Armed Forces DNA Identification Laboratory

Dover, DE, United States

Armed Forces DNA Identification Laboratory

Dover, DE, United States
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Reedy C.R.,University of Virginia | Bienvenue J.M.,Armed Forces DNA Identification Laboratory | Coletta L.,University of Virginia | Strachan B.C.,University of Virginia | And 2 more authors.
Forensic Science International: Genetics | Year: 2010

Microdevices are often designed to process sample volumes on the order of tens of microliters and cannot typically accommodate larger volume samples without adversely affecting efficiency and greatly increasing analysis time. However, dilute, large-volume biological samples are frequently encountered, especially in forensic or clinical laboratories. A microdevice, capable of efficiently processing 0.5-1 mL samples has been developed for solid phase extraction (SPE) of DNA. SPE was carried out on a microdevice utilizing magnetic silica particles and an optimized volumetric flow rate and elution buffer, resulting in a 50-fold decrease in volume and a 15-fold increase in DNA concentration. Device characterization studies showed DNA extraction efficiencies comparable with previously reported silica-based purification methods, with robust performance demonstrated by the successful amplification of a fragment from the gelsolin gene extracted from dilute whole blood. In addition, the microchip-based method for SPE of large volume, dilute samples was also used to demonstrate the first successful on-chip purification of mitochondrial DNA (mtDNA) from both dilute whole blood and a degraded blood stain. © 2009 Elsevier Ireland Ltd.

Lyons E.A.,American Registry of Pathology | Lyons E.A.,Armed Forces DNA Identification Laboratory | Scheible M.K.,American Registry of Pathology | Scheible M.K.,Armed Forces DNA Identification Laboratory | And 7 more authors.
BMC Genomics | Year: 2013

Background: A population reference database of complete human mitochondrial genome (mtGenome) sequences is needed to enable the use of mitochondrial DNA (mtDNA) coding region data in forensic casework applications. However, the development of entire mtGenome haplotypes to forensic data quality standards is difficult and laborious. A Sanger-based amplification and sequencing strategy that is designed for automated processing, yet routinely produces high quality sequences, is needed to facilitate high-volume production of these mtGenome data sets. Results: We developed a robust 8-amplicon Sanger sequencing strategy that regularly produces complete, forensic-quality mtGenome haplotypes in the first pass of data generation. The protocol works equally well on samples representing diverse mtDNA haplogroups and DNA input quantities ranging from 50 pg to 1 ng, and can be applied to specimens of varying DNA quality. The complete workflow was specifically designed for implementation on robotic instrumentation, which increases throughput and reduces both the opportunities for error inherent to manual processing and the cost of generating full mtGenome sequences. Conclusions: The described strategy will assist efforts to generate complete mtGenome haplotypes which meet the highest data quality expectations for forensic genetic and other applications. Additionally, high-quality data produced using this protocol can be used to assess mtDNA data developed using newer technologies and chemistries. Further, the amplification strategy can be used to enrich for mtDNA as a first step in sample preparation for targeted next-generation sequencing. © 2013 Lyons et al.; licensee BioMed Central Ltd.

Tillmar A.O.,National Board of Forensic Medicine | Coble M.D.,Armed Forces DNA Identification Laboratory | Wallerstrom T.,Norwegian University of Science and Technology | Holmlund G.,National Board of Forensic Medicine
International Journal of Legal Medicine | Year: 2010

In order to promote mitochondrial DNA (mtDNA) testing in Sweden we have typed 296 Swedish males, which will serve as a Swedish mtDNA frequency database. The tested males were taken from seven geographically different regions representing the contemporary Swedish population. The complete mtDNA control region was typed and the Swedish population was shown to have high haplotype diversity with a random match probability of 0.5%. Almost 47% of the tested samples belonged to haplogroup H and further haplogroup comparison with worldwide populations clustered the Swedish mtDNA data together with other European populations. AMOVA analysis of the seven Swedish subregions displayed no significant maternal substructure in Sweden (F ST∈=∈0. 002). Our conclusion from this study is that the typed Swedish individuals serve as good representatives for a Swedish forensic mtDNA database. Some caution should, however, be taken for individuals from the northernmost part of Sweden (provinces of Norrbotten and Lapland) due to specific demographic conditions. Furthermore, our analysis of a small sample set of a Swedish Saami population confirmed earlier findings that the Swedish Saami population is an outlier among European populations. © 2009 Springer-Verlag.

