The invention provides nutrient soil for tobacco two-stage seedling cultivation and a preparation method of the nutrient soil. The nutrient soil related to by the invention is prepared from 100 partsof disinfected soil, 20-50 parts of decomposed straw and 1-2 parts of special seedling fertilizer. The materials are easy to obtain, the process is simple, the cost is low, and the nutrient soil is green and environmentally friendly, and is easy for large-scale production. The preparation method related to by the invention comprises two methods. The first method comprises the following steps: S1,decomposing straw, S2, carrying out soil disinfection, S3, preparing the special seedling fertilizer, and S4, preparing the nutrient soil. The second method comprises the following steps: S1, decomposing corn, S2, carrying out soil disinfection, S3, preparing the special seedling fertilizer, and S4, preparing the nutrient soil. The nutrient soil provided by the invention provides a nontoxic environment for the conversion of tobacco seedling aquatic roots to xerophilic roots, the morbidity of tobacco seedlings can be effectively reduced, the root activity of the tobacco seedlings can be obviously improved, the development of the root system is promoted, and the seedling recover stage after transplanting is shortened. In addition, according to the invention, a new application channel is alsoprovided for agricultural wastes, and the nutrient soil conforms to green agriculture.