The vibrational transition moment directions of 1,3-dimethyluracil (DMU), 1,3-dimethyluracil-5-d (DMU-5-d), and 1,3-dimethyluracil-6-d (DMU-6-d) have been determined from measurements of polarized IR spectra on samples partially aligned in stretched poly(vinyl alcohol) and by ab initio calculations at the HF/6-31G* level of approximation. The agreement between experiments and theory is good. IR spectra of DMU, DMU-5-d, and DMU-6-d in KBr and in D2O form, together with the calculated spectra and the IR polarizations, the basis for some reassignments. Special attention is paid to the double-bond region, where the high-frequency carbonyl stretching mode is assigned to an in-phase vibration of the C4O and the C2O. This vibration is polarized in a direction almost bisecting the angle between the two carbonyl bonds, in agreement with an in-phase assignment. The low-frequency mode is the out-of-phase vibration of the two carbonyl groups, and it is consequently polarized perpendicular to the high-frequency mode. In order to determine the orientation axis of DMU in the polymer matrix, the UV linear dichroism (LD) spectrum was measured and the polarizations of the electronic transitions were estimated from a semiempirical self-consistent reaction field (SCRF) calculation including solvent effects. The combined information from IR LD and UV LD shows that the molecular orientation axis in the stretched film experiment is parallel to the first pi --> pi* transition in DMU.