Extraction of DNA from Human Skeletal Material

Pajnič, Irena Zupanič

doi:10.1007/978-1-4939-3597-0_7

Cited by 51 publications

(45 citation statements)

References 60 publications

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“…Temperature, moisture, and oxygen levels in the depositional environment, soil composition, and pH affect the ability to obtain analyzable DNA from biological specimens [23]. When severe degradation and prolonged aging are present, DNA contained within the bone and teeth is protected [24][25][26][27][28][29][30] and typically persists for much longer than in soft tissues. The type of bone being analyzed also plays a role in DNA typing success, with denser and more compact bones yielding more DNA than non-weight-bearing structures [31,32].…”

Section: Resultsmentioning

confidence: 99%

Identification of human remains using Rapid DNA analysis

Turingan¹,

Brown²,

Kaplun³

et al. 2019

Int J Legal Med

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Rapid identification of human remains following mass casualty events is essential to bring closure to family members and friends of the victims. Unfortunately, disaster victim identification, missing persons identification, and forensic casework analysis are often complicated by sample degradation due to exposure to harsh environmental conditions. Following a mass disaster, forensic laboratories may be overwhelmed by the number of dissociated portions that require identification and reassociation or compromised by the event itself. The interval between the disaster and receipt of victim samples at a laboratory is critical in that sample quality deteriorates as the postmortem interval increases. When bodies decompose due to delay in collection, transport, and sample processing, DNA becomes progressively fragmented, adversely impacting identification. We have previously developed a fully automated, field-forward Rapid DNA identification system that produces STR profiles (also referred to as DNA IDs or DNA fingerprints) from buccal and crime scene samples. The system performs all sample processing and data interpretation in less than 2 h. Here, we present results on Rapid DNA identification performed on several tissue types (including buccal, muscle, liver, brain, tooth, and bone) from exposed human bodies placed above ground or stored in a morgue/cooler, two scenarios commonly encountered following mass disasters. We demonstrate that for exposed remains, buccal swabs are the sample of choice for up to 11 days exposure and bone and tooth samples generated excellent DNA IDs for the 1-year duration of the study. For refrigerated remains, all sample types generated excellent DNA IDs for the 3-month testing period.

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Section: Resultsmentioning

confidence: 99%

Identification of human remains using Rapid DNA analysis

Turingan¹,

Brown²,

Kaplun³

et al. 2019

Int J Legal Med

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“…For extracting genomic DNA from the Babna Gora mass grave tooth and bones, a highly efficient extraction protocol that comprises generation of fine powder, complete demineralisation, and efficient purification was used [10] and successful autosomal STR typing using the Precision ID GlobalFiler NGS STR Panel (TFS) allowed the identification of the aunt with sufficient statistical support and with high confidence of correct identification (Fig. 1).…”

Section: Resultsmentioning

confidence: 99%

Next generation sequencing technology in Second World War victim identification

Pajnič

Pogorelc

Zupanc

2019

Forensic Science International: Genetics Supplement Series

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The killings during the Second World War (WWII), with nearly 100,000 victims, is one of the greatest losses of life in Slovenia's modern history and most of the victims are still buried in hidden mass graves and remain unidentified. Identity, ancestry, and phenotypic SNPs, as well as STR markers are already used for solving various cases with Next Generation Sequencing (NGS) technology. In this study, the Precision ID GlobalFiler NGS STR panel was used to identify the WWII victim that could not be identified with capillary electrophoresis (CE) analyses because limited statistical support was obtained after amplification of autosomal STRs using CE STR kits. Bones and teeth were analysed and compared to family references (nephew and niece on paternal line). Prior to DNA isolation 0.5 g of powder was decalcified. The DNA was purified in a Biorobot EZ1 device. The nuclear DNA of the samples was quantified with the PowerQuant kit. Because the recommended posterior probability (PP) of 99.9% was followed with the goal of high confidence of correct identification, the NGS STR Panel was used, and after the analysis of additional STR loci the statistical calculation showed a PP of 99.99986%, showing that a large enough number of genetic markers were analysed when identifying the skeletal remains of the aunt. PP value endorsed the hypothesis that the tooth and bone samples were from individual related to the family references rather than from unrelated individual. In presented case, NGS technology proved to be a powerful tool for increasing the number of autosomal STRs needed for identification of WWII victims when linear markers cannot be used for comparison and only distant relatives are available for analyses.

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“…The laboratory procedure consisted of three stages: preparation of bone samples, isolation of DNA and determination of DNA quantity. Cleaning, grinding, decalcification, and purification of DNA were performed following Zupanič Pajnič [7]. Briefly, the bones were cleaned, both, mechanically and chemically and dried over night.…”

Section: Methodsmentioning

confidence: 99%

Comparison of DNA yield after long-term storage of Second World War bone samples

Friš

Grdina

Podovšovnik

et al. 2019

Forensic Science International: Genetics Supplement Series

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Sample storage is of paramount importance in forensic genetics laboratories since only optimal storage enables successful recovery of DNA from old bones that contain very low amount of severely degraded DNA. When identification of missing persons from skeletal remains is completed, bone sample is routinely stored at -20°C for long-term storage for retesting in future, if necessary. After molecular genetic analyses of Slovenian Second World War (WWII) victims, small fragments of femurs were stored at -20°C. Reduction in DNA recovery has been observed in frozen liquid DNA extracts by some authors and the goal of our study was to explore how freezing of bone samples affects the preservation of DNA. To achieve this goal, the difference in DNA yield in extracts obtained from WWII bones analyzed in 2009 (data from published paper) and DNA yield in extracts obtained from the same bones (piece sampled next to the one used in 2009) taken out of the freezer after long-term storage on -20°C for 10 years was examined, using the same extraction method and the same quantification kit. Up to 100 ng DNA/g of bone powder was obtained from 57 WWII femurs and up to 31 ng DNA/g of bone powder from the same femurs investigated after long-term storage in this study. 0,5 g of bone powder was decalcified using full demineralization extraction method. The DNA was purified in a Biorobot EZ1 device (Qiagen) and DNA quantity determined with the Human Quantifiler kit (TFS). Statistical analysis showed significant difference in DNA yield in extracts obtained from WWII bones in 2009 and extracts obtained from the same bones stored at -20°C after 10 years. As reported for frozen liquid DNA extracts, reduction in recovery of DNA was confirmed for frozen bone samples as well.

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Extraction of DNA from Human Skeletal Material

Cited by 51 publications

References 60 publications

Identification of human remains using Rapid DNA analysis

Identification of human remains using Rapid DNA analysis

Next generation sequencing technology in Second World War victim identification

Comparison of DNA yield after long-term storage of Second World War bone samples

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