Serpent Abstracts
Austin Buchanan – The Effects of Essential Oils on Planaria Regeneration
Essential oils have been used to promote wound healing as a form of alternative medicine, but their effect on stem cells and regeneration is unknown. Using morphometric and gene expression analysis, I explore the effects of several essential oils on Dugesia dorotocephala, an important model organism for regeneration studies. Groups of planaria were bisected and separately treated with thyme oil, lavender oil, rosemary oil, or a water control and allowed to regenerate over a period of three weeks. The average length and growth rate for each group were collected over the regeneration period, showing that thyme was toxic, rosemary hindered regeneration, and lavender was equivalent to the control. Gene expression was tested by analyzing a control gene, collagen, and an experimental gene, piwi-1, that is associated with differentiating stem cells. Primers for collagen and piwi-1 were designed using online tools. RNA was successfully extracted from a planarian that had 12 hours of regeneration time. One-step reverse transcriptase-polymerase chain reaction (RT-PCR) and gel electrophoresis using five sets of primers each for collagen and piwi-1 were successfully conducted using the control planaria, and further analysis is underway for experimental planaria.
Dominic Carrano – Selective Cytotoxicity of Laetiporus sulphureus on Acute Lymphocytic Leukemia
Cancer is the second leading cause of death in the United States and globally, with cases continuing to rise all around the world. This disease is one that has no permanent cure and is only managed by the treatment team. One of the most common forms of treatment for managing cancer is chemotherapy, and while effective at killing the cancer cells, these treatments do not selectively target them and result in the death of non-cancerous cells. This project seeks to address this issue by looking to natural therapy, that selectively kills cancer cells, while leaving the healthy cells unaffected. Laetiporus sulphureous is a bracket fungus native to both Europe and North America. It has been used in traditional medicine to treat a variety of diseases and has been shown in laboratory studies to have anti-inflammatory as well as antitumor properties. The secretions will be analyzed using NGC chromatography and the secretions gathered from the mycelia will be used to test for selective cytotoxicity against acute lymphocytic leukemia.
McKenna Fant – Exploring the Effects of Artificial Sweetener on Stem Cells
Modern diets have changed significantly through the popularization of low-calorie substitutes such as artificial sweeteners. Many of the long-term effects of these sweeteners are unknown. We aim to explore the effects of artificial sweeteners on stem cells using planaria as a model organism. Each group has been fed a different type of artificial sweetener including stevia, monk fruit, aspartame, and saccharin before being bisected. The average growth rate of each group will serve as an indicator of stem cell activity. Gene expression related to stem cell proliferation will be quantified by semi-quantitative RT-polymerase chain reaction (PCR) using the SuperScript IV One-Step System. This method will test three specific aims: the effect of different sweeteners on Dugesia physiology, morphometric analysis of planaria regeneration following exposure to artificial sweeteners, and semi-quantitative RT-PCR to compare the regenerative gene expression between the control and experimental groups. Data so far suggest that artificial sweeteners delay planaria growth and regeneration. The control group exhibits a growth rate of 0.283 mm/day and peaked on day 4, while the experimental groups show rates of 0.053-1.94 mm/day and peaked on day 7. RT-PCR is in progress, and will serve as another marker for measuring stem cell activity.
Samirah Jackson – The Community Analysis of Mouse Gut Microbiota Exposed to Triglycerides
This research project will focus on the identification of microbes present in the mice gut microbiota with and without triglyceride (a more specific form of lipid than olive oil) treatment. This will allow for the investigation of the microbiome community compositions and its changes relative to the presence of triglycerides. We hope to find out exactly how the human gut microbiome community is affected by the triglycerides. We believe that this can help us understand how to possibly prevent CDI in the future.
Phoebe Nguyen – Evaluation of the Effects of Estradiol Implants on Presence of Estrogen Receptors in the Arcuate Nucleus of the Thirteen-lined Ground Squirrel
The goal of this research is to elucidate how estradiol impacts the expression of estrogen receptors in the arcuate nucleus region of the hypothalamus in the thirteen-lined ground squirrel (. As a hibernating species, TLGS proceeds through cycles of extreme fat accumulation and hibernation periods through a circannual rhythm (Florant and Healy 2011). Several papers have studied the mechanisms of energy balance in hibernators; however, the mechanisms that govern the balance between these extreme periods of food accumulation and reproductive cycles has yet to be fully understood. In turn, this project will test the effects of estradiol implants on the arcuate nucleus, a region known to contain neurons responsible for regulating food intake, metabolism, and reproduction by using immunohistochemistry and in situ hybridization to label estrogen receptors in the brains of TGLS.
