Characterization of NIST Human Mitochondrial DNA SRM-2392 and SRM-2392I Standard Reference Materials by Next Generation Sequencing

Published

July 1, 2017

Author(s)

Peter M. Vallone, Kevin M. Kiesler, Sarah Riman, Lisa A. Borsuk

Abstract

Standard Reference Materials SRM 2392 and 2392-I are intended to provide quality control when amplifying and sequencing human mitochondrial genome sequences. The National Institute of Standards and Technology (NIST) offers these SRMs to laboratories performing DNA-based forensic human identification, molecular diagnosis of mitochondrial diseases, or mutation detection. The entire mtGenome (~16,569 bp) of SRM 2392 and 2392-I have previously been characterized at NIST by Sanger sequencing. Herein, we used the sensitivity, specificity, and accuracy offered by next generation sequencing (NGS) to: (1) re-sequence the certified values of the SRM 2392 and 2392-I; (2) confirm Sanger data with a high coverage new sequencing technology; (3) detect lower level heteroplasmies (<20%), and thus (4) support adoption of NGS in forensic community. To obtain a consensus sequence for the SRMs as well as identify and control any bias, sequencing was performed using two NGS platforms and data was analyzed using different bioinformatics pipelines. Our results confirm five low level heteroplasmy sites that were not previously observed with Sanger sequencing: three sites in the GM09947A template in SRM 2392 and two sites in the HL-60 template in SRM 2392-I.

Citation

Forensic Science International: Genetics

Volume

29

Pub Type

Journals

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DOI Link

Keywords

Standard Reference Material (SRM), Next generation sequencing, Mitochondrial DNA, Heteroplasmy

Citation

Vallone, P. , Kiesler, K. , Riman, S. and Borsuk, L. (2017), Characterization of NIST Human Mitochondrial DNA SRM-2392 and SRM-2392I Standard Reference Materials by Next Generation Sequencing, Forensic Science International: Genetics, [online], https://doi.org/10.1016/j.fsigen.2017.04.005 (Accessed October 9, 2025)