Abstract: This study explored the effects of spent mushroom substrate(SMS) that were added to the soil on the nursery soil physical and chemical properties and growth of Manchurian ash(Fraxinus mandshurica) seedlings, and provided a scientific basis for the popularization and utilization of SMS added to the soil during cultivation of seedlings. The experiment adopted a completely randomized block design with 3 size classes of SMS that were applied(3 mm, 5 mm, 8 mm) and six ratios of SMS that were applied. The ratio of SMS to soil volume was 0(CK, no added SMS treatment), 1∶10, 1∶8, 1∶ 5, 1∶3, and 1∶2, and there were 16 plots per complete block. The suitable ratio of SMS that was added to the soil(SMS∶soil=1∶10) can significantly increase the content of soil organic matter and available nitrogen. At this time, the root biomass of Manchurian ash seedlings increased significantly. The growth of seedling height and root biomass were significantly higher with respect to the CK by 125.91% and 187.9%, respectively. A large amount of SMS that was added to the soil(SMS∶soil=1∶2) significantly inhibited the effect on soil nutrients and the growth of Manchurian ash seedlings. At this time, compared with the CK, and the soil organic matter and available nitrogen were significantly reduced by 26.62% and 41.03%, respectively. The growth of seedling height and ground diameter significantly were reduced by 27.80% and 35.09%, respectively. The lower ratio of SMS added to the soil can improve the soil physical and chemical properties, and improve the growth of seedlings. The size classes of SMS had no significant impact on seedling growth, but the interaction of ratio and size promotes the growth of seedlings height and improves the quality of seedlings. The soil condition in this study indicated that when the ratio and size was 1∶10 and 8 mm, respectively, it significantly increased soil pH, soil available nitrogen content and soil organic matter content. Meanwhile, it promoted root growth of seedlings, accumulated more biomass, and improved seedling quality.