||This study set out to sequence the hypervariable segment-I (HVS-I) of the mitochondrial genome from prehistoric skeletal remains associated with Aleut, Sadlermiut, Dorset and Thule groups in Northern North America in an effort to gain insight into their genetic prehistories. Sequences obtained from said ancient populations (Aleut n=6; Sadlermiut n=7; Thule (partial sequences) n=3) were compared to each other as well as those from contemporary and prehistoric populations in the surrounding area. The prehistoric populations under investigation harbored matrilineages typically found in circum-Arctic populations throughout time: A2, A2a, A2b1, D2/D2a‚Äôb/D2a/D2a1 and D4b1a2a1. Ancient Aleuts exhibited HVS-I polymorphisms associated with haplogroups A2a, D2 and D2a‚Äôb, while the Sadlermiut were characterized as A2b1 and D4b1a2a1. Partial Thule HVS-I sequences indicate A2 but preclude definitive assignment to A2, A2a or A2b1 until the remaining portion of HVS-I is sequenced. The results indicate ancient Aleuts across time exhibit affinities with the UnangaxÃÇ (modern Aleuts); however, population movement or genetic exchange with neighbors to the east cannot be ruled out at this time. Ancient Aleuts were also found to have a greater matrilineal genetic similarity to Chukotkan populations (Chukchi and Siberian Yuit), rather than those from Kamchatka (Koryak and Itel‚Äômen). This genetic similarity/dissimilarity provides additional corroboration for colonization of the Aleutian archipelago being initiated from the east rather than the west. The isolated eastern Arctic Sadlermiut population, on the other hand, was shown to have affinities with contemporary Eskimo (Inuit and I√±upiat). The implications of this points towards the Sadlermiut having Neo-Eskimo rather than Paleo-Eskimo ancestry and echoes previous findings of matrilineal discontinuity in the eastern Arctic. The mtDNA (mitochondrial deoxyribonucleic acid) profiles of the ancient populations in this study are also congruent with results from other mtDNA studies indicating the genetic prehistory of the Neo-Eskimo was distinct from that of the Paleo-Eskimo and inhabitants of the Aleutians. Overall the findings in this study speak to matrilineal genetic relationships of prehistoric and contemporary populations in the most northern stretches of the New World while touching upon population movements in the region.