Salinity stress is a growing concern in agriculture, particularly as climate change accelerates soil salinization and limits freshwater availability. Here, we evaluated the effectiveness of classic (low-throughput) versus high-throughput physiological phenotyping methods in detecting early salinity tolerance in Brassica juncea cultivars (‘Carolina Broadleaf’ and ‘Southern Giant Curl’). Traditional phenotyping relies on point measurements such as shoot biomass and leaf gas exchange, which, while valuable, are time-intensive, offer limited temporal resolution, and can be destructive. In contrast, high-throughput phenotyping enables continuous, real-time monitoring of plant physiological responses, providing a dynamic and detailed understanding of stress adaptation mechanisms. We conducted a 42-day experiment in a controlled greenhouse environment, exposing mustard green cultivars to three salinity treatments: control (0.397 dS/m), moderate salinity (10.81 dS/m, ~20% of seawater), and high salinity (24.93 dS/m, ~50% of seawater). The high-throughput PlantArray system was used to measure key physiological parameters, transpiration rates, and net plant weight gain, while traditional phenotyping involved weekly surveys of including stomatal conductance, chlorophyll fluorescence, and biomass accumulation. We found that high-throughput phenotyping allows for earlier and more precise detection of salinity tolerance. Classic methods confirmed significant reductions in biomass, with shoot fresh weight decreasing by up to 80% in high-salinity treatments, but these differences were only detectable at harvest and not before. In contrast, high-throughput phenotyping revealed early signs of osmotic adjustment within the first 20 days, as plants initially maintained transpiration before exhibiting a decline due to ion accumulation. ‘Carolina Broadleaf’ resist moderate salinity, maintaining growth comparable to the control for the first 20 days, suggesting that early harvesting could mitigate yield losses. Overall, this study underscores the advantages of high-throughput phenotyping in improving the precision and efficiency of breeding programs. By integrating continuous physiological measurements, this approach enables earlier and more informed selection of salt-tolerant cultivars, reducing time needed for tolerance screening. Future research should focus on expanding these methods to operational conditions and integrating genomic data to enhance genotype-environment modeling for stress adaptation.
