Life and Environmental Sciences Course-based Research Projects

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    UV Light Exposure and Tetrahymena thermophilia: Expression of the PHR2 Gene
    (2018-07-01) Moloney, Molly; Gustavsen, Tanner; Otto-Hitt, Stefanie; Stefanie Otto-Hitt
    The PHR2 (photolyase repair) gene functions in the repair of DNA in Tetrahymena thermophila. It was hypothesized that when Tetrahymena cultures are exposed to UV light, increased expression of PHR2 would occur in response to the damage incurred. UV light has been known to cause the formation of pyrimidine dimers and other disruptions in DNA. Through two rounds of experimentation, the acute impact of UV light exposure was monitored. The cultures underwent RNA extraction, reverse transcription, and semi-quantitative PCR analysis. We predicted that UV exposure would result in the increased expression of PHR2.
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    Effect of Nitrate Pollution on Tetrahymena thermophila
    (2018-07-01) DeBruycker, Reegan; Sylvester, Hannah; Otto-Hitt, Stefanie; Stefanie Otto-Hitt
    The question under investigation in this project was: Does an elevated concentration of nitrates have a negative eect on cell growth and expression of the CDC16 gene in the organism Tetrahymena thermophila? Exposure of Tetrahymena cultures to nitrates mimics the exposure of these organisms to nitrate pollution in the environment, which can occur due to septic tank leakage, nitrogen-rich fertilizer run o, and agricultural processes. The EPA standard for nitrate concentrations in drinking water is a maximum of 10ppm. In our experiment, Tetrahymena cultures were treated with media that contained 30ppm of nitrates for one week. The 30ppm nitrate concentration has been proven acceptable for freshwater fish; therefore, we predicted that it would have more of an impact on our Tetrahymena cells which are far less complex. The CDC16 gene encodes a Cell Division Cycle protein that contains an anaphase-promoting complex (APC). This APC is involved in Cyclin degradation and promotes the movement of the cell out of mitosis. During the oneweek exposure to 30ppm nitrates, the growth rate of Tetrahymena cultures was determined using a hemocytometer. Furthermore, expression of the CDC16 gene was analyzed using semi-quantitative RT-PCR following the weeklong exposure. We predicted that a high concentration of nitrates would decrease the growth rate and expression of CDC16 in our Tetrahymena cultures.
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    The Effect of Exposure to Ionizing Radiation on Cwf16 Expression in Tetrahymena thermophila
    (2018-07-01) Olsen, Shelby; Woolston, David; Stefanie Otto-Hitt
    The following research seeks to determine whether exposure to ultraviolet radiation will induce an increase in the expression of the Cwf16 gene in Tetrahymena thermophila. Cwf16 is a homolog of Ccgc94, whose gene product is a functioning member of the Prp19 complex, a protein complex which protects cells from committing apoptosisprogrammed cell deathdue to exposure to ionizing radiation. The homology between Cwf16 and Ccgc94 yields the hypothesis that Cwf16 expression in T. thermophila will increase in response to UV radiation exposure in a similar mechanism to that observed the Ccgc94 system. In the present research, T. thermophila cultures were randomly assigned into an experimental group and a negative control group, in which the experimental group was subject to incremental exposures to ultraviolet radiation over the course of one week. RT-PCR and gel electrophoresis were used to measure the level of expression of Cwf16 and a hemocytometer was used to measure the growth rate of the control and experimental cultures. Despite the homology between Cwf16 and Ccdc94, we predict that UV-radiation exposure will not raise T. thermophilas expression of Cwf16, because homology does not guarantee that the two genes products serve the same function. The Prp19 complex (the protein complex where we observe Ccdc94s expression) has not been observed in T. thermophila, either because researchers have not looked for it, or because T. thermophila do not have the Prp19 complex in its biochemical repertoire.
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    The Effects of Warming Temperatures on Rad51 Expression in Tetrahymena thermophila
    (2018-07-01) Brunson, Timothy; Lefstad, Rachael; Otto-Hitt, Stefanie; Stefanie Otto-Hitt
    Our experiment explored whether raised temperatures affected expression of the Rad51 gene in Tetrahymena thermophila. Our hypothesis was that raised temperatures would result in a decrease in expression of the Rad51 gene. The protein product of this gene functions in double stranded DNA repair. We chose this experiment to see if the rising average temperatures around the world would affect expression of genes involved in DNA repair. Our test samples were kept in a growth medium at 37C while our control samples were kept in a growth medium at 24C. Afterwards, the transcribed mRNA was reverse transcribed into DNA and primers were used to amplify the Rad51 gene. Semi-quantitative PCR was then used to analyze expression of the Rad51 gene. Our prediction was that higher temperatures would decrease the expression of the Rad51 gene.
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    Effects of the Everyday Toxin, Titanium Dioxide, on Drosophila melanogaster Nervous System Development
    (2018-07-01) Carparelli, Maria; Ripley, Kelsey; Otto-Hitt, Stefanie; Stefanie Otto-Hitt
    Titanium dioxide (TiO2) is a widely used compound found in everything from food packaging to sunscreens. When ingested, TiO2 is readily transported across membranes and efficiently stored within cells. Previous studies showed that exposure to TiO2 results in underdeveloped nervous systems. For our study, we attempted to answer the following question: Will exposing Drosophila melanogaster larvae to TiO2 affect expression of the Neur gene and development of the central nervous system? The Neur gene is crucial during the celldetermination stage of development as its encoded protein helps specify neuroblast development and aids in nervous system and sensory organ development. It was hypothesized that expression of the Neur gene would decrease in Drosophila larvae exposed to TiO2 and that nervous system development would be abnormal compared to control larvae. To test this hypothesis, Drosophila larvae were randomly assigned to either a control group, which was cultured under ideal conditions, or a treatment group, which was exposed to a non-lethal concentration of TiO2. Following exposure, RNA extraction and Reverse Transcription Polymerase Chain Reaction (RT-PCR) was conducted to analyze expression of Neur. To quantify nervous system development, Drosophila larvae were subjected to a touch-response assay. Because TiO2 likely interferes with neural development, it was predicted that Drosophila larvae would show decreased expression of Neur and that they would respond poorly to a mechanical touch-response assay.