We resolve a longstanding question regarding the kinship of two high-status Egyptians from the 12th Dynasty, Nakht-Ankh and Khnum-Nakht, whose mummies were discovered in 1907 by Egyptian workmen directed by Flinders Petrie and Ernest Mackay. Although their coffin inscriptions indicate that Nakht-Ankh and Khnum-Nakht were brothers, when the mummies were unwrapped in 1908 the skeletal morphologies were found to be quite different, suggesting an absence of family relationship. We extracted ancient DNA from the teeth of the two mummies and, following hybridization capture of the mitochondrial and Y chromosome fractions, sequenced the DNA by a next generation method. Analysis of single nucleotide polymorphisms showed that both Nakht-Ankh and Khnum-Nakht belonged to mitochondrial haplotype M1a1, suggesting a maternal relationship. The Y chromosome sequences were less complete but showed variations between the two mummies, indicating that Nakht-Ankh and Khnum-Nakht had different fathers. Our study emphasizes the importance of kinship in ancient Egypt, and represents the first successful typing of both mitochondrial and Y chromosomal DNA in Egyptian mummies.