrowth (16.7%) of explants encapsulated in 5% alginate beads. Cryopreserved shoot tips of both genotypes multiplied in the three subcultures had normal morphology and similar multiplication capacity in comparison with non-cryopreserved shoots.