The thermogravimetric properties and chemical characterization of rice straw (RS) pretreated by mixed culture of white-rot fungi Phanerochaete chrysosporium (P. chrysosporium) and brown-rot fungi Gloeophyllum trabeum (G. trabeum) were investigated. The mixed fungal pretreatment showed a synergistic effect, which resulted in an energy-efficient pyrolysis of pretreated rice straw. The differences in thermochemical conversion of rice straw before and after fungal pretreatment were investigated using thermogravimetric analysis and the Flynn–Wall–Ozawa (FWO) method. Furthermore, the pretreated samples were also analyzed by Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), and scanning electron microscopy (SEM) to illuminate the changes in chemical composition and pyrolysis behavior. Compared to single fungal pretreatment, the mixed fungal pretreatment worked better and exhibited great potential in biomass pyrolysis.