Saline–alkali stress severely limits crop production worldwide. Soybean [Glycine max (L.) Merr.] is particularly sensitive during seed germination, a stage critical for stand establishment. This complex stress environment encompasses two distinct yet equally critical dimensions: neutral salt stress and alkaline salt stress, each eliciting specialized physiological and metabolic responses. Here, a comparative assessment of four genotypes (tolerant: CN16, CN17; sensitive: Williams 82, K18) under 100 mmol/L Na⁺ revealed that alkaline salt stress exerts a significantly more potent inhibitory effect than neutral salt stress. Tolerant cultivars maintained 75–80% germination under alkaline conditions, whereas sensitive ones dropped below 15%, a divergence primarily driven by superior oxidative mitigation capacity. Integrated multi-omics analysis of the tolerant variety CN16 identified stage-specific regulatory shifts: early alkaline salt stress (60 h) triggers extensive transcriptional reprogramming focused on physical barrier reinforcement, including cell walls and lipid remodeling. By 96 h, regulatory modes between the two stress types diverged: neutral salt elicited a transcriptional surge, while alkaline salt transitioned to a metabolically dominant regulation, shifting flux from growth-related isoflavonoids to defense-related anthocyanins. Crucially, this study uncovers the distinct bioenergetic trade-offs governing these responses: whereas adaptation to neutral salt relies on low-energy galactose metabolism, tolerance to alkaline salt demands energy-intensive processes, specifically the active vacuolar compartmentalization of organic acids and anthocyanins for intracellular buffering. This obligatory energy expenditure restricts biomass accumulation, explaining the severe growth penalties observed in complex saline-alkali environments. Finally, the identification of a core regulatory module driven by key genes, including GmPHOT2b, GmLOG, and GmSHMT08, enriches the metabolic regulatory network under saline-alkali stress, providing core targets and a theoretical framework for precisely balancing metabolic expenditure with biomass accumulation in breeding practice.

Soybean (Glycine max) germination is highly sensitive to neutral salt stress. Although sodium chloride (NaCl) and sodium sulfate (Na₂SO₄) co-exist in nature, their distinct phytotoxic mechanisms remain severely under-investigated. In this study, 50 germplasm accessions were systematically screened, identifying R014 as highly salt-tolerant and R120 as highly sensitive. Phenotypic and dynamic antioxidant monitoring (0–72 h) established 48 h as the critical tolerance window, revealing that Na₂SO₄ induces complex physical damage (crystallization) and osmotic injury, with its ionic toxicity significantly exceeding that induced by NaCl. Crucially, R014 effectively maintained peak activities of antioxidant enzymes (SOD, POD, CAT) to combat these specific stressors. By integrating deep RNA sequencing with weighted gene co-expression network analysis (WGCNA) using 48 h radicle data, significant transcriptomic reprogramming was revealed. WGCNA robustly isolated 35 functional modules, located five key phenotypic clusters, and defined three major hub genes (Glyma.11G101900, Glyma.17G185000, and Glyma.20G247850) that regulate calcium signaling. Verified by qRT-PCR, this study suggests the differential physiological and molecular architectural characteristics between chloride and sulfate toxicities, providing precisely targeted genetic loci for the breeding of salt-tolerant soybean.