Loreille O.,Armed Forces DNA Identification Laboratory | Koshinsky H.,Eureka Genomics Corporation | Fofanov V.Y.,Eureka Genomics Corporation | Irwin J.A.,Armed Forces DNA Identification Laboratory
Forensic Science International: Genetics Supplement Series | Year: 2011

Complete genome studies performed with next generation sequencing technologies are becoming more and more abundant. The potential such technologies could have for DNA identification purposes are obvious, but so far, very few forensic laboratories have tested these new instruments. We decided to evaluate the Illumina GAIIx platform for sequencing mitochondrial DNA extracted from an ancient human bone. We show that using standard Illumina protocols, we obtained a very small coverage of the mtDNA genome and observe a high error rate of 1.44%. We therefore tested various methods to improve the quality and quantity of the data. This report will describe our results when we incorporated a DNA repair and a primer extension capture step in the protocol. © 2011 .

Scheible M.,Armed Forces DNA Identification Laboratory | Kim S.H.,Gangneung - Wonju National University | Sturk-Andreaggi K.,Armed Forces DNA Identification Laboratory | Coble M.D.,Armed Forces DNA Identification Laboratory | And 3 more authors.
International Journal of Legal Medicine | Year: 2014

Mitochondrial DNA (mtDNA) control region (16024-576) sequences were generated from 281 individuals from South Korea. Robotic liquid handling, a redundant sequencing strategy, and a series of quality control checks were implemented to ensure the high quality of the dataset. This population sample showed a low random match probability (0.25 %) and high genetic diversity (0.9933). The haplogroup breakdown was consistent with previous studies describing Korean mtDNA variation. The 224 unique haplotypes (33 shared) presented will supplement the data already publically available. © 2014 Springer-Verlag Berlin Heidelberg (outside the USA).

Edson S.M.,Armed Forces DNA Identification Laboratory | Christensen A.F.,POW Inc
Journal of Forensic Sciences | Year: 2013

The Armed Forces DNA Identification Laboratory reports the mitochondrial DNA (mtDNA) sequences of over 800 skeletal samples a year for the Joint POW/MIA Accounting Command-Central Identification Laboratory. These sequences are generated from degraded skeletal remains that are presumed to belong to U.S. service members missing from past military conflicts. In the laboratory, it is possible to control for contamination of remains; however, in the field, it can be difficult to prevent modern DNA from being transferred to skeletal elements and being carried forward through the analysis process. Four such cases are described here along with the controls in place in the laboratory to eliminate the possibility of the exogenous DNA being reported as authentic. In each case, the controls implemented by the laboratories prevented the false reporting of contaminant exogenous DNA from remains that were either faunal or human, but lacked endogenous DNA. © 2012.

Scheible M.,Armed Forces DNA Identification Laboratory | Loreille O.,Armed Forces DNA Identification Laboratory | Just R.,Armed Forces DNA Identification Laboratory | Irwin J.,Armed Forces DNA Identification Laboratory
Forensic Science International: Genetics Supplement Series | Year: 2011

To investigate the feasibility of next generation sequencing (NGS) technology for the multiplex detection and sequence production of short tandem repeats (STRs), thirteen STR markers (CSF1PO, D2S441, D3S1358, D5S818, D7S820, D8S1179, D10S1248, D13S317, D16S539, D21S11, D22S1045, TPOX, and vWA) were amplified using an optimized multiplex reaction with primer sequences designed for reduced size amplicons. Each sample multiplex was barcoded with a different sample-specific multiplex identifier (MID) for subsequent parallel tagged sequencing on the GS Junior System (454 Life Sciences, Branford, CT). © 2011.

