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ItemMediating Internalin A-dependent entry of microspheres in epithelial cells(Georgia Institute of Technology, 2013-12-19) Bhyravabhotla, KshamaInternalin A, an internalin protein found in the food-borne pathogen Listeria monocytogenes, allows the pathogen to enter host cells through receptor-mediated internalization. Through Internalin A-mediated entry, L. monocytogenes invades enterocytes by binding to the receptor protein E-cadherin (Bergmann et al 2002). In this way, the pathogen is able to cross the intestinal barrier, a highly selective permeable interface that is responsible for allowing nutrients into the intestinal lumen while barring the entry of waste and pathogens. This study seeks to observe Internalin A-mediated entry of a pathogen mimetic system into epithelial cells. We use polystyrene carboxyl-terminated microspheres to display Internalin A, study the effect on internalization of ligand density and the size of the microsphere. A pGEX plasmid containing the inlA gene, which had previously been purified after transformation into MAX Efficiency DH5αF’IQ E. coli competent cells, was transformed into and expressed in OneShot BL21(DE3)pLysS E.coli competent cells. The result of expression of the plasmid was the Internalin A protein (InlA), combined with a glutathione S-transferase (GST) tag, in order to form a 75 kDa InlA-GST fusion protein. This fusion protein was subsequently purified through affinity chromatography. Concurrently, a protocol for labeling protein with fluorescein isothiocyanate dye (FITC) and covalently coupling the protein to 2 µm microspheres was also developed using ovalbumin. The future steps in this experiment are to successfully cleave the GST tag from Internalin A using sequence-specific protease, functionalize microspheres with purified InlA labeled with FITC and perform internalization studies with microspheres of different sizes and different densities of protein coating. Because InlA can effectively facilitate transport of L. monocytogenes into the cells of the intestinal epithelium, this study has important implications for improving the efficiency of drug delivery to the intestinal lumen.
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ItemCharacterizing a novel direct target of the quorum-sensing controlled small RNAs in V. cholerae(Georgia Institute of Technology, 2013-12-13) Elsherbini, Joseph Ahmedn/a
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ItemRibonucleotides in yeast genomic DNA are targets of RNase H2 and nucleotide excision repair(Georgia Institute of Technology, 2013-12-13) Shetty, LahariRibonucleotides can be incorporated into the yeast genome through a variety of mechanisms, including through DNA polymerazation, DNA priming, and oxidative damage. Ribonucleotides contain a reactive 2’ hydroxyl group on the sugar, which can distort the DNA double helix and lead to defective replication and transcription and ultimately mutagenesis. Ribonucleotide excision repair (RER) has been found to remove ribonucleotides through the enzyme RNase H2, though the in vivo substrate specificity is not known. Nucleotide excision repair (NER) removes bulky lesions formed in DNA, however its role in the extraction of ribonucleotides has not yet been determined in eukaryotes. Previously developed oligonucleotide-driven gene correction assays in Saccharomyces cerevisiae, or baker’s yeast, have shown that paired and mispaired rNMPs embedded into genomic DNA, if not removed, serve as templates for DNA synthesis and can result in a genetic alteration. We implemented this assay to examine whether RNase H2 and NER can target specific rNMPs in DNA. Our results deliver new evidence that RNase H2 specifically recognizes isolated paired and mispaired rNMPs embedded in yeast genomic DNA and that the NER mechanism can recognize an isolated paired rNMP as damage during DNA double-strand break repair in yeast.
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ItemRemediation Mercuric Ions via Functionalized Magnetic Nanoparticles(Georgia Institute of Technology, 2013-12-12) Bennett, Austin LandryMercuric ions, Hg2+, display a unique affinity for sulfur-containing biomolecules because of the soft acid/soft base interaction between Hg2+ and sulfur.1 Many of these biomolecules are used in signal processing and response;2 therefore, Hg2+ is a potent neurotoxin. Chronic exposure to high Hg2+ levels can lead to mercury-poisoning and even death. This research focused on removing Hg2+ from water. Cobalt ferrite nanoparticles were used to separate Hg2+ ions from water. A fair amount of research has been conducted using ferromagnetic nanoparticles and various nitrogenous and oxygen-based ligands, such as triazene compounds and tartrate ligands, to remediate mercuric ions.3,4 Magnetic nanoparticles (MNPs) were coated with thiol-containing carboxylic acid ligands to bind Hg2+ ions. To combat thiol oxidation, the weak reducing agent dithiothreitol (DTT) was introduced into the Hg2+ solutions. The density of the particles in the Hg2+ solutions and exposure times were varied in order to determine the optimal density and exposure time for Hg2+ removal. In future work, thiol-containing organosilane ligands will be coated onto the particles and tested for Hg2+ removal to be compared against the carboxylic acid ligands.
