The invention discloses a method for preparing vegetable cultivation substrates from needle mushroom fungus chaff, and belongs to the technical field of agricultural production. The specific method isas follows: using corn steep liquor, molasses, straw and urea to adjust the ratio of carbon to nitrogen of the needle mushroom fungus chaff to (22-28):1, adding a microbial bacterium agent to performstacking fermentation, and regulating aerobic fermentation and anaerobic fermentation time until the fermentation material forms a black brown fluffy substrate with a soil smell, so that a fungus chaff fermented material is obtained; mixing the fungus chaff fermented material with a PGPR (plant-growth-promoting stress-resistant beneficial microorganisms) bacterium agent, stirring uniformly, and performing stacking activation on beneficial bacteria until the water content of the substrate is 15-20 %, so that a base material is prepared; and uniformly mixing the base material with a biologicalactivated carbon powder and diatomaceous earth according to certain ratios, so that the vegetable cultivation substrates are obtained. According to the method, soil environment can be effectively improved, soil fertility is improved, rich and comprehensive nutrient elements are provided for vegetable cultivation, and waste in agricultural production is fully utilized as a substrate material, so that the method is green and environmentally friendly, and production cost of the vegetable cultivation substrates is reduced.