Development of a Forensic Quality Mitogenome Haplotype Database from 5000 High Quality Samples Using Next-Generation Sequencing Methods

The forensics community has long been cognizant of the enhanced discriminatory value of mitochondrial genome (mitogenome) analysis compared to that of the commonly interrogated control region (CR). Although historically met with hurdles due to sequencing impracticality, the introduction of next generation sequencing (NGS) has now provided the forensic community with the capacity to produce reliable mitogenome data from forensic samples. However, an assessment of statistical significance is required to give weight to genetic evidence in a forensic context. To date, publicly available databases focusing only on the CR are inadequate for estimating mitogenome haplotype frequencies and match probabilities because the coding region data are not available for consideration. Thus the limitation to utilizing mitogenome sequencing data has shifted from the ability to produce the data to the availability of appropriate databases for comparison. To address this issue and support the use of the entire sequence range for forensic applications, a database that contains full mitogenome data is required.

This proposal seeks to provide a representative, public database of 5,000 mitogenome haplotypes from individuals of diverse maternal ancestry, making mitogenome sequencing in forensic DNA laboratories within the United States and worldwide a viable reality. This databasing project will utilize high-quality, single-source samples from anonymized donors representing the five major United States racial population groups, as well as a sampling of several global populations. NGS data will be generated for the mitogenome using a validated method of long range PCR, dual-indexed library preparation, and Illumina sequencing. A bioinformatic data analysis pipeline developed specifically for forensic mitogenome analysis, in part by scientists at the AFDIL, will be utilized to produce accurate haplotypes. All procedures involve the application of rigorous quality control procedures to affirm the integrity of the data obtained. A partnership with Prof. Walther Parson (Institute of Legal Medicine, Innsbruck Medical University) will facilitate external review of the data and submission to EMPOP. Access to a large high-quality mitogenome database will enable statistical inferences to be drawn from probative mitogenome data. Presentations at national and international conferences as well as peer-reviewed publications are planned to reach the widest possible audience of forensic practitioners. Interpretation guidelines for mitogenome profile reporting and comparison will be established from the data and disseminated to the forensics community.