Lane Paul – Annotation of Level 6 contig41 in D. willistoni
Genome annotation is a process that helps to identify genes contained in sequenced contigs, define the gene structures and chromosomal location. Annotation enables further work including prediction of possible changes in gene function and structural changes by comparing orthologs to a known model genome, Drosophila melanogaster. Using Drosophila melanogaster genes, along with several bioinformatic programs, to establish a null hypothesis of evolutionary change from the model melanogaster as investigated on the Flybase Gene Record Finder to newly constructed contig41 of D. willistoni as mirrored on the Genomics Education Project (GEP) genome browser. The bioinformatic gene prediction track programs tentatively predicted upwards of twenty-six coding genes but upon investigation of CDS open reading frame (ORF) possible usage, five genuine genes were annotated out of the total twenty-six possibilities in contig 41. These genes are Cortactin, GC12213, aurA, aurB, and bor. By comparing the contig41 sequence at Basic Local Alignment Search Tool (BLAST) to the orthologs (Cortactin, GC12213, aurA, aurB, and bor) the structural edges of each CDS exon were identified and located in the novel genome to adjust annotations compared to the null depending on phase compatibility, isoform number maintenance closer to null conservation and adjustments due to splicing or nonsense mutation where necessary. There were another two genes (CG31211 and Spartin) that stood out as potential genes albeit with lower conservation alignment scores. Similar procedures were used to investigate the gene structure of the genes. Overall, the orthologs proposed to be coded for within contig41 of the Muller E chromosome in D. willistoni ranged from 100% conserved gene structural boundaries to 100% changes in structural annotation boundaries of the intron-exon CDS junctions.
Lucy Pickens – JAM-A/ZO-3 Expression in PA-1 and SKOV-3 Ovarian Cancer Cell Lines
TJs adhere the apical regions of epithelial cells and play a role in maintaining cellular polarity, stability, and cell signaling. TJ integral membrane proteins claudins (CLDNs), occludin (OCLN) and junctional adhesion molecules (JAMs) make up the tight junction protein complex along with their cytosolic adapter proteins: zona occludins (ZO-1, ZO-2, ZO-3). Previously, out lab has shown overexpression of JAM-A and ZO-3 RNA in endometrial cancer cell lines using microarray analyses. We analyzed a panel of human ovarian cancer cell lines: PA-1 (ovarian teratocarcinoma), SKOV-3 (ovarian adenocarcinoma), and a Normal Ovarian cell line H-6036 for the expression of JAM-A and ZO-3 using immunoblotting. The presence of 84 TJ and TJ-associated genes through microarray analysis of H-6036 RNA. This study found that there was__ difference in the levels of expression of protein in PA-1 and SKOV-3, as opposed to the levels expressed in the H-6036. The microarray analysis of the H-6036 normal ovarian cells provide a baseline of TJ gene expression for comparison in our future microarray analyses of a panel of ovarian cancer cell lines.
Darren Powers – AMPK-Mediated Survival Strategies: A Study of Autophagy in Nutrient-Deprived Cancer Cells
Cancer is the second-leading cause of mortality, with about 10 million deaths annually worldwide. Cancer is characterized by uncontrolled growth of mutated cells in the body. The rapid proliferation of cancer cells results in microenvironmental stresses, including low nutrient and oxygen levels. Cancer cells are characterized by increased glucose uptake. However, cancer cells can exhibit remarkable tolerance for nutrient deprivation, including glucose suppression. Cancer cells strive to acquire additional nutrients by hook or crook to survive metabolic stress and use multiple mechanisms to combat cell death. Nutrient sensing by the signaling molecules plays an important role in maintaining and regulating metabolic processes. Moreover, to compensate for the nutritional and metabolic deficiencies, cells can undergo autophagy when they "eat" and "digest" their cytoplasmic components to provide building blocks for the metabolic processes. This project aims to understand the relationship between autophagy and metabolic signaling under nutrient-stress conditions in cancers. Using the nutrient starvation approach, immunoblotting, and cell viability assays, we are currently studying the intricacies of nutrient sensing cascade and autophagy in response to nutrient stress. Our preliminary data indicates that cancer cells appear to upregulate autophagic functions under glucose- and glutamine-deprived conditions via the AMPK signaling cascade.
