The invention discloses a seedling culture matrix and a preparation method thereof and relates to the field of plant seedling culture. The seedling culture matrix is prepared from, by mass, sandy soil, magnesium sulfate, thoroughly decomposed cow dung, a water absorption water-retaining agent, carbonized rice husks, palm bark fibers, calcium magnesium phosphate fertilizer, nitrogen-phosphorus-potassium mixed fertilizer, ammonium sulfate, monoammonium phosphate, boric acid, polyaspartic acid, glucosamine oligosaccharides, a plant disease protective agent and shock-resistant bacteria. By the adoption of the technical scheme, the matrix has the functions that fertilizer efficiency is sufficient, nutrients are slowly released, disease resistance and stress tolerance are achieved, the nutrients are comprehensive, and pertinence is high, and all indexes reach industrial standards. Through volume production and use in multiple seedling culture factories, the seed germination rate and emergence rate can be increased, seedlings emerge in order and grow vigorously, the seedlings are strong and are free of excessive growth, root macerating, dwarfing and ossification, flower bud differentiation of the seedlings is normal, and the properties, yield and quality of crop varieties are guaranteed.