Team:Virginia/Project

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<a href="https://2008.igem.org/Team:Virginia"><img
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src="http://people.virginia.edu/~drt5p/VGEM/finalwiki/icons/home.gif">Home</a><br>
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<a href="https://2008.igem.org/Team:Virginia/People"><img
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src="http://people.virginia.edu/~drt5p/VGEM/finalwiki/icons/team.gif">People</a><br>
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<a class="active" href="https://2008.igem.org/Team:Virginia/Project"><img
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src="http://people.virginia.edu/~drt5p/VGEM/finalwiki/icons/project.gif">Projects&nbsp;</a><span onClick="showHide('projects')"
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class=expander>&raquo;</span><br>
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<div id=projects class=hide>
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<a href="https://2008.igem.org/Team:Virginia/Project#ga">Genetic Attenuators</a><br>
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<a href="https://2008.igem.org/Team:Virginia/Project#ph">BioBrick Placeholders</a><br>
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<a href="https://2008.igem.org/Team:Virginia/Project#bp">BioPlastic</a><br>
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<a href="https://2008.igem.org/Team:Virginia/Project#rsbp">Adding to the RSBP</a><br>
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<a href="https://2008.igem.org/Team:Virginia/Parts"><img
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src="http://people.virginia.edu/~drt5p/VGEM/finalwiki/icons/parts.gif">BioBricks</a><br>
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<a href="https://2008.igem.org/Team:Virginia/Results"><img
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src="http://people.virginia.edu/~drt5p/VGEM/finalwiki/icons/modeling.gif">Results</a><br>
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<a href="https://2008.igem.org/Team:Virginia/Notebook"><img
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src="http://people.virginia.edu/~drt5p/VGEM/finalwiki/icons/notebook.gif">Notebook</a><br>
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<br><span>We'd like to thank our generous sponsors for making our work possible:</span><br>
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<img class="logos" src="http://people.virginia.edu/~drt5p/VGEM/finalwiki/uva-logo-patch.png"
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alt="University of Virginia" title="University of Virginia" /><img class="logos" src="http://people.virginia.edu/~drt5p/VGEM/finalwiki/dupont.gif" alt="duPont"
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title="duPont" />
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<h2>The Projects</h2>
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<center><img src="https://static.igem.org/mediawiki/2008/thumb/d/d5/Board.JPG/800px-Board.JPG" alt="VGEM's brainstorming whiteboard" width="650"></center>
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<b>Fundemental Project</b> - Prokaryotic transcription attenutation by terminator engineering
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<p>Translating or otherwise implementing a metabolic pathway poses many problems. A major one is the need to tune the relative expression levels of the genes involved in the pathway. The goal is usually to optimize the flux of the intermediates involved. There are many levels at which regulation of expression can occur. Transcriptional, translational and post-translational regulation all exist in natural biological systems. Synthetic biologists have been working on gaining control of these mechanisms at each level to have better control of the novel systems they are designing. In transcriptional regulation a major approach for synthetic biology has been promoter engineering. Modifying natural promoters has yielded a library of promoters of varying strengths which can be used to tweak the transcription rate of genes of interest. However this approach is limited in that each gene that one hopes to affect needs to be placed under the control of its own promoter.</p>
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<p>The 2008 team at the University of Virginia will be pursuing another approach to the problem of transcriptional regulation by trying to create a Genetic Attenuator.  Terminators are not 100% efficient. When a polymerase working its way down a DNA strand runs into a terminator not all of them are kicked off the strand, some of them keep going. By changing the structure of the terminator we hope to create Genetic Attenuators of varying strengths which could be placed between genes to achieve any desired ratio of transcription.</p>
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<b>Applied Project</b> - Controlled polyhydroxybutyrate (PHB) bioplastic synthesis in E. coli.
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<h2>Projects</h2>
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<img src="http://people.virginia.edu/~drt5p/VGEM/finalwiki/vgem-projects.jpg" width=650>
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<p>As the world’s fossil fuel supply starts to decline, new methods of producing fossil-fuel derived products need to be established. Plastics are a major category of fossil-fuel derivatives that make modern life possible. Looking for a solution in the biological realm is an obvious start to this problem as “plastic” is a general term that encompasses “a wide range of synthetic or semisynthetic polymerization products.” Many examples of polymerization products can be found in nature. Among the pathways producing these products is the PHA synthesis pathway found in R. eutropha. This pathway has recently been sequenced allowing the tools of synthetic biology to be applied to translating and optimizing it in E. coli. This is a crucial step towards making this pathway industrially useful as E. coli are a standard chassis used to harness nature’s synthesis capabilities. </p>
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<h3>Genetic Attenuator</h3>
