Abstract: To investigate the physiology and molecular mechanism of grafted tobacco in response to drought, taking drought-tolerant cultivar Longjiang 851 (LJ851) as a rootstock and drought-sensitive cultivar Jinxing 6007 as a scion, self-grafted (JX6007/JX6007) and rootstock-grafted (JX6007/LJ851) tobacco plants were prepared. A pot culture experiment was carried out to study the effects of grafting on the biomass, antioxidant enzyme activities, H₂O₂ and membrane lipid peroxidation in tobacco under drought stress. The results showed that under drought stress, the biomass, chlorophyll contents and antioxidant enzyme activities of JX6007/LJ851 were higher than those of JX6007/JX6007 by different extents, while the membrane lipid peroxidation degree and H₂O₂ content were lower than those of JX6007/JX6007. qRT-PCR indicated that the expressions of stress-responsive genes (LEA5, AREB, CDPK2 and ERD10C) in rootstock-grafted plants were significantly up-regulated. Therefore, grafted tobacco with drought-tolerant rootstock genotype might improve drought tolerance in tobacco via regulating antioxidant enzyme activities and stress-responsive gene expression.