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Description
Many factors are at play within the genome of an organism, contributing to much of the diversity and variation across the tree of life. While the genome is generally encoded by four nucleotides, A, C, T, and G, this code can be expanded. One particular mechanism that we examine in this thesis is modification of bases—more specifically, methylation of Adenine (m6A) within the GATC motif of Escherichia coli. These methylated adenines are especially important in a process called methyl-directed mismatch repair (MMR), a pathway responsible for repairing errors in the DNA sequence produced by replication. In this pathway, methylated adenines identify the parent strand and direct the repair proteins to correct the erroneous base in the daughter strand. While the primary role of methylated adenines at GATC sites is to direct the MMR pathway, this methylation has also been found to affect other processes, such as gene expression, the activity of transposable elements, and the timing of DNA replication. However, in the absence of MMR, the ability of these other processes to maintain adenine methylation and its targets is unknown.
To determine if the disruption of the MMR pathway results in the reduced conservation of methylated adenines as well as an increased tolerance for mutations that result in the loss or gain of new GATC sites, we surveyed individual clones isolated from experimentally evolving wild-type and MMR-deficient (mutL- ;conferring an 150x increase in mutation rate) populations of E. coli with whole-genome sequencing. Initial analysis revealed a lack of mutations affecting methylation sites (GATC tetranucleotides) in wild-type clones. However, the inherent low mutation rates conferred by the wild-type background render this result inconclusive, due to a lack of statistical power, and reveal a need for a more direct measure of changes in methylation status. Thus as a first step to comparative methylomics, we benchmarked four different methylation-calling pipelines on three biological replicates of the wildtype progenitor strain for our evolved populations.
While it is understood that these methylated sites play a role in the MMR pathway, it is not fully understood the full extent of their effect on the genome. Thus the goal of this thesis was to better understand the forces which maintain the genome, specifically concerning m6A within the GATC motif.
To determine if the disruption of the MMR pathway results in the reduced conservation of methylated adenines as well as an increased tolerance for mutations that result in the loss or gain of new GATC sites, we surveyed individual clones isolated from experimentally evolving wild-type and MMR-deficient (mutL- ;conferring an 150x increase in mutation rate) populations of E. coli with whole-genome sequencing. Initial analysis revealed a lack of mutations affecting methylation sites (GATC tetranucleotides) in wild-type clones. However, the inherent low mutation rates conferred by the wild-type background render this result inconclusive, due to a lack of statistical power, and reveal a need for a more direct measure of changes in methylation status. Thus as a first step to comparative methylomics, we benchmarked four different methylation-calling pipelines on three biological replicates of the wildtype progenitor strain for our evolved populations.
While it is understood that these methylated sites play a role in the MMR pathway, it is not fully understood the full extent of their effect on the genome. Thus the goal of this thesis was to better understand the forces which maintain the genome, specifically concerning m6A within the GATC motif.
ContributorsBoyer, Gwyneth (Author) / Lynch, Michael (Thesis director) / Behringer, Megan (Committee member) / Geiler-Samerotte, Kerry (Committee member) / School of Life Sciences (Contributor) / Department of Psychology (Contributor) / Barrett, The Honors College (Contributor)
Created2020-05
DescriptionAn auto-ethnographic overview of the author's personal experiences in professional and educational stage management in Arizona. Provides a critique of ASU's stage management program and offers solutions to improve the sustainability and safety of student stage managers.
ContributorsMattson, Jordan Clare (Author) / Hunt, Kristin (Thesis director) / Kirkham, William (Committee member) / School of Music, Dance and Theatre (Contributor) / Department of Psychology (Contributor) / Barrett, The Honors College (Contributor)
Created2020-12
Description
Arachnids belong to the phylum Arthropoda, the largest phylum in the animal kingdom. Ticks are blood-feeding arachnids that vector numerous pathogens of significant medical and veterinary importance, while scorpions have become a common concern in urban desert cities due to the high level of toxicity in their venom. To date, viruses associated with arachnids have been under sampled and understudied. Here viral metagenomics was used to explore the diversity of viruses present in ticks and scorpions. American dog ticks (Dermacentor variabilis) and blacklegged ticks (Ixodes scapularis) were collected in Pennsylvania while one hairy scorpion (Hadrurus arizonensis) and four bark scorpions (Centruroides sculpturatus) were collected in Phoenix. Novel viral genomes described here belong to the families Polyomaviridae, Anelloviridae, Genomoviridae, and a newly proposed family, Arthropolviridae.
Polyomaviruses are non-enveloped viruses with a small, circular double-stranded DNA (dsDNA) genomes that have been identified in a variety of mammals, birds and fish and are known to cause various diseases. Arthropolviridae is a proposed family of circular, large tumor antigen encoding dsDNA viruses that have a unidirectional genome organization. Genomoviruses and anelloviruses are ssDNA viruses that have circular genomes ranging in size from 2–2.4 kb and 2.1–3.8 kb, respectively. Genomoviruses are ubiquitous in the environment, having been identified in a wide range of animal, plant and environmental samples, while anelloviruses have been associated with a plethora of animals.
Here, 16 novel viruses are reported that span four viral families. Eight novel polyomaviruses were recovered from bark scorpions, three arthropolviruses were recovered from dog ticks and one arthropolvirus from a hairy scorpion. Viruses belonging to the families Polyomaviridae and Arthropolviridae are highly divergent. This is the first more extensive study of these viruses in arachnids. Three genomoviruses were recovered from both dog and deer ticks and one anellovirus was recovered from deer ticks, which are the first records of these viruses being recovered from ticks. This work highlights the diversity of dsDNA and ssDNA viruses in the arachnid population and emphasizes the importance of performing viral surveys on these populations.
Polyomaviruses are non-enveloped viruses with a small, circular double-stranded DNA (dsDNA) genomes that have been identified in a variety of mammals, birds and fish and are known to cause various diseases. Arthropolviridae is a proposed family of circular, large tumor antigen encoding dsDNA viruses that have a unidirectional genome organization. Genomoviruses and anelloviruses are ssDNA viruses that have circular genomes ranging in size from 2–2.4 kb and 2.1–3.8 kb, respectively. Genomoviruses are ubiquitous in the environment, having been identified in a wide range of animal, plant and environmental samples, while anelloviruses have been associated with a plethora of animals.
Here, 16 novel viruses are reported that span four viral families. Eight novel polyomaviruses were recovered from bark scorpions, three arthropolviruses were recovered from dog ticks and one arthropolvirus from a hairy scorpion. Viruses belonging to the families Polyomaviridae and Arthropolviridae are highly divergent. This is the first more extensive study of these viruses in arachnids. Three genomoviruses were recovered from both dog and deer ticks and one anellovirus was recovered from deer ticks, which are the first records of these viruses being recovered from ticks. This work highlights the diversity of dsDNA and ssDNA viruses in the arachnid population and emphasizes the importance of performing viral surveys on these populations.
ContributorsSchmidlin, Kara (Author) / Varsani, Arvind (Thesis advisor) / Van Doorslaer, Koenraad (Committee member) / Stenglein, Mark (Committee member) / Arizona State University (Publisher)
Created2019