*4.1. Plant Materials and Growth Conditions*

Rice (*Oryza sativa* cv. Qian You No. 0508) seeds were purchased from the Seed Production Unit, Jiangsu Academy of Agricultural Sciences, China. Before priming, seeds were sterilized with 0.5% NaClO solution for 15 min and washed several times to remove the traces of the disinfectant. The seeds were then primed with GABA at the optimized concentration (0 and 0.5 mM) at 15 ◦C in darkness for 24 h. The time and concentration of the priming agent were initially selected based on a preliminary study. In the preliminary experiment, several concentrations of GABA, i.e., 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6 and 0.7 mM were used for seed priming. GABA at 0.5 mM significantly improved rice germination and seedling growth as compared to other concentrations. The seed were primed for 24 h as this time was sufficient enough for rice seeds to trigger the activation of various metabolic processes such as the synthesis of hydrolytic enzymes which resulted in hydrolysis of reserve food into a simple available form for embryo uptake as stated recently in our previous study [82–85]. The primed seeds were dried at room temperature to maintain their original moisture content [82]. Thereafter, the primed and unprimed seeds were germinated for two weeks in a plastic germination box containing two layers of germination paper moistened with water. Fifty seeds and three replications for each treatment were used. Then, seeds were incubated in a germination chamber at 25 ◦C with 80 % relative humidity under alternating cycles of 16 h illumination and 8 h darkness for 14 days.

The salinity and osmotic stress and their combination were applied to 7-day-old rice seedlings, in which the salinity stress (150 mM NaCl) and the osmotic stress (30 g/L PEG, 6000) and their combination (150 mM NaCl+30 g/L PEG) were supplied to plants for one week. The seedlings without GABA priming and stress treatments were used as the control (Ck). After fourteen days, the germination

percentage, germination energy, root and shoot length, seedling fresh and dry weight and seedling vigor index were measured according to the methods of Hu et al. [86].
