A method of fabricating a semiconductor device, the method including etching a portion of a substrate including a first region and a second region to form a device isolation trench; forming a device isolation layer defining active regions by sequentially stacking a first insulating layer, a second insulating layer, and a third insulating layer on an inner surface of the device isolation trench; forming word lines buried in the substrate of the first region, the word lines extending in a first direction to intersect the active region of the first region, the word lines being spaced apart from each other; forming a first mask layer covering the word lines on the substrate of the first region, the first mask layer exposing the substrate of the second region; forming a channel layer on the substrate of the second region; and forming a gate electrode on the channel layer.