2, 5-Dimethylfuran (DMF) has been receiving increasing interest as a potential alternative fuel to fossil fuels, owing to the recent development of new production technology. However, the influence of DMF properties on the in-cylinder fuel spray and its evaporation, subsequent combustion processes as well as emission formation in current gasoline direct injection (GDI) engines is still not well understood, due to the lack of comprehensive understanding of its physical and chemical characteristics. To better understand the spray characteristics of DMF and its application to the IC engine, the fuel sprays of DMF and gasoline were investigated by experimental and computational methods. The shadowgraph and Phase Doppler Particle Analyzer (PDPA) techniques were used for measuring spray penetration, droplet velocity and size distribution of both fuels. In parallel, numerical simulation was carried out by using KIVA3V code with improvements in the nozzle flow model, spray atomization and secondary breakup models. As compared to the experiment results, the Cascade Atomization Breakup (CAB) model is shown to have a better droplet size prediction than the Kelvin-Helmholtz (KH) and Taylor Analogy Breakup (TAB) spray models. The CAB secondary breakup model together with the Max Planck Institute (MPI) primary atomization model was therefore chosen to investigate the in-cylinder mixture preparation in a GDI engine. It was shown that, both insufficient mixing time and significant spray-wall interaction contribute to relatively poor Air/Fuel distribution with DMF fuel. As compared to the nearly homogeneous mixture with the gasoline fuel spray, up to 9.3% in extra-rich mixture and 4.8% in very-lean mixture were observed in the DMF fuel spray case at the end of compression stroke, and the mixture equivalence ratio around the spark plug was leaner than 0.9.