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<p>Transcriptional attenuation is a very promising new tool for the specification and control of genetic transcription rates. The basic mechanism is simple: stop some fraction of the polymerases before they reach the desired gene to reduce transcription (and thereby translation) of the gene.  Natural terminators already serve this role in their capacity to prevent transcription of undesired DNA, but a heretofore untapped resource is the potential inefficiency of this termination.  For instance, imagine a terminator that interrupts only 50% of the polymerases transcribing it. If placed between two genes, this will result in 100% more copies of mRNA corresponding to the first gene than the second in the total transcript output. Without any sophisticated empirical testing and tuning (as is often required for promoter engineering), using a tool directly out of the BioBrick toolbox, an accurate and reliable ratio of gene transcription can be established.</p>
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<span>&nbsp;</span>
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</div>
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<div class="pagebox" id=ph>
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<h3>BioBrick Placeholders</h3>
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<p>A new technical standard!</p>
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<p>Although standardized, assembling composite BioBricks from basic parts has its limitations. For example, parts are added to the ends of a BioBrick. In other words, there is no easy way to insert a new part within an already constructed BioBrick. That is, until now! We've developed a new technical standard called BioBrick Placeholders. Simple and elegant, these BioBricks serve as placeholders by providing unique but compatible multiple cloning sites. BioBrick Placeholders are simply BioBrick-compatible restriction sites flanked by the standard BioBrick prefix and suffix. Next time you go to build a gene but you're not sure which RBS to use, go ahead and put in a BioBrick Placeholder to reserve a spot for that RBS. Later, after you've amplified your construct, you can insert any RBS you'd like.
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<br><br>
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The following 8 BioBrick Placeholders have been submitted to the Registry.  These parts include compatible restriction sites as listed in "Engineering BioBrick Vectors from BioBrick parts."
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<ol>
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<li>EX - ApoI (NotI) AvrII - SP</li>
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<li>EX - ApoI (NotI) NheI - SP</li>
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<li>EX - ApoI (NotI) NsiI - SP</li>
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<li>EX - ApoI (NotI) SbfI - SP</li>
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<li>EX - MfeI (NotI) AvrII - SP</li>
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<li>EX - MfeI (NotI) NheI - SP</li>
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<li>EX - MfeI (NotI) Nsil - SP</li>
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<li>EX - MfeI (NotI) SbfI - SP</li>
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</ol>
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<span>&nbsp;</span>
</div>
</div>
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<div class="pagebox" id=bp>
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<h3>BioPlastic</h3>
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<p>Making plastic a renewable resource.</p>
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<p><i>Rastonia eutropha</i> naturally produces PHB for long-term carbon storage. Taking advantage of this microbes metabolic abilities, we've codon-optimized the 3 essential genes that code for the PHB biosynthesis pathway for expression in E. coli. The enzymes that are produced are PhaA, PhaB1 and PhaC1, which take acetyl-CoA to acetoacetyl-CoA, acetoacetyl-CoA to 3-hydroxybutyryl-CoA, and 3-hydroxybutyryl-CoA to poly-3-hydroxybutyrate (PHB), respectively. Why bother synthesizing PHB? It has the potential to replace polypropylene, a petroleum derivative, as main component of plastic materials.</p>
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<span>&nbsp;</span>
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<div class="pagebox" id=rsbp>
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<h3>Adding to the Registry</h3>
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<p>More tools in the toolbox.</p>
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<p>Over the course of our research we identified a few key areas in the registry where we felt it was lacking. Namely we saw a dearth of reporters as well as a limited number of anti-biotic resistances to work with. Expanding the scope of the registry is a key aspect of iGEM and we feel glad to contribute.</p>
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<p><b>Reporters</b> are a staple of modern biology. This year's <a href="http://nobelprize.org/nobel_prizes/chemistry/laureates/2008/">Nobel Prize in Chemistry</a> was awarded for the discovery of GFP. Reporters such as GFP allow the visualization and monitoring of synthetic biological systems. However, there are only a handful of fluorescent proteins available in the Registry at this time. Diversity in reporters allowed projects such as <a href="http://www.nature.com/news/2007/071031/full/news.2007.209.html">Brainbow</a> to be created. We have thus added two new reporters to the registry: Strongly Enhanced Blue Fluorescent Protein (<a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K156010">SBFP2</a>) and Orange Fluorescent Protein (<a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K156009">OFP</a>).</p>
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<img src="http://people.virginia.edu/~drt5p/VGEM/finalwiki/vgem-ofp.jpg" width=400/>
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<p>OFP in action. As synthetic biological systems become more complex, there will be a need for many more FPs and, eventually, better ways to visualize what's going on inside the cell. </p><br>
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<p>We also introduce a new <b>antibiotic resistance</b> (to streptomycin) to allow more flexibility and diversity in plasmid construction (<a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K156011">aadA</a>).</p>
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<span>&nbsp;</span>
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Latest revision as of 03:58, 30 October 2008


