To analyze the effect of polyol on polyurethane (PU)-modified asphalt, three different soft segments of polyurethane were synthesized, and we utilized the reaction of MDI (diphenylmethane diisocyanate) with PU650, PU1000, and PU1400. With respect to molecular weight, the effect of polyol on the performance of modified asphalt was analyzed, and the asphalt was modified by using three different polyurethanes. To analyze the PU samples, the Fourier transform infrared spectroscopy (FTIR) tests and gel permeation chromatography (GPC) tests were selected; by contrast, to analyze the rheological properties and modification mechanism of asphalt, the dynamic rheology test (DSR), low-temperature bending creep test (BBR), multi-stress repetitive creep test (MSCR), FTIR, and differential scanning calorimetry (DSC) were selected. The results indicate that the molecular weight of polyol affects the molecular structure of polyurethane, the distribution of soft and hard segments, the content of soft segments, and the distribution of asphaltene in asphalt; thus, the asphalt modification effect occurs differently. The storage stability and high-temperature stability of the polyurethane-modified asphalts that were synthesized using three different polyols (i.e., polyols that exhibit different molecular weights) did not differ considerably, and the PU1400-modified asphalt exhibited the best low-temperature performance.