Matching Items (10)
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- All Subjects: Genetics
- Creators: School of Molecular Sciences
- Member of: Barrett, The Honors College Thesis/Creative Project Collection
- Member of: Theses and Dissertations
Description
This study focused on the connection between the EnvZ/OmpR two-component regulatory system and the iron homeostasis system in Escherichia coli, specifically how a mutant form of EnvZ11/OmpR is able to reduce the expression of fepA::lacZ, a reporter gene fusion in E. coli. FepA is one of several outer membrane siderophore receptors that allow extracellular siderophores bound to iron to enter the cells to power various biological processes. Previous studies have shown that in E. coli cells that expressed a mutant allele of envZ, called envZ11, which led to altered expression of various iron genes including down regulation of fepA::lacZ. The wild type EnvZ/OmpR system is not considered to regulate iron genes, but because these envz11 strains had downregulated fepA::lacZ, this study was undertaken to understand the connection and mechanisms of this downregulation. A large number of Lac+ revertants were obtained from the B32-2483 strain (envz11 and fepA::lacZ) and 7 Lac+ revertants that had reversion mutations not directly correcting the envZ11 allele were further characterized. With P1 phage transduction genetic mapping that involved moving a kanamycin resistance marker linked to fepA::lacZ, two Lac+ revertants were found to have their reversion mutations in the fepA promoter region, while the other five revertants had their mutations mapping outside the fepA region. These two revertants underwent DNA sequencing and found to carry two different single base pair mutations in two different locations of the fepA promoter region. Each one is in the Fur repressor binding region, but one also may have affected the Shine-Dalgarno region involved in translation initiation. All 7 reveratants underwent beta-galactosidase assays to measure fepA::lacZ expression. The two revertants that had mutations in the fepA promoter region had significantly increased fepA activity, with the revertant with the Shine-Dalgarno mutation having the most elevated fepA expression. The other 5 revertants that did not map in the fepA region had fepA expression elevated to the same level as that found in the wild type EnvZ/OmpR background. The data suggest that the negative effect of envZ11 can be overcome by multiple mechanisms, including directly correcting the envZ11 allele or changing the fepA promoter region.
ContributorsKalinkin, Victor Arkady (Co-author) / Misra, Rajeev (Co-author, Thesis director) / Mason, Hugh (Committee member) / Foy, Joseph (Committee member) / Biomedical Informatics Program (Contributor) / School of Life Sciences (Contributor) / W. P. Carey School of Business (Contributor) / Barrett, The Honors College (Contributor)
Created2016-05
Description
Gle1 is an mRNP export mediator with major activity localized to the nuclear pore complex in eukaryotic cells. The protein's high preservation across vast phylogenetic distances allows us to approximate research on the properties of yeast Gle1 (yGle1) with those of human Gle1 (hGle1). Research at Vanderbilt University in 2016, which provides the research basis of this thesis, suggests that the coiled-coil domain of yGle1 is best crystallized in dicationic aqueous conditions of pH ~8.0 and 10\u201420% PEG 8000. Further exploration of crystallizable microconditions revealed a favorability toward lower pH and lower PEG concentration. Following the discovery of the protein's native crystallography conditions, a comprehensive meta-analysis of scientific literature on Gle1 was conducted on the association of Gle1 mutations with neuron disease.
ContributorsGaetano, Philip Pasquale (Author) / Foy, Joseph (Thesis director) / Dawson, T. Renee (Committee member) / School of Molecular Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2016-12
Description
The goal of this project was to design and create a genetic construct that would allow for <br/>tumor growth to be induced in the center of the wing imaginal disc of Drosophila larvae, the <br/>R85E08 domain, using a heat shock. The resulting transgene would be combined with other <br/>transgenes in a single fly that would allow for simultaneous expression of the oncogene and, in <br/>the surrounding cells, other genes of interest. This system would help establish Drosophila as a <br/>more versatile and reliable model organism for cancer research. Furthermore, pilot studies were <br/>performed, using elements of the final proposed system, to determine if tumor growth is possible <br/>in the center of the disc, which oncogene produces the best results, and if oncogene expression <br/>induced later in development causes tumor growth. Three different candidate genes were <br/>investigated: RasV12, PvrACT, and Avli.