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ItemIdentifying Promoters of Hepatic Regeneration in Zebrafish (Danio rerio) Following an Acetaminophen Liver Ablation(Georgia Institute of Technology, 2013-12-12) Dattilo, ZacharyIdentifying chemicals that can promote regeneration in damaged liver tissue could be critical for curing various liver diseases and accelerating the healing of liver damage. In order to study the regeneration of developing livers in zebrafish, acetaminophen was explored as a possible method for liver ablation. A chemical screening of over 250 novel compounds with unknown cellular targets and 75 compounds with stem cell targets was performed in order to identify some promising promoters of regeneration. Acetaminophen was found to successfully destroy the liver tissue of developing embryos, demonstrating its usefulness as a method of ablation in order to study regeneration. The chemical screening revealed several novel compounds and cell signalling pathways that show promise for successfully promoting liver regeneration.
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ItemThe Effects of Flow on Swimming Behavior of Brachionus manjavacas (Rotifera)(Georgia Institute of Technology, 2013-12-11) Rittweger, ShelbyRotifers serve as model species and are crucial to the zooplankton communities in terms of feeding and nutrition as well as their overall contribution to aquatic food webs (Wallace et al., 2010). Rotifers experience fluid flow in their natural environments of lakes and streams. Fluid velocity acts as stimulus to rotifers, causing them to adjust their swimming speed and direction. I am interested in how rotifers respond to flow, which is known as rheotaxis (Marcos, 2012). Brachionus manjavacas is the rotifer species employed in my experiments. This study simulates fluid flow at rates similar to that rotifers may experience in a riverine ecosystem with unidirectional flow. My intention is to uncover the ways in which the animals respond to flow in these tightly controlled conditions. Rotifers are categorized by age and tested in flow rates ranging from 0.0 to 1.0 mm/sec. Video analysis enables us to quantify swimming velocity and dissect its directionality. The study observes Brachionus manjavacas behavior in terms of aging and analyzes behavior (swimming) from an ecological perspective. It was observed that two-day-old rotifers swim the fastest on average, while four-day-old animals show fastest swimming patterns against the flow. The end result is a behavioral profile that can be useful for understanding how rotifers adapt to flow.
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ItemThe Initiation, Multiplication, and Cryopreservation of Fraser Fir (Abies fraseri [Pursh] Poir.) Embryogenic Tissue for Somatic Embryogenesis(Georgia Institute of Technology, 2013-12-11) Fischer, Susan TaylorFraser fir (Abies fraseri [Pursh] Poir.) is a coniferous tree native to the southern Appalachian Mountains in the United States. Due to its restricted native range in a high-elevation habitat and long reproductive process, the forces of anthropogenic global climate change and invasive pests have made this species vulnerable to extinction (Conifer Specialist Group 1998). Research on ways to propagate mass numbers of conifers like the Fraser fir and restore forest productivity includes clonal propagation through somatic embryogenesis. Such research is critical to help ensure the survival of this species for both environmental and economic reasons. Fraser fir is the most popular Christmas tree in the United States and the primary Christmas tree species grown in North Carolina, where Christmas tree sales alone brought in a revenue of over $75 million dollars in 2011 (NCDA 2012). To explore potential methods of increasing embryogenic tissue initiation and growth, embryogenic tissue initiation and capture media were supplemented with the redox chemical sodium thiosulfate (158.09 mg/L) and were compared to control media. Although the redox medium yielded a higher average percent initiation (29.3% versus 26.9%), the results were not statistically significant (p > 0.05). To assess the effects of toxic carbohydrate hydrolysis products in autoclaved media, growth of embryogenic tissue was recorded for capture media with autoclaved sucrose and compared to the growth of tissue on media with filter-sterilized sucrose. The non-significant results suggest that filter-sterilization of sucrose is not necessary and does not inhibit embryonic tissue proliferation. High-mass initiations were selected for cryopreservation and were analyzed for new growth after removal from cryogenic storage. Ongoing research includes production of somatic embryos from designated high-yielding cultures removed from cryostorage, propagation of those cultures on maturation media, and germination of normal somatic embryos on germination media to effectively create highly efficient protocols for the somatic embryogenesis of Fraser fir.