Emma Seiler – An Exploration of Efficient Essential Oil Extraction
Essential oils have gained popularity due to their beneficial properties as an antibacterial agent. These oils are extracted mainly from plant matter with traces of essential oils being discovered in brewer spent hops (BSH) and brewers spent grains (BSG), common by-products of beer brewing. There is potential to repurpose these nutrient-rich by-products as sources of macronutrients and essential oils. The purpose of the current study is to identify efficient extraction methods based on product yield. Samples of BSG and BSH were set in water, acetone, and hexanes. The samples were soaked for a week and then magnetically stirred for a week. Water and acetone samples were steam distilled for approximately two hours and attempted extractions using washes and evaporation of solvent, while hexanes were gravity filtered. Lipid extracts were analyzed with Thin Layer Chromatography (TLC). Results show that preliminary research methods were not reproducible, and hexanes/acetone extractions have shown the possibility of extracting higher amounts of oils. These new methods must be investigated to determine the purity and composition of the extracted oils. Further research will analyze, separate, and identify the oils using high performance liquid chromatography (HPLC) and steam distillation.
Davry Amparano, Taylor Spence & Maverick Stephenson – The Annotation of contig53 of D. willistoni
Genome annotation is the process of deriving the structural and functional information of a gene from a raw, newly sequenced genome using different analysis techniques, comparison, estimation and other mining techniques. To characterize a gene’s role in phenotypic expression, gene chromosomal location, structure, and mutational changes must be categorized and sorted for better understanding of the evolutionary process. This project involved use of the Genome Education Partnership’s (GEP) mirror genome browser and associated tools to compare the genome of Drosophila melanogaster along with several other prediction programs to annotate a novel contig from the D. willistoni Muller E chromosome. Within the assembled DNA of contig53, three genes were tentatively identified by homological comparisons to the model melanogaster system (BLAST, Flybase). An additional fourth gene at the 3’ end of the contig was proposed based on RNA obtained from male flies. These comparisons were used to locate the exon/intron boundaries along with coding sequence start/ stops in the orthologous predicted genes. Modifications of boundary edges resulted from possible evolutionary changes in the D. willistoni sequence compared to our model system which were then adjusted to give the final gene structure predictions. Overall, the accurate coordinates of each exon were modified slightly from that of the predicted coordinates. All coordinates for D. willistoni Muller E-contig53 were verified and checked using RNA sequencing data and the GEP gene model checker.
Essential oils have been used to promote wound healing as a form of alternative medicine, but their effect on stem cells and regeneration is unknown. Using morphometric and gene expression analysis, I explore the effects of several essential oils on Dugesia dorotocephala, an important model organism for regeneration studies. Groups of planaria were bisected and separately treated with thyme oil, lavender oil, rosemary oil, or a water control and allowed to regenerate over a period of three weeks. The average length and growth rate for each group were collected over the regeneration period, showing that thyme was toxic, rosemary hindered regeneration, and lavender was equivalent to the control. Gene expression was tested by analyzing a control gene, collagen, and an experimental gene, piwi-1, that is associated with differentiating stem cells. Primers for collagen and piwi-1 were designed using online tools. RNA was successfully extracted from a planarian that had 12 hours of regeneration time. One-step reverse transcriptase-polymerase chain reaction (RT-PCR) and gel electrophoresis using five sets of primers each for collagen and piwi-1 were successfully conducted using the control planaria, and further analysis is underway for experimental planaria.
Dominic Carrano – Selective Cytotoxicity of Laetiporus sulphureus on Acute Lymphocytic Leukemia
Cancer is the second leading cause of death in the United States and globally, with cases continuing to rise all around the world. This disease is one that has no permanent cure and is only managed by the treatment team. One of the most common forms of treatment for managing cancer is chemotherapy, and while effective at killing the cancer cells, these treatments do not selectively target them and result in the death of non-cancerous cells. This project seeks to address this issue by looking to natural therapy, that selectively kills cancer cells, while leaving the healthy cells unaffected. Laetiporus sulphureous is a bracket fungus native to both Europe and North America. It has been used in traditional medicine to treat a variety of diseases and has been shown in laboratory studies to have anti-inflammatory as well as antitumor properties. The secretions will be analyzed using NGC chromatography and the secretions gathered from the mycelia will be used to test for selective cytotoxicity against acute lymphocytic leukemia.
