Bioarchaeological research must balance scholarly commitment to the generation of new knowledge, descendants’ interests in their collective past, and the now common practice of rapid re-interment of excavated human remains. This paper documents the first results of a negotiated protocol built on the retention of one tooth per archaeologically derived skeleton, teeth that can then be used for destructive testing associated with ancient DNA and stable isotope investigations. Seven archaeological sites dating from the 13th to 16th centuries provided 53 teeth, 10 of which were subdivided between DNA and isotope labs. All tooth roots yielded haplogroup results, and five provided more detailed sequence results. Stable isotopes of carbon and nitrogen document heavy reliance on maize among all individuals, as well as reliance on a diverse range of fish. This work establishes baseline mtDNA information for Northern Iroquoians, and confirms the value of using dental tissues for dietary reconstruction. Particularly when human remains are fragmentary or co-mingled, this approach holds promise for ongoing incorporation of bioarchaeology into reconstructions of past peoples’ lives.