Team:PennState
From 2008.igem.org
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<p> The <em>Nuclear Fusion</em> project currently has two similar directions that it may turn but both involve a plasmid construct very generously donated to our iGEM team from David W. Wood, Department of Chemical Engineering at Princeton University. Research in their lab has constructed a biosensor containing just the ligand binding domain (LBD) of the estrogen receptor (ER). The ER is very similar to the PPAR and other hormone receptors. Previous attempts at isolating the LBD failed due to the specific folding pattern of this region required to maintain similar binding characteristics to natural ER. The folding pattern was kept by inserting the LBD into a minimal splicing intein domain. This construct was also made more soluble by addition of a maltose-binding tag. The ER was fused with a thymidylate synthase enzyme (TS) that remains deactivated until homodimerization of the ER after binding ligand. The cells are grown on thymine-free plates allowing for recognition of strength and function of ER ligands.</p> | <p> The <em>Nuclear Fusion</em> project currently has two similar directions that it may turn but both involve a plasmid construct very generously donated to our iGEM team from David W. Wood, Department of Chemical Engineering at Princeton University. Research in their lab has constructed a biosensor containing just the ligand binding domain (LBD) of the estrogen receptor (ER). The ER is very similar to the PPAR and other hormone receptors. Previous attempts at isolating the LBD failed due to the specific folding pattern of this region required to maintain similar binding characteristics to natural ER. The folding pattern was kept by inserting the LBD into a minimal splicing intein domain. This construct was also made more soluble by addition of a maltose-binding tag. The ER was fused with a thymidylate synthase enzyme (TS) that remains deactivated until homodimerization of the ER after binding ligand. The cells are grown on thymine-free plates allowing for recognition of strength and function of ER ligands.</p> | ||
- | <p>Our plan for this project is to work on the sensitivity of the biosensor in hopes of using this for water prescreens, similar to the <em>Smart Fold Reporter</em> project. The sensitivity will be focused | + | <p>Our plan for this project is to work on the sensitivity of the biosensor in hopes of using this for water prescreens, similar to the <em>Smart Fold Reporter</em> project. The sensitivity will be focused for BPA which has a very different conformation than the natural agonist of the ER system. This difference causes BPA to bind weakly but still disturbs normal ER function. One idea is to replace the ER LBD from Wood’s biosensor with the estrogen-related receptor (ERR) LBD. The ERR is similar to ER and binds many of the same ligands and has a tendency to bind to the estrogen response element (ERE) in the human body. The one benefit of ERR for our project is that it binds BPA very strongly. Another direction that this project could take would be to analyze the LBD of ER and perform directed evolution to increase BPA sensitivity. During directed evolution, certain regions of the ER LBD would be targeted for random mutagenesis providing a library of mutants in the trillions. The mutant library would be induced with BPA and the best growing colony would be selected, tested, and mutated for further sensitivity. |
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<td colspan="2" style="padding-top:30px; padding-right:30px" valign="top" width="45%"><span style="font-size: 14pt">Diauxie Elimination: <em>Two</em> spoons full of sugar.</span> | <td colspan="2" style="padding-top:30px; padding-right:30px" valign="top" width="45%"><span style="font-size: 14pt">Diauxie Elimination: <em>Two</em> spoons full of sugar.</span> | ||
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- | <p><img src="picture here" alt="[img]" style="float:left; margin:5px;"/>Cellulosic biomass is an abundant and inexpensive energy source, coming from plant waste: ideal for Ethanol production through fermentation. However, biomass contains glucose and xylose sugars in relatively equal ratios, | + | <p><img src="picture here" alt="[img]" style="float:left; margin:5px;"/>Cellulosic biomass is an abundant and inexpensive energy source, coming from plant waste: ideal for Ethanol production through fermentation. However, biomass contains glucose and xylose sugars in relatively equal ratios, preferentially <em>e. coli</em> metabolizes glucose before any other sugar. In this project we attempt to eliminate this phenomenon, called <em>diauxie</em>, and get our cells to utilize both sugars at the same time. Solving this problem will lead to more efficent use of cellulosic biomass including moving towards the future of bioproduction continous processes.</p> |
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Revision as of 17:43, 30 June 2008
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PENN STATE iGEM 2008
Welcome to the Penn State iGEM 2008 team’s website. We are currently working hard at a few different projects for this year's competition. In early May we began brainstorming and came up with a couple of ideas to create biosensors that use human nuclear hormone receptors to recognize potentially harmful ligands. These receptor systems occur naturally in the human body, but our goal is to retain and utilize their functions in Escherichia Coli. We are also finishing up one of last year's projects which is aimed at creating a more efficient bioproduction process by altering how E. Coli selects between the utilization of 5 and 6 carbon sugars. Please explore our website to find out more about us and our projects! If there are any questions or comments about the information on this site please contact us at gjt5001@psu.edu.
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