Home
People
Projects »
BioBricks
Results
Notebook
Locations of visitors to this page
We'd like to thank our generous sponsors for making our work possible:
University of VirginiaduPont

Projects

Genetic Attenuator

Transcriptional attenuation is a very promising new tool for the specification and control of genetic transcription rates. The basic mechanism is simple: stop some fraction of the polymerases before they reach the desired gene to reduce transcription (and thereby translation) of the gene. Natural terminators already serve this role in their capacity to prevent transcription of undesired DNA, but a heretofore untapped resource is the potential inefficiency of this termination. For instance, imagine a terminator that interrupts only 50% of the polymerases transcribing it. If placed between two genes, this will result in 100% more copies of mRNA corresponding to the first gene than the second in the total transcript output. Without any sophisticated empirical testing and tuning (as is often required for promoter engineering), using a tool directly out of the BioBrick toolbox, an accurate and reliable ratio of gene transcription can be established.

 

BioBrick Placeholders

A new technical standard!

Although standardized, assembling composite BioBricks from basic parts has its limitations. For example, parts are added to the ends of a BioBrick. In other words, there is no easy way to insert a new part within an already constructed BioBrick. That is, until now! We've developed a new technical standard called BioBrick Placeholders. Simple and elegant, these BioBricks serve as placeholders by providing unique but compatible multiple cloning sites. BioBrick Placeholders are simply BioBrick-compatible restriction sites flanked by the standard BioBrick prefix and suffix. Next time you go to build a gene but you're not sure which RBS to use, go ahead and put in a BioBrick Placeholder to reserve a spot for that RBS. Later, after you've amplified your construct, you can insert any RBS you'd like.

The following 8 BioBrick Placeholders have been submitted to the Registry. These parts include compatible restriction sites as listed in "Engineering BioBrick Vectors from BioBrick parts."

  1. EX - ApoI (NotI) AvrII - SP
  2. EX - ApoI (NotI) NheI - SP
  3. EX - ApoI (NotI) NsiI - SP
  4. EX - ApoI (NotI) SbfI - SP
  5. EX - MfeI (NotI) AvrII - SP
  6. EX - MfeI (NotI) NheI - SP
  7. EX - MfeI (NotI) Nsil - SP
  8. EX - MfeI (NotI) SbfI - SP
 

BioPlastic

Making plastic a renewable resource.

Rastonia eutropha naturally produces PHB for long-term carbon storage. Taking advantage of this microbes metabolic abilities, we've codon-optimized the 3 essential genes that code for the PHB biosynthesis pathway for expression in E. coli. The enzymes that are produced are PhaA, PhaB1 and PhaC1, which take acetyl-CoA to acetoacetyl-CoA, acetoacetyl-CoA to 3-hydroxybutyryl-CoA, and 3-hydroxybutyryl-CoA to poly-3-hydroxybutyrate (PHB), respectively. Why bother synthesizing PHB? It has the potential to replace polypropylene, a petroleum derivative, as main component of plastic materials.

 

Adding to the Registry

More tools in the toolbox.

Over the course of our research we identified a few key areas in the registry where we felt it was lacking. Namely we saw a dearth of reporters as well as a limited number of anti-biotic resistances to work with. Expanding the scope of the registry is a key aspect of iGEM and we feel glad to contribute.

Reporters are a staple of modern biology. This year's Nobel Prize in Chemistry was awarded for the discovery of GFP. Reporters such as GFP allow the visualization and monitoring of synthetic biological systems. However, there are only a handful of fluorescent proteins available in the Registry at this time. Diversity in reporters allowed projects such as Brainbow to be created. We have thus added two new reporters to the registry: Strongly Enhanced Blue Fluorescent Protein (SBFP2) and Orange Fluorescent Protein (OFP).

OFP in action. As synthetic biological systems become more complex, there will be a need for many more FPs and, eventually, better ways to visualize what's going on inside the cell.


We also introduce a new antibiotic resistance (to streptomycin) to allow more flexibility and diversity in plasmid construction (aadA).

 

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