Crop lignocellulose residues represent a renewable and abundant biomass resource for biofuels and other chemicals. In terms of bioethanol production, biomass process principally involves in three major steps: physical and chemical pretreatments leading to wall polymer destruction, enzymatic digestion releasing soluble sugar and yeast fermentation producing bioethanol. However, lignocellulose recalcitrance leads to an unacceptably costly biomass conversion, due to complicated structures and diverse biological functions of plant cell walls. To meet the accelerating worldwide demand for liquid transportation fuels, conversion of cellulosic biomass into advanced biofuels will require a good and low cost pretreatment methods. Our group have performed systems biology analyses between cell wall composition/structure and biomass digestibility in four representative wheat natural-germplasms and 2 typical rice genetic-mutants. Both plant samples displayed a diverse cell wall structure and varied biomass saccharification, suggesting that cell wall structure can fundamentally determine biomass enzymatic digestibility upon various chemical and physical pretreatments. Firstly, a combined pretreatment was performed in four wheat accessions using steam explosion followed with different concentrations of H2SO4 or NaOH, leading to increased hexoses yields by 3–6 folds from enzymatic hydrolysis. Secondly, two rice cultivars were collected from experimental fields with seven nitrogen fertilizer treatments. All biomass samples contained significantly increased cellulose contents and reduced silica levels, with variable amounts of hemicellulose and lignin from different nitrogen treatments. Hence, these studies have demonstrated that nitrogen fertilizer supply could largely enhance biomass saccharification in rice cultivars, while mild pretreatment technology for a complete saccharification and high ethanol production in wheat crop.