Abstract: This study examined whether exogenous melatonin (MT) promotes adventitious root induction in tissue-cultured elongated shoots of Sapium sebiferum (L.) Roxb. under salt stress and clarified the associated physiological and biochemical responses. Explants were cultured on 1/2 MS medium containing 3.0 g/L NaCl and supplemented with MT at concentrations of 10, 25, 50, and 100 μmol/L. Morphological traits, soluble sugar and soluble protein contents, antioxidant enzyme activities, including superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT), and chlorophyll fluorescence parameters were assessed. Results revealed that MT markedly improved adventitious root formation under salt stress, with 25 μmol/L producing the strongest rooting response. MT treatment across different stages effectively enhanced salt tolerance of seedlings. The maximum increases in soluble sugar and soluble protein contents exceeded 30%, and the maximum increases in SOD, POD, and CAT activities were 61.21%, 12.65%, and 51.37%, respectively. MT also improved photosynthetic performance under salt stress. Maximum quantum yield (QY_max) increased by 7.34%–9.42%, steady-state light-adapted photochemical quenching (qp_Lss) increased by a maximum of 124.01%, and the steady-state fluorescence decline ratio (Rfd_Lss) increased by up to 91.96%. In contrast, steady-state non-photochemical quenching (qN_Lss) decreased by at least 35.01%.