ContributorsSt Peter, John Daniel (Author) / Harris, Rob (Thesis director) / Varsani, Arvind (Committee member) / School of Molecular Sciences (Contributor) / Department of Psychology (Contributor) / Barrett, The Honors College (Contributor)
Created2021-05
Description
The transcriptome of an organism is a collection of the various messenger RNAs that the genes of an organism produce. As the level of gene expression is different between different tissues of an organism, understanding the transcriptome serves as a way to better understand the differences between the functions and abilities of tissues and cells in an organism. This understanding of the transcriptome can aid further research in targeted disease treatments and indentifying new biomarkers. This study aims to gather the transcriptome from various tissues of the organism Daphnia pulex. This will be done by using a combination of single cell RNA sequencing (scRNA-seq), which involves the isolation and sequencing of single cells, and single nuclei RNA sequencing (snRNA-seq), which involves the isolation and sequencing of single nuclei. Here we show the viability of isolating single cells and single nuclei from various Daphnia pulex tissues using different techniques and enzymes including trypLE, trypsin EDTA, accutase, etc by using microscopy and automatic cell counting. The results show that each tissue is best isolated using different techniques.
ContributorsShahriari, Ryan (Author) / Lynch, Michael (Thesis director) / Ye, Zhiqiang (Committee member) / Barrett, The Honors College (Contributor) / School of Molecular Sciences (Contributor)
Created2023-05
Description
Immediate early genes (IEGs) are rapidly activated in response to an environmental stimulus, and most code for transcription factors that mediate processes of synaptic plasticity, learning, and memory. EGR3, an immediate early gene transcription factor, is a mediator of biological processes that are disrupted in patients with schizophrenia (SCZ). A microarray experiment conducted by our lab revealed that Egr3 also regulates genes involved in DNA damage response. A recent study revealed that physiological neuronal activity results in the formation of DNA double-stranded breaks (DSBs) in the promoters of IEGs. Additionally, they showed that these DSBs are essential for inducing the expression of IEGs, and failure to repair these DSBs results in the persistent expression of IEGs. We hypothesize that Egr3 plays a role in repairing activity- induced DNA DSBs, and mice lacking Egr3 should have an abnormal accumulation of these DSBs. Before proceeding with that experiment, we conducted a preliminary investigation to determine if electroconvulsive stimulation (ECS) is a reliable method of inducing activity- dependent DNA damage, and to measure this DNA damage in three subregions of the hippocampus: CA1, CA3, and dentate gyrus (DG). We asked the question, are levels of DNA DSBs different between these hippocampal subregions in animals at baseline and following electroconvulsive stimulation (ECS)? To answer this question, we quantified γ-H2AX, a biomarker of DNA DSBs, in the hippocampal subregions of wildtype mice. Due to technical errors and small sample size, we were unable to substantiate our preliminary findings. Despite these shortcomings, our experimental design can be modified in future studies that investigate the role of Egr3 in activity-induced DNA damage repair.
