Deep eutectic solvent (DES) pretreatment is attractive for the delignification of lignocellulosic biomass, while unable to circumvent the trenchant demand for the higher-temperature operating conditions. Herein, an electro-assisted DES (choline chloride/ethylene glycol = 1:2) strategy was developed for wheat straw pretreatment at room temperature. The rate of lignin removal, hemicellulose removal, cellulose recovery, and cellulose saccharification reached 68.1%, 60.8%, 95.1%, and 82.5%, respectively, which were comparable or superior to the reported efficiency of traditional DES pretreatment methods. The destruction of lignin by electricity and in-situ dissolution of released lignin components with DES enabled effectively the separation of the full components. Notably, the evolution rate of hydrogen in-situ produced during electro-driven DES pretreatment of wheat straw was 50 μmol cm⁻² h⁻¹, and 4.6 g/100 g lipids could be obtained with Trichosporon cutaneum grown on the fractionated cellulose and hemicellulose components. The electro-assisted DES process offers a potential platform for lignocellulosic biomass fractionation at ambient conditions. According to the life cycle cost analysis (LCCA), the estimated cost of producing hydrogen from 100 g of wheat straw is only $37.24, demonstrating its potential for commercial viability.