Title

Approaches to mitochondrial genome sequencing using the oxford nanopore minion device

Date of Award

5-1-2017

Degree Type

Thesis

Degree Name

Master of Science

Field of Study

Forensic Genetics

Department

Graduate School of Biomedical Sciences

First Advisor

John V. Planz

Second Advisor

Rance E. Berg

Third Advisor

J. Thomas Cunningham

Abstract

Current DNA sequencing methods rely on polymerase chain reaction (PCR) to create sufficient copies of targeted DNA fragments to serve as a library. PCR and subsequent clean-up steps add considerable time and cost to the process and provide opportunity for introduction of amplification errors or contamination. The aim of this study was to develop a reliable method for mitochondrial genome sequencing sans PCR using the Oxford Nanopore MinION. Our hypothesis was that results generated from native DNA sequencing would be concordant to sequencing results generated from PCR-enriched libraries. Error-rates of 1.47% (enriched) and 1.26% (native) were observed when sequencing the control DNA. Consensus sequences generated from native and PCR-enriched libraries show an average of 98.7% identity between the two methods.

Comments

Available May 2018.

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