Abstract: Cultivating strong seedlings is an important guarantee for the production of high-quality tobacco leaves, while there are many disadvantages in tobacco floating system that was commonly adopted in China. To improve the tobacco floating system, ‘Xiangyan No.3’ was used as material, the effects of water control before transplanting and rooting powder treatments on tobacco seedling quality and physiological properties at green stage were studied. The results showed that: (1) the effect of water control on tobacco seedling quality was little, while the effects of rooting powder treatment and water control rooting powder treatment were significant, mainly shown as plant height decreasing, stem thickening and dry matter accumulation increasing; (2) water control before transplanting and rooting powder treatment could improve leaf chlorophyll content and roots activity, and the effect of water control rooting powder treatment was the best, and that of rooting powder treatment was the next, and that of water control was the worst; (3) all treatments could increase activities of nitrate reductase and converting enzyme of tobacco seedlings, and decrease malondialdehyde content in tobacco seedlings, and the effect of water control rooting powder treatment was the best, and that of rooting powder treatment was the next, and that of water control was worst. Meanwhile, the effects of treatment 10 days before transplanting were better than those of treatment 5 days before transplanting. Comprehensively, the improvement effects of water control 10 days before transplanting rooting powder treatment on quality of tobacco seedling and physiological properties at green stage were the best.