Egypt, located on the isthmus of Africa, is an ideal region to study historical population dynamics due to its geographic location and documented interactions with ancient civilizations in Africa, Asia, and Europe. Particularly, in the first millennium BCE Egypt endured foreign domination leading to growing numbers of foreigners living within its borders possibly contributing genetically to the local population. Here we mtDNA and nuclear DNA from mummified humans recovered from Middle Egypt that span around 1,300 years of ancient Egyptian history from the Third Intermediate to the Roman Period. Our analyses reveal that ancient Egyptians shared more Near Eastern ancestry than present-day Egyptians, who received additional Sub-Saharan admixture in more recent times. This analysis establishes ancient Egyptian mummies as a genetic source to study ancient human history and offers the perspective of deciphering Egypt’s past at a genome-wide level.