Abstract: Taking Morchella sextelata as the test material, the effects of mycelial characteristics and TTC-dehydrogenase activity were investigated by repeated subculture, in order to provide reference for the relationship between repeated subculture and strain degradation.The results showed that under different cultivation conditions, the growth rate of mycelial, biomass and TTC-dehydrogenase activity decreased with repeated subculture increased.In order to maintain high enzyme activity and mycelial activity, repeated subculture in five times, and that mycelial germination was fast, thick and radiation in front.In addition, TTC-dehydrogenase activity was strongly and significantly affected by different methods of strain culture, and the relationship between species characteristics and mode of production should be fully considered.