McKenna Fant – Exploring the Effects of Artificial Sweetener on Stem Cells
Modern diets have changed significantly through the popularization of low-calorie substitutes such as artificial sweeteners. Many of the long-term effects of these sweeteners are unknown. We aim to explore the effects of artificial sweeteners on stem cells using planaria as a model organism. Each group has been fed a different type of artificial sweetener including stevia, monk fruit, aspartame, and saccharin before being bisected. The average growth rate of each group will serve as an indicator of stem cell activity. Gene expression related to stem cell proliferation will be quantified by semi-quantitative RT-polymerase chain reaction (PCR) using the SuperScript IV One-Step System. This method will test three specific aims: the effect of different sweeteners on Dugesia physiology, morphometric analysis of planaria regeneration following exposure to artificial sweeteners, and semi-quantitative RT-PCR to compare the regenerative gene expression between the control and experimental groups. Data so far suggest that artificial sweeteners delay planaria growth and regeneration. The control group exhibits a growth rate of 0.283 mm/day and peaked on day 4, while the experimental groups show rates of 0.053-1.94 mm/day and peaked on day 7. RT-PCR is in progress, and will serve as another marker for measuring stem cell activity.
Samirah Jackson – The Community Analysis of Mouse Gut Microbiota Exposed to Triglycerides
This research project will focus on the identification of microbes present in the mice gut microbiota with and without triglyceride (a more specific form of lipid than olive oil) treatment. This will allow for the investigation of the microbiome community compositions and its changes relative to the presence of triglycerides. We hope to find out exactly how the human gut microbiome community is affected by the triglycerides. We believe that this can help us understand how to possibly prevent CDI in the future.
Phoebe Nguyen – Evaluation of the Effects of Estradiol Implants on Presence of Estrogen Receptors in the Arcuate Nucleus of the Thirteen-lined Ground Squirrel
The goal of this research is to elucidate how estradiol impacts the expression of estrogen receptors in the arcuate nucleus region of the hypothalamus in the thirteen-lined ground squirrel (. As a hibernating species, TLGS proceeds through cycles of extreme fat accumulation and hibernation periods through a circannual rhythm (Florant and Healy 2011). Several papers have studied the mechanisms of energy balance in hibernators; however, the mechanisms that govern the balance between these extreme periods of food accumulation and reproductive cycles has yet to be fully understood. In turn, this project will test the effects of estradiol implants on the arcuate nucleus, a region known to contain neurons responsible for regulating food intake, metabolism, and reproduction by using immunohistochemistry and in situ hybridization to label estrogen receptors in the brains of TGLS.
Lane Paul – Annotation of Level 6 contig41 in D. willistoni
Genome annotation is a process that helps to identify genes contained in sequenced contigs, define the gene structures and chromosomal location. Annotation enables further work including prediction of possible changes in gene function and structural changes by comparing orthologs to a known model genome, Drosophila melanogaster. Using Drosophila melanogaster genes, along with several bioinformatic programs, to establish a null hypothesis of evolutionary change from the model melanogaster as investigated on the Flybase Gene Record Finder to newly constructed contig41 of D. willistoni as mirrored on the Genomics Education Project (GEP) genome browser. The bioinformatic gene prediction track programs tentatively predicted upwards of twenty-six coding genes but upon investigation of CDS open reading frame (ORF) possible usage, five genuine genes were annotated out of the total twenty-six possibilities in contig 41. These genes are Cortactin, GC12213, aurA, aurB, and bor. By comparing the contig41 sequence at Basic Local Alignment Search Tool (BLAST) to the orthologs (Cortactin, GC12213, aurA, aurB, and bor) the structural edges of each CDS exon were identified and located in the novel genome to adjust annotations compared to the null depending on phase compatibility, isoform number maintenance closer to null conservation and adjustments due to splicing or nonsense mutation where necessary. There were another two genes (CG31211 and Spartin) that stood out as potential genes albeit with lower conservation alignment scores. Similar procedures were used to investigate the gene structure of the genes. Overall, the orthologs proposed to be coded for within contig41 of the Muller E chromosome in D. willistoni ranged from 100% conserved gene structural boundaries to 100% changes in structural annotation boundaries of the intron-exon CDS junctions.
