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

Date

2020-11-13

Authors

Cihlar, Jennifer Churchill
Amory, Christina
Lagace, Robert
Roth, Chantal
Parson, Walther
Budowle, Bruce

ORCID

0000-0001-9796-193X (Cihlar, Jennifer Churchill)

Journal Title

Journal ISSN

Volume Title

Publisher

MDPI

Abstract

For 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.

Description

Citation

Cihlar, 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

Rights

© 2020 by the authors.

License

Attribution 4.0 International (CC BY 4.0)