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Assessing the Effects of DNA Damage on Next Generation mtDNA Sequencing

NCJ Number
252862
Author(s)
Mitchell M. Holland
Date Published
September 2018
Length
14 pages
Annotation
This study assessed the impact of DNA damage on the interpretation of massively parallel sequencing mitochondrial (mt) DNA heteroplasmy from a variety of perspectives.
Abstract
The first part of the study focused on methods development. This involved a mtDNA qPCR assay to assess both quantity and quality of mtDNA; amplification schemes for production of mtDNA products for massively parallel sequencing (MPS) analysis; and bioinorganic pipelines and software development for analysis of MPS mtDNA data, along with initial considerations of DNA damage. These achievements were critical for completion of the project and resulted in multiple publications. The second part of the study focused on the evaluation of various approaches to modeling deamination, depurination, and oxidation-based DNA damage, including whether approaches of biological repair can reverse the damage. These studies have led to targeted experiments on mock-evidence-type samples. The low template study provided a valuable perspective on the influence of low template samples when considering the impact of DNA damage on the interpretation of mtDNA MPS data. In addition, the findings confirmed previous observation that as the amplicon size being evaluated for damage increases, the amount of damage decreases. This is an important outcome, since most of the studies in forensic laboratories involve the analysis of smaller amplification products. Final data analysis has been completed, and a manuscript is being prepared. 8-item bibliography