Plant-based renewable biofuels guarantee sustainable solutions to food and energy demands. High-biomass C4 grasses including sugarcane, corn, and sorghum are potential candidates for bioenergy. Among these, sorghum enjoys the status of a highly diverse food, feed, and biofuel source worldwide. The natural attributes like abiotic stress tolerance, diverse genetic base, viable seed industry, and sound breeding system make sorghum a perfect candidate for establishing an efficient and low-cost biofuel industry. Scientists are exploring ways to exploit forage, sweet, and biomass sorghums as climate-smart energy crops. In this context, conventional breeding has played a significant role in developing high-yielding sorghum varieties. For biomass sorghum, stem compositional analysis helps screen low lignin and high polysaccharide types as feedstocks for biofuels. Recent tools of phenomics, genomics, proteomics, and genome editing are key players of designing eco-friendly bioenergy sorghum. Here, we report stem compositional analysis and proteomicsbased evaluation of USDA sorghum germplasm as a baseline to develop sorghum as a biofuel feedstock.