Team:Clemson/Project

From 2012.igem.org

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             <li><a href="/Team:Clemson/Safety"><img src="http://f.cl.ly/items/3P1e1G2V073P462h2a0j/saftey.png" alt="" width="100" /></a><br />Safety</li>
             <li><a href="/Team:Clemson/Safety"><img src="http://f.cl.ly/items/3P1e1G2V073P462h2a0j/saftey.png" alt="" width="100" /></a><br />Safety</li>
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        <li>Introduction</li>
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            <li><a href="/Team:Clemson/Bio">Biosurfactant - Rhamnolipid</a></li>
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            <li><a href="/Team:Clemson/Pcb">PCB Degradation</a></li>
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            <li><a href="/Team:Clemson/Future">Future Directions</a></li>
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      <p align="justify"><font size="5"><strong> <u>Motivation</u></strong></font></p><br><br>
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             In an effort to fight pollution through the process of bioremediation, our team proposes to develop bacteria that will be highly effective in degrading Polychlorinated Biphenyls (PCBs) with a long term goal of being able to help clean Lake Hartwell, a major recreational area for people of all ages in the upstate of South Carolina. To accomplish this, we have adapted and expanded the model constructed by the 2011 UT Tokyo team. The focus of their project was to engineer a division of bacteria that would increase the efficiency of bioremediation in any given environment. Their two-component system is comprised of a cell assembling and a cell arrest system. Within the parameters of the cell assembling system, their employed mechanism was to use chemoattraction to increase the density of bacteria at a desired location. To accomplish this, aspartate was used as a natural chemoattractant, and its production was induced by the presence of the substrate. The cell arrest system, a secondary extension of system one, is used to maintain a high density of bacteria once they have reached the desired location since it is likely that the concentration of bacteria will dissipate as the substrate is degraded. This system operates by “arresting” or prohibiting the rotation of the flagellum by regulation of the cheZ gene and thereby decreasing cell motility. Following a few key modifications, such as a change in promoters, this system will be adjusted to work perfectly for our project.</strong></p>
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             From 1955 to 1987 in Pickens, South Carolina (the neighboring town to Clemson, SC) Sangamo-Weston Inc. had multiple factories, where they built capacitors. The dielectric fluid used in these capacitors contained Polychlorinated Biphenyls (PCBs). Due to improper land-burial of off-specification capacitors and wastewater treatment sludges that were built both on-site and off-site, PCBs contaminated the water supply of Lake Hartwell and its tributaries. Since PCBs have low water solubility, they remain mostly in the organic sediments and as particulate matter. Furthermore, PCB sediment concentrations are within the range of injury for benthic macroinvertabrates and exceed healthy levels in the fish of Lake Hartwell – a 87.5 square mile recreational lake that serves both South Carolina and Georgia areas.             </strong></p>
   <p>&nbsp;</p>           
   <p>&nbsp;</p>           
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  <p align="justify"><font size="5"><strong> <u>Introduction</u></strong></font></p><br><br>
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  Overall, our project goal is to create a three component bacterial system: signaling, biosurfactant, and PCB degradation. First is an expansion of the UT-Tokyo 2011 Team’s bioremediation specific system, consisting of guider and worker bacteria. The system (currently activated by UV light) causes the guider bacteria to produce AspartateA, which is converted to L-Asp, a natural chemoattractant for E. coli. Thus, both guiders and workers will respond when the bacteria come into contact with the substrate. However, the reaction producing L-Asp is reversible and would cause the bacteria to no longer stay with the substrate. In order to circumvent this, UT-Tokyo created a system with cheZ knockout E. coli and initiated substrate-dependent cheZ expression. Thus, in the presence of the substrate, C1 is produced, binding to the c1-repressed promoter, inhibiting cheZ expression, and halting bacterial movement. In order for this system to work for our purposes, we are to replace the UV light promoter with a PCB specific promoter. Ideally we would like to implement this system in magnetotactic bacteria since PCBs are insoluble particles that remain in the sediments of Lake Hartwell, we will need the guiders to dive to the bottom. The workers in this system will be comprised of two genetically different bacteria: one programmed to produce a biosurfactant and the other to degrade the PCBs to less harmful byproducts. </strong></p>
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             <img src="http://f.cl.ly/items/2Q401m1X0N3i2c1F1C3s/sec1.png" width="450" height="400" frameborder="0"><br><p align="center"> <font size="2"><strong>Signaling Guider Bacteria</strong></font></p>
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            In an effort to fight pollution through the process of bioremediation, our team proposes to develop bacteria that will be highly effective in degrading Polychlorinated Biphenyls (PCBs) with a long term goal of being able to help clean Lake Hartwell, a major recreational area for people of all ages in the upstate of South Carolina. To accomplish this, we have adapted and expanded the model constructed by the 2011 UT Tokyo team. The focus of their project was to engineer a division of bacteria that would increase the efficiency of bioremediation in any given environment. Their two-component system is comprised of a cell assembling and a cell arrest system. Within the parameters of the cell assembling system, their employed mechanism was to use chemoattraction to increase the density of bacteria at a desired location. To accomplish this, aspartate was used as a natural chemoattractant, and its production was induced by the presence of the substrate. The cell arrest system, a secondary extension of system one, is used to maintain a high density of bacteria once they have reached the desired location since it is likely that the concentration of bacteria will dissipate as the substrate is degraded. This system operates by “arresting” or prohibiting the rotation of the flagellum by regulation of the cheZ gene and thereby decreasing cell motility. Following a few key modifications, such as a change in promoters, this system will be adjusted to work perfectly for our project.</strong></p>
 
