Brown

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== Welcome to our World ==
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[[/Outreach]] (Tito, 10/20/07)
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[[Image:brown_website.jpg|frame|left| [http://www.brownigem.com The Website: Our Public Image]]]
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[[Image:brown_oww.jpg|frame|right| [http://openwetware.org/wiki/IGEM:Brown/2007 The Resource: Our Calculations]]]
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==Our 2 Projects==
 
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===Lead Detector===
 
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Lead poisoning is a public health concern - there is lead in soil, paint, water, and dust. Lead Poisoning is often caused by ingesting contaminated drinking water, or soil. It can cause neurological and gastrointestinal disorders, especially among children.
 
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The legal limit of lead in drinking water is 15 parts per billion.
 
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Current ways of testing for lead either require expensive chemical lab analysis or involve inaccurate home kits.
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A lead detector, based on E. Col, is cheap, sensitive, quick, and specific
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Our system involves a Lead detecting promoter, an amplifier, and an output of GFP. We're also working on a system to remove false positives.
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===Tri-stable Toggle Switch===
 
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The purpose of the Tri-stable Toggle Switch is to produce three distinct, continuous, and stable outputs in response to three distinct inputs.  These three inputs are three separate chemicals which will each induce one state of the switch.  [[Image:Tristable_Toggle_Switch_2007.jpg|thumb|left|The Tri-stable Toggle Switch Architecture]] In order to achieve this goal, we are constructing three constructs, each of which consists of a repressible, constitutively-on promoter attached to two repressors. Specifically, our three constructs are pBAD->LacI->TetR, pLacI->AraC->TetR, and pTet->AraC->LacI, where [http://en.wikipedia.org/wiki/Bcl-2-associated_death_promoter AraC] represses pBAD, [http://en.wikipedia.org/wiki/Lac_repressor LacI] represses pLac and [http://en.wikipedia.org/wiki/Tetracycline_controlled_transcriptional_activation TetR] represses pTet.  Each of the three repressors are inactivated by one of three chemicals, the three inducer chemicals mentioned earlier. These three([http://en.wikipedia.org/wiki/Arabinose arabinose], [http://en.wikipedia.org/wiki/IPTG IPTG] (Isopropyl β-D-1-thiogalactopyranoside) and [http://en.wikipedia.org/wiki/Tetracycline Tetracycline], respectively), cause conformational changes in their respective repressor proteins which leads to gene expression.  For example, in the presence of arabinose, AraC cannot repress pBAD so LacI and TetR are produced which in turn repress pTet and pLac.
 
