The identification process of human skeletal remains exhumed from a mass grave from the Spanish Civil War (1936–1939) is presented. Information regarding the presumptive location of the grave and the presumptive number and identities of the persons buried in the grave was collected from interviews and written records from relatives and witnesses, as well as from research at the penitentiary archive. Antemortem individual data were collected from testimonies, and from research from penitentiary, army and civil archives. The consistency between data obtained from testimonies, archives, archeology and osteology allowed a targeted approach to DNA typing based on the assumption of the finding of a closed synchronic group. Two were the first genetic studies requested: the first study focused in the identification of a family group presumptively buried in the grave, compatible with a group of four skeletons that were associated on the basis of dental non-metric traits; the second study focused on the identification of the youngest person presumptively buried at the grave, compatible with the biologically youngest skeleton exhumed. A complete match between 16 Y-STR loci was observed for the four skeletons, as well as a match between mtDNA profiles of the biologically youngest skeleton and the sister of the youngest person presumptively known to be buried in the grave. These results, together with the accumulated evidence, led to propose the identification of these five persons. To date, identifications have been proposed for 17 out of 46 skeletons exhumed from the grave.