SNP analysis of challenging bone DNA samples using the HID‑Ion AmpliSeq™ Identity Panel : facts and artefacts

Fattorini, Paolo (author), Previderè, Carlo (author), Livieri, Tommaso (author), Zupanc, Tomaž (author), Zupanič-Pajnič, Irena (author)

PCR-MPS is an emerging tool for the analysis of low-quality DNA samples. In this study, we used PCR-MPS to analyse 32 challenging bone DNA samples from three Second World War victims, which previously yielded no results in conventional STR PCR-CE typing. The Identity Panel was used with 27 cycles of PCR. Despite that we only had an average of 6.8 pg of degraded DNA as template, 30 out of 32 libraries (93.8%) produced sequencing data for about 63/90 autosomal markers per sample. Out of the 30 libraries, 14 (46.7%) yielded single source genetic profiles in agreement with the biological identity of the donor, whereas 12 cases (40.0%) resulted in SNP profiles that did not match or were mixed. The misleading outcomes for those 12 cases were likely due to hidden exogenous human contamination, as shown by the higher frequencies of allelic imbalance, unusual high frequencies of allelic drop-ins, high heterozygosity levels in the consensus profiles generated from challenging samples, and traces of amplified molecular products in four out of eight extraction negative controls. Even if the source and the time of the contamination were not identified, it is likely that it occurred along the multi-step bone processing workflow. Our results suggest that only positive identification by statistical tools (e.g. likelihood ratio) should be accepted as reliable; oppositely, the results leading to exclusion should be treated as inconclusive because of potential contamination issues. Finally, strategies are discussed for monitoring the workflow of extremely challenging bone samples in PCR-MPS experiments with an increased number of PCR cycles.

• Language: English
• Keywords: bone DNA, SNP analysis, DNA degradation, SNPs, PCR-MPS, LT-DNA
• Work type: Article
• Typology: 1.01 - Original Scientific Article
• Organization: MF - Faculty of Medicine
• Publication status: Published
• Publication version: Version of Record
• Year: 2023
• Number of pages: Str. 981-993
• Numbering: Vol. 137, iss. 4
• PID: 20.500.12556/RUL-148106
• UDC: 61
• ISSN on article: 0937-9827
• DOI: 10.1007/s00414-023-03019-9
• COBISS.SI-ID: 154212355
• Publication date in RUL: 27.07.2023

Record is a part of a journal

• Title: International journal of legal medicine
• Shortened title: Int. j. leg. med.
• Publisher: Springer Nature, International Academy of Legal Medicine
• ISSN: 0937-9827
• COBISS.SI-ID: 527892

Licences

• License: CC BY 4.0, Creative Commons Attribution 4.0 International
• Link: http://creativecommons.org/licenses/by/4.0/
• Description: This is the standard Creative Commons license that gives others maximum freedom to do what they want with the work as long as they credit the author.

Secondary language

• Language: Slovenian
• Keywords: DNK kosti, analiza SNP, razgradnja DNK

Projects

• Funder: ARRS - Slovenian Research Agency
• Project number: J3-3080
• Name: Določitev predniškega porekla iz DNA za identifikacijske namene