A semiconductor device including a plurality of active regions extending in a first direction on a substrate; a device isolation layer between the plurality of active regions such that upper portions of the plurality of active regions protrude from the device isolation layer; a first gate electrode and a second gate electrode extending in a second direction crossing the first direction and intersecting the plurality of active regions, respectively, on the substrate, the first gate electrode being spaced apart from the second gate electrode in the second direction; a first gate separation layer between the first gate electrode and the second gate electrode; and a second gate separation layer under the first gate separation layer and between the first gate electrode and the second gate electrode, the second gate separation layer extending into the device isolation layer in a third direction crossing the first direction and the second direction.