The molecular alignment of polymer film has attracted a lot of attentions due to its complicated process affected by processing conditions and dominating polymer applications. Here we demonstrate that the annealing process can not only change the surface morphology of annealed poly(ethylene oxide) (PEO) film significantly but also induce the formation of highly oriented lamellar structure. It indicates that sheaf-like and cluster banding morphology can be both found obviously on the surface of pristine film. However, there is only sheaf-like morphology and sporadical cluster banding cannot be found obviously on the surface of annealed film. Then, there are both tilt and flat-on orientations of PEO crystalline for pristine film. But, single flat-on orientation of PEO crystalline is present and tilt orientation mode is absent for annealed films. Furthermore, the transition of molecular alignment of PEO film via thermal annealing cannot be affected by the thickness and temperature.