ContributorsKhoshaba, Rami Samuel (Author) / Newbern, Jason (Thesis director) / Gallitano, Amelia (Committee member) / Marballi, Ketan (Committee member) / School of Molecular Sciences (Contributor) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2020-05
Description
In most bird species, females disperse prior to their first breeding attempt, while males remain close to the place they were hatched for their entire lives (Greenwood and Harvey (1982)). Explanations for such female bias in natal dispersal have focused on the potential benefits that males derive from knowing the local environment to establish territories, while females search for suitable mates (Greenwood (1980)). However, the variables shaping dispersal decisions appear more complex (Mabry et al. (2013), Végvári et al. (2018)). There are a number of different variables that could act as a driving force behind dispersal including the social mating system, food competition, inbreeding avoidance, predation, and others. Here, we investigate whether females are the dispersing sex in great-tailed grackles, which have a mating system where the males hold territories and the females choose which territory to place their nest in (Johnson et al. (2000)). We used genetic approaches to identify sex biases in the propensity to disperse. In the experiment, we found that the male grackles were less related to each other while the female grackles were more related to each other. Building on that, the average distance between closely related individuals of the male group was longer than the average distance of closely related females. But, the mantel correlograms for the males and females both lack a consistent trend. Overall, the results indicated suggest that the males are the dispersing sex while the females are potentially philopatric and that the average dispersal distance for the grackle is greater than 2000 meters, the size of the sampling range used in the experiment. These results will inform our long-term study on the relationship between behavioral flexibility and rapid geographic range expansion by elucidating which individuals are likely to experience similar conditions across their lives, and which are likely to face new conditions when they become breeders.
ContributorsSevchik, August L (Author) / Langergraber, Kevin (Thesis director) / Logan, Corina (Committee member) / College of Integrative Sciences and Arts (Contributor) / School of Molecular Sciences (Contributor) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2020-05
Description
When limited for iron, Escherichia coli secretes a siderophore, enterobactin, to solubilize and intake extracellular Fe3+ by a TonB-dependent high-affinity pathway. Consequently, E. coli tonB mutants grow poorly on a medium limited for iron. Upon longer incubation, however, faster growing colonies emerge and overcome this growth defect. The work presented in this paper reports and characterizes these faster growing colonies (revertants) in an attempt to dissect the mechanism by which they overcome the TonB deficiency. Genomic analysis revealed mutations in yejM, a putative inner-to-outer membrane cardiolipin transporter, which are responsible for the faster growth phenotype in a tonB mutant background. Further characterization of the revertants revealed that they display hypersensitivity to vancomycin, a large antibiotic that is normally precluded from entering E. coli cells, and leaked periplasmic proteins into the culture supernatant, indicating a compromised outer membrane permeability barrier. All phenotypes were reversed by supplying the wild type copy of yejM on a plasmid, suggesting that yejM mutations are solely responsible for the observed phenotypes. In the absence of wild type tonB, however, the deletion of all known of cardiolipin synthase genes (clsABC) did not produce the phenotype similar to mutations in the yejM gene, suggesting the absence of cardiolipin from the outer membrane per se is not responsible for the increased outer membrane permeability. These data show that a defect in lipid biogenesis and transport can compromise outer membrane permeability barrier to allow siderophore intake and that YejM may have additional roles other than transporting cardiolipin.
ContributorsQiu, Nan (Author) / Misra, Rajeev (Thesis director) / Bean, Heather (Committee member) / Yu, Julian (Committee member) / School of Molecular Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2019-05
Description
This thesis summarizes the process of writing a children's book about achondroplasia directed at children without genetic disorders. The thesis also includes the children's book The Genetics of Little People that was created during the project.
ContributorsBinsfeld, Allison (Author) / Watkins, Payton (Co-author) / Wilson, Melissa (Thesis director) / Hunt Brendish, Katherine (Committee member) / Barrett, The Honors College (Contributor) / School of Molecular Sciences (Contributor) / School of Human Evolution & Social Change (Contributor)
Created2022-05
ContributorsBinsfeld, Allison (Author) / Watkins, Payton (Co-author) / Wilson, Melissa (Thesis director) / Hunt Brendish, Katherine (Committee member) / Barrett, The Honors College (Contributor) / School of Molecular Sciences (Contributor)
Created2022-05
ContributorsBinsfeld, Allison (Author) / Watkins, Payton (Co-author) / Wilson, Melissa (Thesis director) / Hunt Brendish, Katherine (Committee member) / Barrett, The Honors College (Contributor) / School of Molecular Sciences (Contributor)
Created2022-05