[Background] Stropharia rugosoannulata and Hypsizigus marmoreus are rare edible and medicinal fungi with great development potential. The medium plays an important role in mycelial growth and fruit body development. Optimizing the medium of the two mushroom is particularly important. [Objective] Screen out the new medium more suitable for S. rugosoannulata and H. marmoreus cultivation. [Methods] Using new media containing trehalose for the cultivation of S. rugosoannulata and H. marmoreus. The mycelial growth status, growth rate and biomass, cellulase and laccase activities were measured and analyzed in different culture media. [Results] Trehalose-added media promoted the growth rate of mycelia and increased the biomass compared with PDA medium. The proportion of trehalose content had a great effect on cellulase and laccase activity. Trehalose-added media could significantly promote the growth of hyphae, but the protein composition did not change. [Conclusion] PTA-5 was more suitable for the cultivation of S. rugosoannulata mycelia; PDTA was more suitable for the cultivation of H. marmoreus mycelia.