Keywords: Artificial seeds, Slow growth storage, Storage conditions, Sucrose concentration, Synseeds
Phytophthora cinnamomi is a threat to chestnut fruit production. For improving resistance to this pathogen, a breeding program was implemented based on controlled crosses between Castanea species (Europe x Asia), using the latter as donor of resistant genes. The offspring with altered susceptibility to Phytophthora have been micropropagated. Plantlets were acclimatized to obtain copies of each genotype for phenotyping to Phytophthora susceptibility through inoculation tests. In this work, different methods of in vitro conservation, slow-growth storage and synthetic seed, during 9 and 12 months (at 4 ºC, darkness) were compared. For slow-growth storage, different types and concentrations of carbon sources (sucrose and mannitol at 0.09, 0.16, 0.22 M) were compared. Conversely, for synseeds, nodal segments were encapsulated using different concentrations of Na-alginate (2.75, 3%) and released in a solution of CaCl22H2O (50, 75, 100 mM). For conservation, three conditions were tested: empty tubes; tubes filled (15 mL) with sterile distilled water, and tubes with a 30% glycerol solution. Both methods tested were found suitable for medium-term conservation (12 months). For slow-growth storage, the best results of survival (93.3%) and multiplication rate (2.10, at 2nd subculture) were achieved with 0.22 M sucrose. To the best of our knowledge, our studies show for the first time that the storage of the synseeds in tubes with sterile distilled water was the most effective method for conservation, allowing higher survival (97.5%) and germination (92.5%). The use of the highest concentration of Na-alginate demonstrated to promote the multiplication rate (3.18, at 2nd subculture).
This work was supported by the project ProDeR, co-financed by the FEADER / European Union, Cooperation for Innovation, Ref. N. 53.590. We thank to Rosemeyre Cordeiro for the contribution to the manuscript.