Rodriguez, DavidMolis, Toriann Ayzlynn2023-05-152023-05-152022-05Molis, T. A. (2022). Nanopore sequencing of the Cachabi robber frog (Pristimantis achatinus) mitochondrial genome. Honors College, Texas State University.https://hdl.handle.net/10877/16784Due to the devastating impacts of chytridiomycosis caused by Batrachochytrium dendrobatidis on tropical amphibians, it is vital to understand their host-pathogen dynamics. These studies can be better informed by measuring genetic diversity in the hosts, especially where cryptic diversity is present, such as in the tropics. Measuring diversity can now be performed in situ with portable nanopore DNA sequencing technology. To facilitate this, universal PCR primers have been designed to generate mitochondrial DNA (mtDNA) fragments of ~3400 base pairs (bp) when applied to most amphibians; however, in Pristimantis achatinus, they only generate fragments of ~900 bp, likely due to a rearrangement in its mitogenome. To investigate the cause of the decreased amplicon size, we performed a series of tests using nanopore sequencing, wherein we attempted to sequence the entire mitochondrial genome of P. achatinus. Different methods of mtDNA extraction were tested as well as different means of whole genome amplification. The sequence data obtained from these nanopore runs will provide necessary insights into the genetic diversity of these amphibians, while the tests will help to optimize a protocol for nanopore sequencing of whole mitochondrial genomes of amphibians, furthering our ability to uncover genetic diversity and understand the dynamics between fungal pathogens and their amphibian hosts.Text44 pages1 file (.pdf)encryptic diversitywhole mitogenome sequencingEcuadorian amphibiansOxford nanopore technologieswhole genome amplificationHonors College