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====AraC/BAD====
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The gene AraC one of several genes (AraA, AraB, AraD, etc) originally for the metabolism of arabinose.[http://www.mun.ca/biochem/courses/3107/Topics/Ara_operon.html]  [[Image:Two_Dimers_of_AraC.jpg|thumb|left|Dimer structure with arabinose on the left (yellow)]]
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[[Image:AraC_Promoters.gif|left|thumb|The left image shows the araC dimer repressing transcription, while the right conformation enables transcription]]The protein forms a dimer in with and without arabinose but the structural change activates or represses the pBAD ([http://en.wikipedia.org/wiki/Bcl-2-associated_death_promoter Bcl-2-associated death promoter], an apoptotic regulator in humans).
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<img src="https://static.igem.org/mediawiki/2007/a/a0/Brownlogo.gif" alt="Brown University">
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<a href="https://2007.igem.org/Brown"><image src="https://static.igem.org/mediawiki/2007/1/1e/Home_pressed.gif" alt="Home" border=0></a>
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<a href="lead"><image src="https://static.igem.org/mediawiki/2007/2/2a/Lead_reg.gif" alt="Lead Sensor" border=0></a>
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<a href="tristable"><image src="https://static.igem.org/mediawiki/2007/c/c5/Tri_reg.gif" alt="Tristable Switch" border=0></a>
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<a href="about"><image src="https://static.igem.org/mediawiki/2007/c/ca/About_reg.gif" alt="About us" border=0></a>
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<center> <script type="text/javascript" src="http://blip.tv/scripts/pokkariPlayer.js?ver=2007100301"></script><script type="text/javascript" src="http://blip.tv/syndication/write_player?skin=js&posts_id=302880&source=3&autoplay=true&file_type=flv&player_width=&player_height="></script><div id="blip_movie_content_302880"><a rel="enclosure" href="http://blip.tv/file/get/Titok16-BrownIGEM2007588.flv" onclick="play_blip_movie_302880(); return false;"><img src="http://www.brownigem.com/images/Wiki_Teampicture.jpg" title="(left to right) Tito, Rohan, Adam, Deepa, Jeff, Norris, Kyle"></a><br /><a rel="enclosure" href="http://blip.tv/file/get/Titok16-BrownIGEM2007588.flv" onclick="play_blip_movie_302880(); return false;"></a></div> </center>
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<a href="http://blip.tv/file/298237"></a>
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<span style="font-size:18pt">Cellular Lead Sensor</span>
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About 40% of the world does not have access to clean water. Lead is a major contaminant worldwide. In the US alone, over 1 million children ages 1 through 5 have elevated levels of lead in their blood. Current lead detection systems are expensive and require lab analysis. Home lead testing kits are inaccurate and only detect lead at very high levels. We have created a genetic circuit in E Coli that responds to lead. The promoter and lead binding protein we use are ten times more selective for lead than for other similar heavy metals. We have also incorporated a genetic amplifier into our circuit to allow us to detect fairly low concentrations of lead.
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<span style="font-size:18pt">Tristable Switch</span>
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Wouldn't you like to have a stable signal in the continuously fluctuating environment of cells?
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And if, say, your system doesn't work the very first try, wouldn't you like to have a methodical means to go about debugging it?
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The Tri-Stable Toggle Switch represents a continuation on the theme of the Toggle Switch begun by Gardner, et al to produce stable outputs in response to transient inputs.  Applications such as a memory circuit and a drug delivery system are a few suggestions, but perhaps the most promising innovation lies in the design process. Our novel approach to  the Tri-Stable Switch development is founded on quantitative principles, pioneering a technique to remove the guesswork from designing and debugging biological systems.
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====LacI====
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http://www.brownigem.com/images/pfizer.gif
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In nature, LacI represses pLac which promotes LacYZA genes that metabolize lactose, thus LacI represses pLac except in the presence of lactose (or lactose mimics, eg IPTG).  [[Image:LacI_repressor.gif|thumb|left|Image[http://www.mun.ca/biochem/courses/3107/Topics/Lac_genetics.html]. LacI forms a tetramer and represses pLac.  However, an inducer, such as IPTG, causes a conformation change that removes LacI from the operator site.]]    Lactose causes a conformational change which inhibits LacI from binding to the operator site of pLac.  Four LacI proteins form a tetramer to inhibit pLac and four inducer molecules are required to cause the full conformational change in the inhibitor.[http://www.mun.ca/biochem/courses/3107/Topics/Lac_genetics.html]
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<span style="font-size: 14pt; color: #4b8ada">&nbsp; Links
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                                                <td class="news">
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<a href="http://openwetware.org/wiki/IGEM:Brown/2007">Brown iGem on Openwetware</a><br>
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<a href="http://www.brownigem.com">Brown iGem Official Website</a><br>
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<a href="Supplemental">Check out our Videos!</a><br>
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<span style="font-size: 14pt; color: #4b8ada">&nbsp;News Updates
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<b>09.06.07</b><hr size=1 color=#696969>
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Brown University students love the Brown iGEM Booth at the Student Activities Fair! <p>
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<b>09.04.07</b><hr size=1 color=#696969>
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Brown iGEM visits Pfizer HQ in Groton, Conneticut<p>
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<b>08.28.07</b><hr size=1 color=#696969>
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Nanodrop donates ND-1000 Spectrophotometer and ND-3300 Fluorospectrophotometer to Brown iGEM
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<p>
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<b>08.22.07</b><hr size=1 color=#696969>
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Brown iGEM Team presents its August Update to faculty and friends
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<p>
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<b>08.20.07</b><hr size=1 color=#696969>
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Nanodrop it like it's hot! http://youtube.com/watch?v=pKT4Zrjeh6s
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<p>
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<b>08.18.07</b><hr size=1 color=#696969>
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iGEM 2007 Meetup. Teams from MIT, BU, McGill, and Cold Spring Harbor presented their progress to each other and discussed the future of Synthetic Biology.
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<p>
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<b>08.04.07</b><hr size=1 color=#696969>
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Brown iGEM presents summer research at the Brown University UTRA (Undergraduate Research) Symposium
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<p>
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<b>07.30.07</b><hr size=1 color=#696969>
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Deepa and Tito gave a presentation to Brown Summer students and discussed the applications of synthetic biology
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<p>
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<b>07.15.07</b><hr size=1 color=#696969>
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Brown iGEM presents June Update to faculty and friends of iGEM
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<p>
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<b>07.03.07</b><hr size=1 color=#696969>
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The iGEM Team met up with Jody Hall's Biotechnology class on July 3rd. Afterwards we toured the lab and discussed the ethics of synthetic biology.
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<p><b>06.15.07</b><hr size=1 color=#696969>
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Brown iGEM presents June Update to Pfizer Team and Brown community<p>
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<b>06.07.07</b><hr size=1 color=#696969>
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$25,000 grant from Pfizer for equipment purchases
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<p>
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<b>06.01.07</b><hr size=1 color=#696969>
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Labnet sponsors Brown iGEM with vortex mixers, a microcentrifuge, pipettes, and autopipettes.
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====TetR====
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TetR represses the constitutive promoter pTet.  In the presence of tetracycline, an antibiotic, a conformational change in TetR inhibits the protein from binding to the operator region.  In nature, pTet promotes TetR and TetA.  The latter which acts to pump tetracycline out of the cell, thus the pump is only activated in the presence of Tetracycline.[http://en.wikipedia.org/wiki/Tetracycline_controlled_transcriptional_activation]
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The TetR, as it turns out is a very tight repressor and a range of 0 to 1 ug/ml has been shown to cause a 5 order of magnitude change in luciferase production.[http://www.ncbi.nlm.nih.gov/sites/entrez?db=pubmed&cmd=Retrieve&dopt=AbstractPlus&list_uids=1319065&query_hl=1&itool=pubmed_docsum]
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[[Image:Tc_bound_to_TetR.jpg|thumb|left|A tetracycline molecule binds to each of the two TetR monomers to form a dimer]]
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Tetracycline is highly diffusable through cell membrane (permeation coeficient or 5.6±1.9 * 10^-9 cm/s or half equilibrium time = 35 ± 15 min) and TetR shows a very high affinity for the molecule. The binding constant of TetR to [tc-Mg+] is Ka ~ 10^9 M^-1.  When bound to tc, TetR has a low binding level to DNA of 10^5 M^-1. [http://content.febsjournal.org/cgi/content/abstract/270/15/3109]
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===Modeling===
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Brown iGEM 2006 Matlab model code [[Media:tristable2006.txt]]
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[http://parts2.mit.edu/wiki/index.php/Table_of_preliminary_model_constants Initial Table of Constants]
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[http://parts2.mit.edu/wiki/index.php/Derivation_of_the_Model_Equations Derivation of Model Equations]
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A simpler model based on the bistable paper was developed that takes the relative transcription/translation rates into account [[Media:tristable2007.txt]].
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Latest revision as of 03:55, 27 October 2007

