The invention discloses a seedling hardening method for significantly improving the transplanting survival rate of cassava tissue cultured seedlings, which comprises the following steps: retaining the mature stem segments of leaves of cassava, subcultured on a seedling culture medium for 33 to 37 days to obtain tissue cultured seedlings; Take out the tissue-cultured seedlings and wash them, clamp the stems with planting cotton at a place 0.5cm higher than the root system, and transplant them into peat soil pits for planting; cover the potted plants with fresh-keeping bags, shade them all the way, and keep moisture After a few days, the bag is removed, and the planting process and 6-12 hours after planting are carried out indoors; after removing the bag, cultivate for 12-16 days, transfer the cassava plants to the formula soil and remove the planted cotton, and cultivate for 14-21 days to complete refining. Seedlings, the formula soil is 4:1:1:1:1 peat soil, humus soil, vermiculite, perlite and red soil. Through the controllable subculture of the mature stems of the cassava leaves, and then using the cotton planted in the peat soil, and then transferred to a specific formula soil to cultivate good quality seedlings, the survival rate was significantly increased from 50% to 95%. %.