Date of Award

Summer 7-30-2024

Document Type

Thesis

Degree Name

Master of Science (MS)

Department

Biology

First Advisor

Dr. Arnab Sengupta

Second Advisor

Dr. Bruce Snyder

Abstract

Despite the vast diversity of millipedes, complete genomic and mitogenomic data available in public databases is limited, hindering comprehensive studies on their classification, evolution, and genetics. Among the underrepresented taxa are the Xystodesmidae, a family comprising approximately 500 species with minimal mitogenomic data, and no complete genomes sequenced. Our research involves sequencing the mitochondrial genome of Cherokia georgiana, a representative of this diverse family with interesting traits that are not clearly defined at a genetic level. This research also serves as a pilot study to develop DNA extraction methods and protocols for whole genome sequencing. Multiple DNA extraction kits and techniques were evaluated to optimize the yield of high-quality, high molecular weight (HMW) DNA suitable for sequencing. We assess the quality of next-generation sequencing (NGS) libraries using spectrophotometry (ThermoFisher Scientific Nanodrop), fluorometry (Invitrogen Qubit 4.0), and automated electrophoresis (Agilent TapeStation 4200). This work outlines the research strategies and protocols employing the Oxford Nanopore Technologies (ONT) MinION Mk1B platform for long read sequencing. We employed Dotmatics Geneious Prime software (Version 2023.2.1) and used the reference species data from Appalachioria falcifera to assemble the mitogenome. We generated a mitogenome map of C. georgiana, analyzed predicted structures of mitochondrial tRNA, and conducted basic phylogenetic analysis using the cytochrome c oxidase subunit I (COX1) genes and full mitogenomes published to GenBank. Future experiments will use the extraction methods described in this paper and will apply the PacBIO platform for whole genome sequencing to further enhance our knowledge of millipede genomics.

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