Strategy for STR typing of bones from the Second World War combining CE and NGS technology: A pilot study

The genetic identification of skeletal remains found in Second World War mass graves is complicated because of the poor quality of the samples. The aim of this study was to set up a workflow for STR typing of such samples combining PCR/CE and PCR/NGS technologies. To this end, 57 DNA samples from an equal number of 75-year-old femurs were studied. After a first round of PCR typing using GlobalFiler CE, 42 samples yielded a full profile and were therefore submitted to our standard workflow. The 15 samples that yielded no or a limited number (2-17/21) of autosomal STR markers as well four bone control samples that provided a full profile with the conventional PCR/CE test were typed in duplicate by the GlobalFiler NGS kit. Despite the degradation of the samples, which resulted in lower coverage and a lower % of on-target reads, reliable sequencing data were obtained from 16/19 samples. The use of a threshold of 30× for the locus call led to a consensus profile (cp) of 20-31/31 STR autosomal loci in 10 samples and to a cp of 8-10/31 loci in two samples, whereas the four control samples yielded a cp of 26-31/31 loci. Finally, the data of the NGS typing were combined with those of the CE typing. This last task allowed us to recover (on average) three alleles per sample and to increase the number of the heterozygous patterns in 37 cases. In total, the combined approach proposed here made possible the genetic typing of 65-100% of the autosomal STR markers in 10/15 (66.6 %) skeletal remains that yielded no or very poor results with the conventional PCR/CE approach. However, because several artefacts (such as allelic drop-out and allelic drop-in) were scored, the risk of mistyping cannot be neglected.