Abstract:[Background] The contradiction between edible fungi and forest is increasingly prominent, and the agricultural residues are rich in resources, which can be used as the main substrate for the cultivation of edible fungi. [Objective] To screen out the agricultural residue formula suitable for the growth of Auricularia auricula mycelia by using common agricultural residues. [Methods] Six substrates, including soybean straw, rape straw, corn straw, peanut straw, wheat straw, and sawdust, were used as raw materials in this study, and the formula was designed using the simplex-lattice method. The effects of different substrate interactions on the mycelial growth rate, mycelial growth index, laccase activity, polyphenol oxidase activity, and cellulase activity of A. auricula were analyzed. [Results] Among the substrates of agricultural residues, soybean straw was the most suitable for the growth of A. auricula mycelia, followed by rape straw. The combination of the three main ingredients optimized the substrate ratio which was most suitable for the growth of A. auricula mycelia. [Conclusion] This study has finally optimized an agricultural residue formula suitable for the growth of A. auricula mycelia (49.4% sawdust, 16.4% rape straw, 12.2% soybean straw, 20% wheat bran, 1% sugar, and 1% CaSO4). This study provides a theoretical basis for the cultivation of A. auricula with grass instead of wood.