Developmental Validation of a MPS Workflow with a PCR-Based Short Amplicon Whole Mitochondrial Genome Panel

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dc.creatorCihlar, Jennifer Churchill
dc.creatorAmory, Christina
dc.creatorLagace, Robert
dc.creatorRoth, Chantal
dc.creatorParson, Walther
dc.creatorBudowle, Bruce
dc.creator.orcid0000-0001-9796-193X (Cihlar, Jennifer Churchill)
dc.date.accessioned2022-08-23T19:18:56Z
dc.date.available2022-08-23T19:18:56Z
dc.date.issued2020-11-13
dc.description.abstractFor the adoption of massively parallel sequencing (MPS) systems by forensic laboratories, validation studies on specific workflows are needed to support the feasibility of implementation and the reliability of the data they produce. As such, the whole mitochondrial genome sequencing methodology-Precision ID mtDNA Whole Genome Panel, Ion Chef, Ion S5, and Converge-has been subjected to a variety of developmental validation studies. These validation studies were completed in accordance with the Scientific Working Group on DNA Analysis Methods (SWGDAM) validation guidelines and assessed reproducibility, repeatability, accuracy, sensitivity, specificity to human DNA, and ability to analyze challenging (e.g., mixed, degraded, or low quantity) samples. Intra- and inter-run replicates produced an average maximum pairwise difference in variant frequency of 1.2%. Concordance with data generated with traditional Sanger sequencing and an orthogonal MPS platform methodology was used to assess accuracy, and generation of complete and concordant haplotypes at DNA input levels as low as 37.5 pg of nuclear DNA or 187.5 mitochondrial genome copies illustrated the sensitivity of the system. Overall, data presented herein demonstrate that highly accurate and reproducible results were generated for a variety of sample qualities and quantities, supporting the reliability of this specific whole genome mitochondrial DNA MPS system for analysis of forensic biological evidence.
dc.description.sponsorshipThermo Fisher Scientific provided reagents necessary to perform this study. This work was supported in part by the European Union, grant agreement number 779485-STEFA-ISFP-2016-AG-IBA-ENFSI and the National Institute of Justice, Office of Justice Programs, U.S. Department of Justice, grant number 2016-DN-BX-K001. The opinions, findings, and conclusions or recommendations expressed in this publication are those of the authors and do not necessarily reflect those of the U.S. Department of Justice.
dc.identifier.citationCihlar, J. C., Amory, C., Lagacé, R., Roth, C., Parson, W., & Budowle, B. (2020). Developmental Validation of a MPS Workflow with a PCR-Based Short Amplicon Whole Mitochondrial Genome Panel. Genes, 11(11), 1345. https://doi.org/10.3390/genes11111345
dc.identifier.issn2073-4425
dc.identifier.issue11
dc.identifier.urihttp://hdl.handle.net/20.500.12503/31623
dc.identifier.volume11
dc.publisherMDPI
dc.relation.urihttps://doi.org/10.3390/genes11111345
dc.rights.holder© 2020 by the authors.
dc.rights.licenseAttribution 4.0 International (CC BY 4.0)
dc.rights.urihttp://creativecommons.org/licenses/by/4.0/
dc.sourceGenes (Basel)
dc.subjectIon Torrent
dc.subjectSWGDAM guidelines
dc.subjectdevelopmental validation
dc.subjectmassively parallel sequencing
dc.subjectmitochondrial DNA
dc.subject.meshAnimals
dc.subject.meshComputational Biology
dc.subject.meshForensic Genetics / methods
dc.subject.meshGenome, Mitochondrial
dc.subject.meshHair
dc.subject.meshHaplotypes
dc.subject.meshHigh-Throughput Nucleotide Sequencing / methods
dc.subject.meshHumans
dc.subject.meshMale
dc.subject.meshPolymerase Chain Reaction / methods
dc.subject.meshSaliva
dc.subject.meshSemen
dc.subject.meshSensitivity and Specificity
dc.subject.meshSpecies Specificity
dc.subject.meshWorkflow
dc.titleDevelopmental Validation of a MPS Workflow with a PCR-Based Short Amplicon Whole Mitochondrial Genome Panel
dc.typeArticle
dc.type.materialtext

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