Plants
Tomato plants Solanum lycopersicum cv. Better Boy (BB) were grown in cubic plastic pots (3.5” x 3.5” x 3.5”) filled with Metromix 400 potting mix (Griffin Greenhouse & Nursery Supplies, Tewksbury, MA, USA) in a greenhouse at the Pennsylvania State University (25 ± 2°C, 70% ± 10% R.H., 16L: 8D). Plants were fertilized at the one-leaf stage with Osmocote Plus 15-9-12 Fertilizer (ICL Speciality Fertilizers, Summerville, SC, USA). Tomato plants at the four-leaf stage (roughly 4 weeks old) were used in all experiments.
Plants were subjected to three salt treatments:
1. No salt (0 mM): 200 ml of a 0 mM salt solution (distilled water) is added to the base of the plant.
2. Medium/intermediate salt (100 mM): 200 ml of a 100 mM salt solution is added to the base of the plant.
3. High salt (200 mM): 200 ml of a 200 mM salt solution is added to the base of the plant.
100 mM was chosen as an intermediate concentration and 200 mM as the high concentration based on multiple studies (Foolad, 1996; Zhang and Blumwald, 2001; Dombrowski, 2003; Sun et al. , 2010; Munns and Gilliham, 2015; Forieri, Hildebrandt and Rostás, 2016).
Plants were watered with their designated salt solutions every day for three days unless specified otherwise, following which they were used for experiments on the third day. It is worth noting that at the end of the three-day period, the soil salt concentration might be higher than the supplied amount due to salt accumulation in the soil. By the end of the three-day treatment, salt-treated plants looked visibly stressed, dark green in colour, and showed retarded growth.