Physiological changes associated with enhanced cold resistance during Medicago ruthenica germination and seedling growth in response to exogenous γ-aminobutyric acid

Elucidating the effect of γ-aminobutyric acid (GABA) on seed germination and seedling growth of Medicago ruthenica under low-temperature could provide a reference for alleviating the harmful effects of low temperatures on legumes in alpine meadows. We set up different temperatures to screen three low temperatures that inhibit seed germination of Medicago ruthenica, and six aqueous concentrations of GABA (0, 0.1, 0.5, 1, 5, 10 mM) were set up to screen out the best GABA seed soaking concentration that can promote the seed germination at low-temperature. The three temperatures of 10 °C, 20/5 °C and 15/5 °C inhibited seed germination of Medicago ruthenica. Soaking seeds with 1 mM GABA could significantly increase seed germination rate of Medicago ruthenica at low-temperature (10 °C). Soaking seeds only with 0.1 mM GABA could promote germination of Medicago ruthenica at 20/5 °C, but there is no significant difference compared with distilled water soaking. Whereas, all GABA seed soaking concentrations delayed seed germination time and inhibited seed germination at 15/5 °C. Pre-spraying seedlings with 1 mM GABA at 10 °C reduced the production of superoxide anion, hydrogen peroxide and malondialdehyde content, and increased the activities of catalase, peroxidase, superoxide dismutase and ascorbate peroxidase in leaves of Medicago ruthenica on days 7. Simultaneously, Pre-spraying seedlings with 1 mM GABA at 10 °C significantly increased the net photosynthetic rate and decreased intercellular CO₂ concentration. These data suggest that GABA could enhance the cold tolerance of Medicago ruthenica by promoting seed germination, regulating the antioxidant system, and increasing photosynthetic efficiency. However, the mitigation effect of GABA on low-temperature is only applicable to a certain temperature. The mitigation effect of GABA on low temperature will be weakened as the low temperature of plants is less than 10 °C