Micropropagation and encapsulation-dehydration method for cryopreservation of Fritillaria imperialis

Fritillaria imperialis from the Liliaceae family is an ornamental species in danger of extinction. Therefore, it is nessessary to conserve the plant genetic pool. Two proper methods to access this goal are micropropagation and germplasm conservation in in vitro freezing conditions. The presence of suitable pre-treatments is necessary for cryopreservation. The most important and the most application of pre-treatment is encapsulation-dehydration. The purpose of current research was in vitro proliferation of F. imperialis using plant growth regulators and its cryopreservation using encapsulation-dehydration pre-treatment. 

Bulb scales as explants and MS medium as basic culture medium were used. For micropropagation, kinetin (Kin) and α_naphthaleneacetic acid in concentrations of 0 (as control), 0.5, 1 and 2 mg l–1 were applied. For cryopreservation, explants were pretreated with encapsulation-dehydration followed by conservation in liquid nitrogen. Encapsulation was done using sodium alginate. Dehydration was carried out in air and using high level of sucrose.

The results of micropropagation showed that the maximum number of leaf (3.5) and root number (5.7) was obtained in explants treated with 0.5 mg l–1 Kin along with 1 mg l–1 NAA. The results of cryopreservation showed that encapsulation-dehydration in air as a pre-treatment had effective role on the survival and regeneration of explants (70.5%) after conservation in liquid nitrogen. Least explant survival was obtained in control (without pre-treatment). In vitro regenerated plantlets were cultivated in plastic pots containing peat moss and perlite (in ration of 1:1) and acclimatized with environmental conditions. The survival rate was 90% and the growth pattern of regenerated plants was similar to that of mother plants.

Present research proposes the use of 0.5 mg l–1 Kin together with 1 mg l–1 NAA for micropropagation and encapsulation-dehydration for germplam cryopreservation of F. imperialis. Micropropagation by direct and indirect organogenesis and embryogenesis is a proper approach for proliferation of ornamentals in danger of extiction. The success of these methods depends on type and concentrations of auxins and cytokinins applied in culture medium, singular or in combination. In order to protect rare and endangered ornamental species, modern biotechnological tools need to be utilized.