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.