Hebda L.M.,Michigan State University | Hebda L.M.,Armed Forces DNA Identification Laboratory | Foran D.R.,Michigan State University
Journal of Forensic Sciences | Year: 2015

DNA identification of human remains is often necessary when decedents are skeletonized; however, poor DNA recovery and polymerase chain reaction (PCR) inhibition are frequently encountered, a situation exacerbated by burial. In this research, the utility of integrating soil DNA isolation kits into buried skeletal DNA analysis was evaluated and compared to a standard human DNA extraction kit and organic extraction. The soil kits successfully extracted skeletal DNA at quantities similar to standard methods, although the two kits tested, which differ mechanistically, were not equivalent. Further, the PCR inhibitors calcium and humic acid were effectively removed using the soil kits, whereas collagen was less so. Finally, concordant control region sequences were obtained from human skeletal remains using all four methods. Based on these comparisons, soil DNA isolation kits, which quickened the extraction process, proved to be a viable extraction technique for skeletal remains that resulted in positive identification of a decedent. © 2015 American Academy of Forensic Sciences.

Scheible M.,American Registry of Pathology | Scheible M.,Armed Forces DNA Identification Laboratory | Loreille O.,American Registry of Pathology | Loreille O.,Armed Forces DNA Identification Laboratory | And 4 more authors.
Forensic Science International: Genetics | Year: 2014

To investigate the feasibility of next generation sequencing technology (NGS) for the multiplex detection and sequence production of short tandem repeats (STRs) from degraded and low DNA quantity samples, standard polymerase chain reaction amplification methods were used to enrich for commonly employed STR markers. Samples were amplified with two multiplexing strategies: a multiplex containing thirteen miniSTR markers and a series of multiplexes containing four miniSTR markers each. Each sample multiplex was barcoded with a sample-specific multiplex identifier for subsequent parallel tagged sequencing on the GS Junior System (454 Life Sciences, a Roche company, Branford, CT). Sequencing results from over fifty DNA extracts representing both pristine samples and low-quality evidentiary specimens reflected known genotypes and were consistent across multiple extracts and/or amplifications of the same sample. Furthermore, the NGS data revealed sequence information not available with standard capillary electrophoresis-based detection alone. For the population samples tested, a total of 152 single nucleotide polymorphisms or insertions/deletions were identified in over 935 recovered alleles, averaging one polymorphism for every six recovered alleles. For three of the loci, the sequence information doubled the number of alleles detected via traditional STR typing by fragment analysis. In addition, twenty-eight of these variants were only seen once within our dataset, highlighting the potential for discrimination among individuals. These additional data are likely to be particularly valuable in missing persons and disaster victim identification cases for which only partial profiles may be recovered and/or only distant kin are available as references. And, considering the opportunity to target only small amplicons with NGS, this type of STR typing will allow for greater information recovery from challenging casework samples. While our results highlight the potential of new technologies for recovering discriminatory genetic information from evidentiary specimens, our data also reveal the complexities of NGS-based STR typing, both in terms of the laboratory assays themselves as well as the downstream data processing and analysis. © 2014 Elsevier Ireland Ltd.

Just R.S.,Armed Forces DNA Identification Laboratory | Irwin J.A.,FBI Laboratory | Parson W.,Innsbruck Medical University | Parson W.,Pennsylvania State University
Forensic Science International: Genetics | Year: 2015

Abstract Long an important and useful tool in forensic genetic investigations, mitochondrial DNA (mtDNA) typing continues to mature. Research in the last few years has demonstrated both that data from the entire molecule will have practical benefits in forensic DNA casework, and that massively parallel sequencing (MPS) methods will make full mitochondrial genome (mtGenome) sequencing of forensic specimens feasible and cost-effective. A spate of recent studies has employed these new technologies to assess intraindividual mtDNA variation. However, in several instances, contamination and other sources of mixed mtDNA data have been erroneously identified as heteroplasmy. Well vetted mtGenome datasets based on both Sanger and MPS sequences have found authentic point heteroplasmy in approximately 25% of individuals when minor component detection thresholds are in the range of 10-20%, along with positional distribution patterns in the coding region that differ from patterns of point heteroplasmy in the well-studied control region. A few recent studies that examined very low-level heteroplasmy are concordant with these observations when the data are examined at a common level of resolution. In this review we provide an overview of considerations related to the use of MPS technologies to detect mtDNA heteroplasmy. In addition, we examine published reports on point heteroplasmy to characterize features of the data that will assist in the evaluation of future mtGenome data developed by any typing method. © 2015 The Authors.

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