Lucy Pickens – JAM-A/ZO-3 Expression in PA-1 and SKOV-3 Ovarian Cancer Cell Lines
TJs adhere the apical regions of epithelial cells and play a role in maintaining cellular polarity, stability, and cell signaling. TJ integral membrane proteins claudins (CLDNs), occludin (OCLN) and junctional adhesion molecules (JAMs) make up the tight junction protein complex along with their cytosolic adapter proteins: zona occludins (ZO-1, ZO-2, ZO-3). Previously, out lab has shown overexpression of JAM-A and ZO-3 RNA in endometrial cancer cell lines using microarray analyses. We analyzed a panel of human ovarian cancer cell lines: PA-1 (ovarian teratocarcinoma), SKOV-3 (ovarian adenocarcinoma), and a Normal Ovarian cell line H-6036 for the expression of JAM-A and ZO-3 using immunoblotting. The presence of 84 TJ and TJ-associated genes through microarray analysis of H-6036 RNA. This study found that there was__ difference in the levels of expression of protein in PA-1 and SKOV-3, as opposed to the levels expressed in the H-6036. The microarray analysis of the H-6036 normal ovarian cells provide a baseline of TJ gene expression for comparison in our future microarray analyses of a panel of ovarian cancer cell lines.
Darren Powers – AMPK-Mediated Survival Strategies: A Study of Autophagy in Nutrient-Deprived Cancer Cells
Cancer is the second-leading cause of mortality, with about 10 million deaths annually worldwide. Cancer is characterized by uncontrolled growth of mutated cells in the body. The rapid proliferation of cancer cells results in microenvironmental stresses, including low nutrient and oxygen levels. Cancer cells are characterized by increased glucose uptake. However, cancer cells can exhibit remarkable tolerance for nutrient deprivation, including glucose suppression. Cancer cells strive to acquire additional nutrients by hook or crook to survive metabolic stress and use multiple mechanisms to combat cell death. Nutrient sensing by the signaling molecules plays an important role in maintaining and regulating metabolic processes. Moreover, to compensate for the nutritional and metabolic deficiencies, cells can undergo autophagy when they "eat" and "digest" their cytoplasmic components to provide building blocks for the metabolic processes. This project aims to understand the relationship between autophagy and metabolic signaling under nutrient-stress conditions in cancers. Using the nutrient starvation approach, immunoblotting, and cell viability assays, we are currently studying the intricacies of nutrient sensing cascade and autophagy in response to nutrient stress. Our preliminary data indicates that cancer cells appear to upregulate autophagic functions under glucose- and glutamine-deprived conditions via the AMPK signaling cascade.
Emma Seiler – An Exploration of Efficient Essential Oil Extraction
Essential oils have gained popularity due to their beneficial properties as an antibacterial agent. These oils are extracted mainly from plant matter with traces of essential oils being discovered in brewer spent hops (BSH) and brewers spent grains (BSG), common by-products of beer brewing. There is potential to repurpose these nutrient-rich by-products as sources of macronutrients and essential oils. The purpose of the current study is to identify efficient extraction methods based on product yield. Samples of BSG and BSH were set in water, acetone, and hexanes. The samples were soaked for a week and then magnetically stirred for a week. Water and acetone samples were steam distilled for approximately two hours and attempted extractions using washes and evaporation of solvent, while hexanes were gravity filtered. Lipid extracts were analyzed with Thin Layer Chromatography (TLC). Results show that preliminary research methods were not reproducible, and hexanes/acetone extractions have shown the possibility of extracting higher amounts of oils. These new methods must be investigated to determine the purity and composition of the extracted oils. Further research will analyze, separate, and identify the oils using high performance liquid chromatography (HPLC) and steam distillation.
Davry Amparano, Taylor Spence & Maverick Stephenson – The Annotation of contig53 of D. willistoni
Genome annotation is the process of deriving the structural and functional information of a gene from a raw, newly sequenced genome using different analysis techniques, comparison, estimation and other mining techniques. To characterize a gene’s role in phenotypic expression, gene chromosomal location, structure, and mutational changes must be categorized and sorted for better understanding of the evolutionary process. This project involved use of the Genome Education Partnership’s (GEP) mirror genome browser and associated tools to compare the genome of Drosophila melanogaster along with several other prediction programs to annotate a novel contig from the D. willistoni Muller E chromosome. Within the assembled DNA of contig53, three genes were tentatively identified by homological comparisons to the model melanogaster system (BLAST, Flybase). An additional fourth gene at the 3’ end of the contig was proposed based on RNA obtained from male flies. These comparisons were used to locate the exon/intron boundaries along with coding sequence start/ stops in the orthologous predicted genes. Modifications of boundary edges resulted from possible evolutionary changes in the D. willistoni sequence compared to our model system which were then adjusted to give the final gene structure predictions. Overall, the accurate coordinates of each exon were modified slightly from that of the predicted coordinates. All coordinates for D. willistoni Muller E-contig53 were verified and checked using RNA sequencing data and the GEP gene model checker.