2004 Summer Undergraduate Research and Creative Achievements Forum (MU)
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The MU Summer Undergraduate Research and Creative Achievements Forum showcases the creative and scholarly activities that undergraduates have been engaged in over the summer. All students engaged in scholarly or creative activity with a faculty mentor are invited to present their work.
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Item Bovine oocyte permeability in 1, 2-propanediol(University of Missouri--Columbia. Office of Undergraduate Research, 2004) Young, Alan; Mullen, Steven Francis, 1968-; Critser, John Kenneth; University of Missouri-Columbia. Office of Undergraduate Research; Summer Undergraduate Research and Creative Achievements Forum (2004 : University of Missouri--Columbia)Cryopreservation of oocytes is of great importance both in medical research and in the agricultural industry. Cryopreservation of oocytes allows for future breeding of selected genetic lines of animals. Effective cryopreservation of bovine oocytes has drawn considerable attention due to its application in the agricultural industry. The reproductive cells of genetic lines of cattle can now be cryopreserved and used in the future to breed superior cattle through in vitro fertilization (IVF). Currently, the preservation of bovine oocytes and other cells has had marginal success due to damage sustained to the cell during freezing and thawing due to volume fluctuations (Mazur et al; Experimental Cell Research 71(1972) 345-355). The Kedem and Katchalsky model (Biochem Biophys Acta, 1958, 27:229-246) can be utilized to model changes in cellular volume during freezing and thawing in cryoprotectant solutions. This model takes into account the specific plasma membrane permeability of the cell that is exposed to a particular solution. The purpose of this study was to determine the hydraulic conductivity, Lp, and the permeability coefficient for the cryoprotective agent, 1, 2-propandiol (PrOH), PCPA, for bovine oocytes. The activation energies of each of these parameters can be determined under the assumption that the plasma membrane permeability parameters follow an Arrhenius relationship. Experimental trials were conducted at temperatures of 30°C, 20°C, 10°C, and 4°C. In order to study the response of a single bovine oocyte to 1.5 M 1, 2-propanediol, a micro-pipette holding device (Gao et al.; Biophysics J, 71:443-450) was used to immobilize the oocyte in a small drop of TL-Hepes media. The oocyte was then abruptly exposed to 1.5 M PrOH media. The volume change of the oocyte (dv/dt) was recorded with a digital camera that was mounted to an inverted light microscope. The area of the cell in each image was calculated with a Fovea Pro Software plug-in to Adobe Photoshop. The volume of the cell was calculated from the calculated area, assuming that the cell was spherical. The constants, Lp and PCPA, were numerically approximated assuming that the cellular volume dynamics followed the Kedem and Katchalsky model. It was found that the oocytes underwent osmotically driven volume changes upon exposure to the cryoprotectant, 1, 2-propanediol. The bovine oocytes contracted more rapidly at higher temperatures. The oocytes also regained their isosmotic volume faster at higher temperatures. The permeability parameters found in this study along with there activation energies will be used in the future to develop an optimal cryopreservation protocol for bovine oocytes through computer-based modeling.Item Examination of the robustness of viscosity sensitive molecular rotors against chemical modification(University of Missouri--Columbia. Office of Undergraduate Research, 2004) Wilson, Joseph P.; Haidekker, Mark A., 1963-; University of Missouri-Columbia. Office of Undergraduate Research; Summer Undergraduate Research and Creative Achievements Forum (2004 : University of Missouri--Columbia)Molecular rotors, a special class of compounds which form twisted intramolecular charge transfer complexes (TICT), have dual deexcitation processes - intramolecular rotation and fluorescence emission. Molecular rotors exhibit a quantum yield that is sensitive to changes in solution viscosity; this property makes molecular rotors highly suitable for optical measurements of viscosity. The goal of this study was to test the idea that increasing the chain length of the molecular rotor increases its viscosity sensitivity. Molecular rotors of varying chain length were selected for experimentation; DCVJ, a thoroughly studied molecular rotor, was chosen as the control. Mixtures of differing ratios of ethylene glycol and glycerol were created to vary viscosity. Fluorescence emission intensity data for the six molecular rotors was measured with a Jobin Yvon Fluoromax-3 spectrofluorometer. Logarithmic graphs of fluorescence intensity versus viscosity were created and displayed a linear relationship with regression values exceeding 0.99. However, the slopes for the various molecular rotors were not significantly different. The results demonstrate that increasing the molecular rotor's chain length does not significantly increase viscosity sensitivity, indicating that additional elements can be attached to the molecular rotor without changing its functionality.Item A multicopy suppressor screen in yeast to look for negative regulators of Ser13 phosphorylation-based trafficking to the pre-vacuolar compartment(University of Missouri--Columbia. Office of Undergraduate Research, 2004) Wilson, Kyle; University of Missouri-Columbia. Office of Undergraduate Research; Summer Undergraduate Research and Creative Achievements Forum (2004 : University of Missouri--Columbia)Resident membrane proteins of the yeast trans-Golgi network (TGN) frequently cycle between the TGN and both the early and late (prevacuolar) endosomal compartments. The model yeast TGN protein, A-ALP, which contains the cytosolic domain of the TGN protein, depeptidyl aminopeptidase A (DPAP A), and the transmembrane and lumenal domains of the vacuolar membrane protein, alkaline phosphatase, have been studied. The cytosolic domain of A-ALP has been shown to be phosphorylated in vivo and in vitro. Of the 25 potentially phosphorylatable residues, only one, Ser13, was observed to influence trafficking between the TGN and endosomes. It has been suggested that phosphorylation of Ser13 is required for trafficking of A-ALP from the TGN to the pre-vacuolar compartment, which implies that phosphorylation of Ser13 may act as a switch for association of A-ALP with vesicular trafficking machinery. It is also important to note that once A-ALP reaches the vacuole by way of the PVC, it is cleaved in the lumenal domain, which allows us to follow the trafficking of the protein. We designed a screen to overexpress negative regulators, such as phosphatase, so that A-ALP will exhibit slower trafficking to the pre-vacuolar compartment. Yeast that seemed to have slower A-ALP trafficking to the PVC were identified by an ALP Overlay Assay. These positives are now being screened by western blot in order to verify that these cells exhibit slower trafficking to the PVC and eventually the vacuole. This can be determined by a western blot by examining the amount of processing of A-ALP in the vacuole. Those that are processed less give the indication of slower trafficking to the PVC, which is possibly the result of high phosphatase activity.Item From pyruvate to PEP...the unknown pathway(University of Missouri--Columbia. Office of Undergraduate Research, 2004) Wigfall, Darian; Oehrle, Nathan Wayne, 1972-; University of Missouri-Columbia. Office of Undergraduate Research; Summer Undergraduate Research and Creative Achievements Forum (2004 : University of Missouri--Columbia)Soybean is a major crop in the United States. Its survival and relationship with its environment are closely monitored. For example, we are looking at the symbiotic relationship between soybean and the soil bacteria. These bacteria can convert nitrogen in the air into usable nitrogen for the soybean plant. This process occurs when the rhizobium forms a nodule on the soybean root. These nodules are complex, hyperplastic tissue masses derived from cortical cells that transport nitrogen as uerides. The soybean plant then in turn acts as a carbon and energy source for the bacteria. The metabolic pathway for the bacteria's reception and consumption of this carbon is unknown. Our specific focus is to use in gel assays to identify the presence of glyceraldehydes-3-P-dehydrogenase, phosphoglycerate kinase, phosohpoglycerate mutase and enolase, used by the plant to synthesize a reaction to convert glyceraldehydes-3-phosphate into PEP (phosphoenolpyruvate) which will be conducted after my departure from the lab. We first conduct a protein extraction to isolate only the desired material from the soybean plant. We then use a one-dimensional gel enzyme assay to determine whether our four main enzymes are active (we received positive results for all four). Also, a two-dimensional gel enzyme assay to determine whether the enzyme we have found is indeed the one we think we have identified in one dimension (we have found positive results for all except phosphoglycerate mutase). Finally, we use an enzyme assay to determine if there is enzyme activity by measuring the absorbance of a solution containing substrate and introducing the enzyme. We found the presence and activity of all of the crucial enzymes involved in the pathway. We have pretty much concluded that this pathway, formerly thought to be a part of the Alanine Transport model, is more likely to be a part of the plant's metabolism.Item Therapy of experimental allergic encephalomyelitis under circumstances relevant to human multiple sclerosis(University of Missouri--Columbia. Office of Undergraduate Research, 2004) Wiehagen, Karla; Bell, J.; Ellis, Jason Scott, 1977-; Zaghouani, Habib; University of Missouri-Columbia. Office of Undergraduate Research; Summer Undergraduate Research and Creative Achievements Forum (2004 : University of Missouri--Columbia)Experimental allergic encephalomyelitis (EAE) is an inflammatory disease of the central nervous system (CNS) that resembles human multiple sclerosis (MS). EAE and MS develop when proteins of the myelin sheath that covers axons are released and encounter cells of the immune system such as T lymphocytes. Activation of these lymphocytes will trigger inflammation that destroys the myelin leading to clinical signs that manifest mostly in the form of motion impairment and muscle paralysis. Inactivation of myelin specific lymphocytes is currently viewed as a means to halt immune attack against the brain and reverse the course of disease. In this study we devised an antigen specific approach against EAE and tested its efficacy in an advanced genetic setting, which would better represent the human genetic polymorphism. Therefore, we have created an F1(SJL/J x Bl/6) mouse for analysis of the reversal of compatible as well as “in trans” EAE where the disease is induced by a peptide restricted to one parent and the treatment uses a fusion protein carrying a peptide restricted to the other parent. The results indicate that the effectiveness of the treatment depends on the method of disease induction and the genetic makeup of the mouse.