Abstract: In order to explore the effects of different cultivation substrate formulations on the activities of extracellular enzymes of Tremella fuciformis, the cottonseed shell and sawdust were used as the control, and the fungi such as fresh Lvzhou No.1 and fresh Pennisetum giganteum were used as the main raw materials for the industrial cultivation of Tremella fuciformis, thus to compare the activities of extracellular enzymes of Tremella fuciformis in different growth stages. The results showed that the activities of extracellular enzymes such as carboxymethyl cellulase, filter paper cellulase, amylase, laccase and xylanase under different cultivation substrates of Tremella fuciformis were as follows:cottonseed shell (C1) > fresh Lvzhou No.1 (L1) > fresh Pennisetum giganteum (J1) > sawdust (C2). The utilization ability of nutrient matrix in the industrial cultivation of Tremella fuciformis by using fresh fungi as the raw material was stronger than that of sawdust, and fresh fungi could replace sawdust as raw material for the industrial cultivation of Tremella fuciformis.