Abstract: To achieve sustainable development of tobacco production, explore new types of substrate for the growth of flue-cured tobacco, and get rid of its dependence on peat soil substrate, in this paper, the influence of alternative substrate on the germination time, emergence rate, planting percent and the quality of tobacco seedlings is studied by taking local wheat straw, leftover bits and pieces of mushroom and coarse sand common in the place investigated as the raw materials and peat soil as the control group. The results showed that in the group of wheat straw substrate, the rate of emergence and part of the seedlings’ agronomic property were significantly different from CK, but the planting percent was basically the same; in the group of mushroom substrate, the emergence rate was consistent with CK, but there was no seedlings; and in the group of coarse sand substrate, the emergence rate, planting percent and agronomic property were superior to or in proximity to CK; The cost of all these three types of alternative substrate is lower than that of CK, among which the cost of coarse sand substrate is the lowest. Therefore, the substitution effect of coarse sand substrate is the best, so it could be properly publicized as an alternative to peat soil as the substrate formulation for floating seedling.