P227 – No Root, no Problem: Comparison of Four DNA Extraction Protocols for mtDNA Recovery fromHhair Shafts in Forensic Casework

10:37

Thursday August 20^th

Station 14

Duration: 12 minutes

04. Forensic biology

Christina Amory

Shed hair shafts are among the most frequently recovered biological traces at crime scenes, yet they remain one of the most challenging sample types for forensic genetic analysis, as DNA quantity is low and degradation is often substantial. High-quality nuclear DNA is frequently absent, making mitochondrial DNA (mtDNA) the primary target. Therefore, the DNA extraction method is critical for obtaining reliable results.

Our study systematically evaluated two manual and two automated DNA extraction approaches for their efficiency in recovering mtDNA from hair shafts. The extraction strategies were compared based on their ability to recover mtDNA fragments of different lengths, which can vary in detectability in degraded samples. All samples comprised hair shafts from which roots and hair sheaths had been removed prior to extraction. The dataset included (i) casework-like archived hairs collected in 2009 and 2012, (ii) hairs from the same donors freshly collected in 2026, and (iii) samples relevant to ancient DNA research. The ancient DNA samples consisted of rootless hair cuttings approximately 150 years old that had been stored at room temperature and included both pigmented (black) and non-pigmented (white/gray) hairs. Because these were cuttings, neither hair orientation nor distance from the scalp could be determined.

Extracts were quantified using SDquants, a quadruplex real-time PCR assay including mtDNA amplicons of 69 bp and 143 bp, to provide information on mtDNA quantity and fragmentation. In addition, a subset of extracts was sequenced to corroborate the quantification results. This combined quantitative and sequencing-based approach supports the evaluation of extraction-method suitability for forensic mtDNA analysis of limited and degraded hair shaft samples.

Authors

Christina Amory (Institute of Legal Medicine, Medical University of Innsbruck, Austria)
Gabriela Huber (Institute of Legal Medicine, Medical University of Innsbruck, Austria)
Harald Niederstätter (Institute of Legal Medicine, Medical University of Innsbruck, Austria)
Walther Parson (Institute of Legal Medicine, Medical University of Innsbruck, Austria)

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P227 – No Root, no Problem: Comparison of Four DNA Extraction Protocols for mtDNA Recovery fromHhair Shafts in Forensic Casework

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