Soybean is an important oilseed crop that provides high-quality protein and vegetable oil. Salinity constitutes a negative abiotic factor that reduces soybean plant growth, production, and quality. The adsorption of Na+ by chitosan-modified biochar (CMB) has a significant effect on salinity but the application of CMB is limited in soybean. In the current study, CMB was used for characterization of physiological, biochemical, and molecular responses of soybean under salt stress. Comparison of CMB and unmodified (as-is) biochar (BR) demonstrated a significant difference between them shown by using Fourier transform infrared spectroscopy (FTIR), scan electron microscopy (SEM), Brunauer–Emmett–Teller (BET), elemental analysis and z-potential measurement. Pseudo-first and second-order better suited for the analysis of Na+ adsorption kinetics. The salt-stress reduced the soybean plants growth, root architecture characteristics, biomass yield, nutrients acquisition, chlorophyll contents, soluble protein, and sugar contents, while CMB with salt-stress significantly increased the above parameters. Moreover, CMB also reduced the salinity-induced increase in the Na+, glycine betaine (GB), proline, hydrogen peroxide (H2O2), and malondialdehyde (MDA) levels in plants. The antioxidant activity and gene expression levels triggered by salinity but with the application of CMB significantly further boosted the expression profile of four genes (CAT, APX, POD and SOD) encoding antioxidant enzyme and two salt-tolerant conferring genes (GmSALT3 and CHS). Overall, these findings demonstrate the crucial role of CMB in minimizing the adverse effects of high salinity on soybean growth and efficiency of the mechanisms enabling plant protection from salinity through a shift of the architecture of the root system and enhancing the antioxidant defense systems and stress-responsive genes for achieving sustainable crop production.