Dissertation/Thesis Abstract

Low-level variant detection in human mitochondrial DNA using the Illumina MiSeq next-generation sequencing (NGS) platform
by Smith, Brandon Chase, M.S., Western Carolina University, 2013, 84; 1537163
Abstract (Summary)

When challenged by difficult biological samples, the forensic analyst is far more likely to obtain useful data by sequencing the human mitochondrial DNA (mtDNA). Nextgeneration sequencing (NGS) technologies are currently being evaluated by the Forensic Science Program at Western Carolina University for their ability to reliably detect lowlevel variants in mixtures of mtDNA. The sequence profiles for twenty individuals were obtained by sequencing amplified DNA derived from the mitochondrial hypervariable (HV) regions using Sanger methods. Two-person mixtures were then constructed by mixing quantified templates, simulating heteroplasmy at discrete sites and in defined ratios. Libraries of unmixed samples, artificial mixtures, and instrument controls were prepared using Illumina® Nextera® XT and deep-sequenced on the Illumina®MiSeq™. Analysis of NGS data using a novel bioinformatics pipeline indicated that minor variants could be detected at the 5, 2, 1, and 0.5% levels of detection. Additional experiments which examined the occurrence of sequence variation in hair tissue demonstrates that a considerable amount of sequence variation can exist between hairs and other tissues derived from a single donor.

Indexing (document details)
Advisor: Wilson, Mark R.
Commitee: Bose, Indrani, Summers, Jack
School: Western Carolina University
Department: Biology
School Location: United States -- North Carolina
Source: MAI 51/06M(E), Masters Abstracts International
Source Type: DISSERTATION
Subjects: Genetics, Bioinformatics
Keywords: Forensic science, Low-level mixtures, Minor variant, Mitochondrial DNA, Next-generation sequencing
Publication Number: 1537163
ISBN: 978-1-303-07196-6
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