Optimization of surface sterilization treatments in two Birch (Betula sp.) species ‎

Birch (Betula sp.) is considered as extinction species and its natural regeneration in Iranian forests ‎is very slow. Difficulties in regeneration as well as natural seed germination of birch tree besides ‎high levels of seed contaminations have justified the application in‏ ‏vitro culture with respect to its ‎conservation and genetic improvement. Hence, the in vitro culture initiation using seed explants of ‎two birch species was attempted. The seeds of B. pendula‏ ‏‎(collected from Sangdeh altitudes, ‎Mazandaran) and B. litwinowii‏ ‏‎(from Siamarzkooh forests, Golestan region) were randomly ‎harvested from the ten adult trees and then subjected to 7 surface sterilization treatments in 3 ‎replications. The experiment was undertaken as completely randomized design in factorial ‎arrangement (two factors) and 15 replications per each treatment. Application of Difnoconazole‏ ‏fungicide (3% for 24 h) followed by warm water (55°C in 30 min) and HgCl2 (0.1% for‎‏ ‏‎10 min) ‎were recognized as the best treatment for seed sterilization as well as germination. With this ‎treatment, 95-100%‎‏ ‏seeds were found to be free from bacterial and fungal contamination. ‎Consequently, seeds of two species were cultured on MS and B5 media. The germination ‎percentage for B. litwinowii and‏ ‏B. pendula‎‏ ‏were recorded as 17.3% and 14%, respectively. In ‎conclusion, these in vitro raised plantlets may be used as a source explants for in vitro culture ‎initiation of birch species, whose seed collection is so difficult in such impassable sites. ‎