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Direct-to-PCR tissue preservation for DNA profiling

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Abstract

Disaster victim identification (DVI) often occurs in remote locations with extremes of temperatures and humidities. Access to mortuary facilities and refrigeration are not always available. An effective and robust DNA sampling and preservation procedure would increase the probability of successful DNA profiling and allow faster repatriation of bodies and body parts. If the act of tissue preservation also released DNA into solution, ready for polymerase chain reaction (PCR), the DVI process could be further streamlined. In this study, we explored the possibility of obtaining DNA profiles without DNA extraction, by adding aliquots of preservative solutions surrounding fresh human muscle and decomposing human muscle and skin tissue samples directly to PCR. The preservatives consisted of two custom preparations and two proprietary solutions. The custom preparations were a salt-saturated solution of dimethyl sulfoxide (DMSO) with ethylenediaminetetraacetic (EDTA) and TENT buffer (Tris, EDTA, NaCl, Tween 20). The proprietary preservatives were DNAgard (Biomatrica®) and Tissue Stabilising Kit (DNA Genotek). We obtained full PowerPlex® 21 (Promega) and GlobalFiler® (Life Technologies) DNA profiles from fresh and decomposed tissue preserved at 35 °C for up to 28 days for all four preservatives. The preservative aliquots removed from the fresh muscle tissue samples had been stored at −80 °C for 4 years, indicating that long-term archival does not diminish the probability of successful DNA typing. Rather, storage at −80 °C seems to reduce PCR inhibition.

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Acknowledgments

Some equipment and consumables were provided by the NSW Forensic and Analytical Science Service (Sydney, Australia). The authors thank Dr Heloise Breton for providing valuable feedback on the manuscript; the staff at STAFS (Sam Houston State University, Huntsville, TX, USA) for their assistance and the individuals and families of those who donated their bodies to STAFS for scientific research.

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Authors and Affiliations

  1. National Centre for Forensic Studies, Faculty of Education, Science, Technology & Mathematics (ESTeM), University of Canberra, Canberra, Australia

    Amy Sorensen, Clare Berry, David Bruce, Michelle Elizabeth Gahan, Sheree Hughes-Stamm & Dennis McNevin

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  1. Amy Sorensen
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  2. Clare Berry
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  3. David Bruce
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  4. Michelle Elizabeth Gahan
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  5. Sheree Hughes-Stamm
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  6. Dennis McNevin
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Corresponding author

Correspondence to Dennis McNevin.

Ethics declarations

Approval for collection and use of human tissue samples was granted by the University of Canberra Human Research Ethics Committee (Project number 09-01with extension 14-71). All samples were collected with the informed consent of the donors. For the decomposed samples used in this study, Code 45 of US Federal Regulations part 46102(f) exempts the requirement for Institutional Review Board (IRB) approval regarding the use of human cadaveric samples. All procedures were in accordance with the 1964 Helsinki Declaration and its later amendments.

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The authors declare that they have no competing interests.

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Sorensen, A., Berry, C., Bruce, D. et al. Direct-to-PCR tissue preservation for DNA profiling. Int J Legal Med 130, 607–613 (2016). https://doi.org/10.1007/s00414-015-1286-z

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  • DOI: https://doi.org/10.1007/s00414-015-1286-z

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