The fodder soybean (Glycine max) is an excellent leguminous forage with a high protein content and hay yield, cultivated comprehensively in alpine regions, but seasonal drought in northern regions severely impacts the growth of seedlings. Melatonin (MT) and strigolactone (SL) are critical in relieving the restraint of plant growth in water-deficient environments, but the mechanisms of MT- and SL-mediated drought resistance in fodder soybean needs to be explored. This study mainly investigated the potential morphophysiological mechanism of MT and SL treatments in protecting fodder soybean from drought stress. The fodder soybean ‘Gongnong 535’ was treated with 100 µM MT or 1 µM SL under normal, moderate, and severe water deficit conditions. The results showed that MT and SL treatments enhanced the plant growth parameters and stomatal aperture under drought stress. Moreover, the observed reductions in superoxide ion (O2.-), malondialdehyde (MDA), and relative electrical conductivity (REC), along with enhancements in the activities of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT), as well as higher levels of ascorbate (AsA), glutathione (GSH), soluble sugar (SS), soluble protein (SP), and free proline (Pro), indicated that MT and SL application effectively alleviated the oxidative damage and prevented the cell membrane disruption caused by drought stress. Additionally, MT and SL treatments improved photosynthesis and growth in fodder soybean seedlings under water stress by adjusting chlorophyll pigments, gas exchange indexes, and chlorophyll fluorescence parameters, as well as endogenous hormone levels. Simultaneously, MT and SL influenced the expression of genes associated with photosynthesis and antioxidant defenses, as well as phytohormone concentrations. Notably, the protective effect of the SL treatment was superior to that of MT in water-deficient conditions. This study contributes to further understanding the defensive mechanism of MT and SL against drought stress.
