Abstract: Cucumber (Cucumis sativus L. ‘Xinchun 4’) was used to explore the relationship between nitric oxide (NO) and calcium (Ca²⁺) during adventitious rooting under drought stress. Rooting parameters, endogenous Ca²⁺ fluorescent intensity and the antioxidant enzymes activity (SOD, CAT and APX) in cucumber explants under drought stress were investigated. The results showed that treatment with 200 μmol·L^-1 CaCl₂ and 0.05% PEG significantly improved the number and length of adventitious root in cucumber explants under drought stress, while the application of Ca²⁺ chelating agent (EGTA) and channel inhibitor (BAPTA/AM) significantly decreased NO-induced number and length of adventitious root under drought stress. Under drought stress, the fluorescence intensity of Ca²⁺ in hypocotyls treated with NO and CaCl₂ was improved, however, the Ca²⁺ fluorescence intensity in the hypocotyls treated with NO scavenger (cPTIO) was significantly lower than that in the hypocotyls treated with NO. Under drought stress, the activities of antioxidant enzymes in the cucumber explants were significantly promoted by the treatments with NO and CaCl₂, however, Ca²⁺ chelating agent and channel inhibitor significantly decreased the activity of antioxidant enzymes induced by NO. In conclusion, Ca²⁺ might be involved in the process of NO-adjusted antioxidant enzymes activity during adventitious rooting under drought stress, which alleviated the negative effects of drought on the adventitious rooting and promoted the formation of adventitious roots.