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ItemUsing interruptions to study associations in prospective memory(Georgia Institute of Technology, 2013-12-10) Kazi, SadafBackground: Prospective memory (ProM) consists of remembering that some action needs to be performed in the future and when (detecting the Intent Trigger), and what the action is (Recalling the Content of the trigger). The Intent Trigger is bound by a forward association to the Content Recall, and the Content Recall has a backward association to the intent Trigger. In situations which present multiple, interleaving ProM tasks to operators it is not known how subsequently-presented ProM tasks interfere with the associations between the Intent Trigger and Content Recall of the original ProM task. Objective: The current study investigated the effect of presenting multiple, interleaved ProM tasks on timely detection of the Intent Trigger and accurate Recall of the Content of the original ProM task. Method: Participants encoded a ProM task (AB) in an air traffic control simulation. They then were interrupted with a second ProM task. The ProM interruption task was different from the original ProM task in either the Intent Trigger (AB, CB), Content Recall (AB, AD), or both Intent Trigger and Content Recall (i.e., a new ProM task, AB, CD). A control condition involved interrupting the participant with a weather report. Results: Detection of the Intent Trigger was significantly worse after a ProM interruption as compared to a weather interruption; a similar pattern of results, but with marginal significance, was also found for Content Recall. Additionally, a ProM task that interfered with backward association (AB, CB) was no better or worse than doing two unrelated ProM tasks (AB, CD) on the detection of the Intent Trigger. However, a task that presented a new forward association (AB, AD) was worse than performing two unrelated ProM tasks (AB, CD) on Recall of the Content. The results are discussed in the context of designing memory aids to support interleaved ProM tasks in dynamic environments.
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ItemStructured Graphene: from Ballistic Transport to Electronics(Georgia Institute of Technology, 2013-12-02) de Heer, Walter A.The original concept of graphene electronics focused on carbon nanotube properties. Carbon nanotubes were known to be high mobility ballistic, phase coherent conductors and quantum confinement effects produced significant bandgaps. However, it turns out to be very difficult to develop nanotube electronics platform for a variety of reasons including fundamental physical constraints related to the quantum mechanical properties of the metal-to-nanotube contacts. Graphene electronics can in principle overcome the major problems because graphene structures can be patterned using conventional lithography and dissipation at contacts can be controlled. However, these developments rely on the premise that narrow, ballistic graphene ribbons can be produced. Experiments on conventionally patterned graphene structures produced from graphene that is deposited on insulating substrates have been discouraging. The graphene ribbon mobilities are so low due to edge roughness effects, to render this direction to be impracticable. On the other hand, graphene produced on silicon carbide turns has been found to be more immune to edge scattering problems. Moreover, recent developments of template grown graphene structures on silicon carbide are promising. Very narrow ballistic graphene ribbons that demonstrate ballistic transport properties, have been produced with these methods which again brings the original concept of graphene based nanoelectronics back into play.
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ItemUnderstanding older adults' perceptions of usefulness of an assistive home robot(Georgia Institute of Technology, 2013-11-21) Beer, Jenay M.Developing robots that are useful to older adults is more than simply creating robots that complete household tasks. To ensure that older adults perceive a robot to be useful, careful consideration of the users’ capabilities, robot autonomy, and task is needed (Venkatesh & Davis, 2000). The purpose of this study was to investigate the construct of perceived usefulness within the context of robot assistance. Mobile older adults (N = 12) and older adults with mobility loss (N=12) participated in an autonomy selection think aloud task, and a persona based interview. Findings suggest that older adults with mobility loss preferred an autonomy level where they command/control the robot themselves. Mobile older adults’ preferences were split between commanding/controlling the robot themselves, or the robot commands/controls itself. Reasons for their preferences were related to decision making, and were task specific. Additionally, findings from the persona base interview study support Technology Acceptance Model (TAM) constructs, as well as adaptability, reliability, and trust as positively correlated with perceptions of usefulness. However, despite the positive correlation, barriers and facilitators of acceptance identified in the interview suggest that perceived usefulness judgments are complex, and some questionnaire constructs were interpreted differently between participants. Thus, care should be taken when applying TAM constructs to other domains, such as robot assistance to promote older adult independence.