Validation of a mitochondrial extraction and isolation technique for use in wildlife forensics.

Abstract

The primary goal of this project is to provide the wildlife forensic science community with a DNA analysis protocol that is consistent across all jurisdictions. To accomplish this goal, three important objectives must be achieved: 1) validation of a mitochondrial DNA (mtDNA) specific extraction, purification, and sequencing method that is robust no matter the species, 2) creation of an internet web interface for the submission and analysis of sequence data from casework samples for data analysis, and 3) creation and curation of a database of mitochondrial reference genomes that are forensically-vouchered. This thesis aims to accomplish the first objective. Current protocols within the field of forensic wildlife DNA analysis require a complete redesign and validation of protocols every time a new species is encountered during casework. This practice increases the turnaround time of casework dramatically, causes challenges when testifying to findings, and relies on data from non-forensic scientists that may not be at the same high standard required for forensic applications. My research focuses on developing a mitochondrial extraction method that is free of nuclear DNA contamination and absent of primer usage and PCR. I therefore offer to the wildlife forensic community the first total mtDNA sequencing method that can be used on any animal species and is independent of prior genetic knowledge of the organism in question. The project seeks to aid the wildlife forensic science community by providing a streamlined method for the analysis of DNA which is both quicker and more robust..