title: Direct-to-PCR tissue preservation for DNA profiling.
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.
authors.affiliation: National Centre for Forensic Studies, Faculty of Education, Science, Technology & Mathematics (ESTeM), University of Canberra, Canberra, Australia.
authors.forename: Amy
authors.initials: A
authors.lastname: Sorensen
authors.affiliation: National Centre for Forensic Studies, Faculty of Education, Science, Technology & Mathematics (ESTeM), University of Canberra, Canberra, Australia.
authors.forename: Clare
authors.initials: C
authors.lastname: Berry
authors.affiliation: National Centre for Forensic Studies, Faculty of Education, Science, Technology & Mathematics (ESTeM), University of Canberra, Canberra, Australia.
authors.forename: David
authors.initials: D
authors.lastname: Bruce
authors.affiliation: National Centre for Forensic Studies, Faculty of Education, Science, Technology & Mathematics (ESTeM), University of Canberra, Canberra, Australia.
authors.forename: Michelle Elizabeth
authors.initials: ME
authors.lastname: Gahan
authors.affiliation: National Centre for Forensic Studies, Faculty of Education, Science, Technology & Mathematics (ESTeM), University of Canberra, Canberra, Australia.
authors.forename: Sheree
authors.initials: S
authors.lastname: Hughes-Stamm
authors.affiliation: National Centre for Forensic Studies, Faculty of Education, Science, Technology & Mathematics (ESTeM), University of Canberra, Canberra, Australia. dennis.mcnevin@canberra.edu.au.
authors.forename: Dennis
authors.initials: D
authors.lastname: McNevin
mesh_terms: D003451:Cryoprotective Agents; D004247:DNA / Q000032:analysis*; D016172:DNA Fingerprinting / Q000379:methods*; D004121:Dimethyl Sulfoxide; D004492:Edetic Acid; D005838:Genotype; D006801:Humans; D018895:Microsatellite Repeats; D018482:Muscle, Skeletal / Q000737:chemistry; D016133:Polymerase Chain Reaction; D012867:Skin / Q000737:chemistry; D013048:Specimen Handling / Q000379:methods; D014021:Tissue Preservation
chemical_list: D003451:Cryoprotective Agents; D004247:DNA; D004492:Edetic Acid; D004121:Dimethyl Sulfoxide
keywords: DNA profile; Direct PCR; Disaster victim identification (DVI); Mass disaster; PCR inhibition; Tissue preservation