Abstract: Dimethyl sulfoxide (DMSO) plays an important role in chemistry, biology and industry. The S=O stretching vibration serves as a sensitive probe of its structure under different chemical environments. However, spectral assignment for the S=O stretching vibration of liquid DMSO remains unclear due to the interference of CH₃ antisymmetric rocking and different aggregates, leading to the confusion in explaining the microstructures of liquid DMSO. Here, the S=O stretching spectra in pure DMSO and in extremely diluted CCl₄ solution were investigated with temperature-dependent Raman spectroscopy. It is shown that CH₃ antisymmetric rocking does not significantly interfere with the S=O band profile due to its low Raman intensity. Three decomposed peaks at 1027, 1042 and 1055 cm⁻¹ in liquid DMSO show obviously different intensity variety upon the temperature increasing. Combined with quantum chemistry calculation on (DMSO)_n (n=1–4) clusters, Raman spectra in the S=O stretching region were assigned and the simultaneous presence of the monomer, dimer and trimmer in liquid DMSO were concluded. These results contribute to a more detailed molecular understanding of the structure of DMSO in the liquid state.