To find out the reasonable ratio of rice seedling substrate prepared by Agaricus bisporus residue，which provides a scientific basis for the resource utilization of mushroom residue. Six kinds of rice seedling substrates were prepared according to different proportions using Agaricus bisporus residue，biochar，vermiculite and perlite as raw materials， and the effects of different raw material ratios on the physical and chemical properties of the substrates，the quality of rice seedlings and the number of microbial communities were explored. The results showed that with the increase of biochar content，the water withdrawal rate of the prepared substrate showed an increasing trend，and the ventilation porosity， water-holding porosity ratio and gas-water ratio of T2 treatment（biochar∶agaricus bisporus residue∶vermiculite∶perlite = 2.5∶8∶1∶1）were increased by 14.25%，5.34% and 6.45%，respectively，compared with that of CK treatment （commercially available peat-containing matrix）. With the increase of biochar content in the raw material ratio，the contents of organic matter and available potassium in the matrix showed an increasing trend，and the contents of organic matter and available potassium in T2 treatment were increased by 1.71% and 1.76%，respectively，compared with CK treatment，while the content of available phosphorus showed a decreasing trend. In terms of seedling quality，stem diameter， plant height and root length of T2 treatment increased by 13.65%，13.53% and 10.74%，respectively，compared with T6 treatment（biochar∶agaricus bisporus residue∶vermiculite∶perlite = 0.5∶8∶1∶1），but there was no significant difference between T2 and CK treatments，and the root vigour and root binding ability of T2 treatment increased by 36.30% and 96.89%，respectively，compared with T6 treatment. In addition，with the increase of biochar content，the content of bacteria and fungi in the substrate showed a decreasing trend，and the content of actinomycetes showed an increasing trend，compared with CK treatment，the content actinomycetes in T2 treatment increased by 7.12%，and the content of bacteria and fungi were decreased by 9.69% and 9.46%. On the whole，when the volume ratio of biochar∶agaricus bisporus residue∶vermiculite∶perlite was 2.5∶8∶1∶1，the prepared rice seedling raising substrate had the best effect.