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            In an effort to fight pollution through the process of bioremediation, our team proposes to develop bacteria that will be highly effective in degrading Polychlorinated Biphenyls (PCBs) with a long term goal of being able to help clean Lake Hartwell, a major recreational area for people of all ages in the upstate of South Carolina. To accomplish this, we have adapted and expanded the model constructed by the 2011 UT Tokyo team. The focus of their project was to engineer a division of bacteria that would increase the efficiency of bioremediation in any given environment. Their two-component system is comprised of a cell assembling and a cell arrest system. Within the parameters of the cell assembling system, their employed mechanism was to use chemoattraction to increase the density of bacteria at a desired location. To accomplish this, aspartate was used as a natural chemoattractant, and its production was induced by the presence of the substrate. The cell arrest system, a secondary extension of system one, is used to maintain a high density of bacteria once they have reached the desired location since it is likely that the concentration of bacteria will dissipate as the substrate is degraded. This system operates by “arresting” or prohibiting the rotation of the flagellum by regulation of the cheZ gene and thereby decreasing cell motility. Following a few key modifications, such as a change in promoters, this system will be adjusted to work perfectly for our project.</strong></p>
 
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            In an effort to fight pollution through the process of bioremediation, our team proposes to develop bacteria that will be highly effective in degrading Polychlorinated Biphenyls (PCBs) with a long term goal of being able to help clean Lake Hartwell, a major recreational area for people of all ages in the upstate of South Carolina. To accomplish this, we have adapted and expanded the model constructed by the 2011 UT Tokyo team. The focus of their project was to engineer a division of bacteria that would increase the efficiency of bioremediation in any given environment. Their two-component system is comprised of a cell assembling and a cell arrest system. Within the parameters of the cell assembling system, their employed mechanism was to use chemoattraction to increase the density of bacteria at a desired location. To accomplish this, aspartate was used as a natural chemoattractant, and its production was induced by the presence of the substrate. The cell arrest system, a secondary extension of system one, is used to maintain a high density of bacteria once they have reached the desired location since it is likely that the concentration of bacteria will dissipate as the substrate is degraded. This system operates by “arresting” or prohibiting the rotation of the flagellum by regulation of the cheZ gene and thereby decreasing cell motility. Following a few key modifications, such as a change in promoters, this system will be adjusted to work perfectly for our project.</strong></p>
 
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Revision as of 05:48, 2 October 2012

CU iGEM



Motivation



From 1955 to 1987 in Pickens, South Carolina (the neighboring town to Clemson, SC) Sangamo-Weston Inc. had multiple factories, where they built capacitors. The dielectric fluid used in these capacitors contained Polychlorinated Biphenyls (PCBs). Due to improper land-burial of off-specification capacitors and wastewater treatment sludges that were built both on-site and off-site, PCBs contaminated the water supply of Lake Hartwell and its tributaries. Since PCBs have low water solubility, they remain mostly in the organic sediments and as particulate matter. Furthermore, PCB sediment concentrations are within the range of injury for benthic macroinvertabrates and exceed healthy levels in the fish of Lake Hartwell – a 87.5 square mile recreational lake that serves both South Carolina and Georgia areas.

 

Introduction



Overall, our project goal is to create a three component bacterial system: signaling, biosurfactant, and PCB degradation. First is an expansion of the UT-Tokyo 2011 Team’s bioremediation specific system, consisting of guider and worker bacteria. The system (currently activated by UV light) causes the guider bacteria to produce AspartateA, which is converted to L-Asp, a natural chemoattractant for E. coli. Thus, both guiders and workers will respond when the bacteria come into contact with the substrate. However, the reaction producing L-Asp is reversible and would cause the bacteria to no longer stay with the substrate. In order to circumvent this, UT-Tokyo created a system with cheZ knockout E. coli and initiated substrate-dependent cheZ expression. Thus, in the presence of the substrate, C1 is produced, binding to the c1-repressed promoter, inhibiting cheZ expression, and halting bacterial movement. In order for this system to work for our purposes, we are to replace the UV light promoter with a PCB specific promoter. Ideally we would like to implement this system in magnetotactic bacteria since PCBs are insoluble particles that remain in the sediments of Lake Hartwell, we will need the guiders to dive to the bottom. The workers in this system will be comprised of two genetically different bacteria: one programmed to produce a biosurfactant and the other to degrade the PCBs to less harmful byproducts.

 




Signaling Guider Bacteria