Brown University
Home Lead Sensor Tristable Switch Community Supplemental About us

Cellular Lead Sensor

About 40% of the world does not have access to clean water. Lead is a major contaminant worldwide. In the US alone, over 1 million children ages 1 through 5 have elevated levels of lead in their blood. Current lead detection systems are expensive and require lab analysis. Home lead testing kits are inaccurate and only detect lead at very high levels. We have created a genetic circuit in E Coli that responds to lead. The promoter and lead binding protein we use are ten times more selective for lead than for other similar heavy metals. We have also incorporated a genetic amplifier into our circuit to allow us to detect fairly low concentrations of lead.

Tristable Switch

The Tri-Stable Toggle Switch represents a continuation on the theme of the Toggle Switch begun by Gardner, et al to produce stable outputs in response to transient inputs. Applications such as a memory circuit and a drug delivery system are a few suggestions, but perhaps the most promising innovation lies in the design process. Our novel approach to the Tri-Stable Switch development is founded on quantitative principles, pioneering a technique to remove the guesswork from designing and debugging biological systems.

http://www.brownigem.com/images/pfizer.gif http://www.mandel.ca/suppliers/images/logo_labnet.gif http://brownigem.com/images/sponsors/nanodrop_small.jpg

  Links
Brown iGem on Openwetware
Brown iGem Official Website
Check out our Videos!
 News Updates
09.06.07
Brown University students love the Brown iGEM Booth at the Student Activities Fair!

09.04.07

Brown iGEM visits Pfizer HQ in Groton, Conneticut

08.28.07

Nanodrop donates ND-1000 Spectrophotometer and ND-3300 Fluorospectrophotometer to Brown iGEM

08.22.07

Brown iGEM Team presents its August Update to faculty and friends

08.20.07

Nanodrop it like it's hot! http://youtube.com/watch?v=pKT4Zrjeh6s

08.18.07

iGEM 2007 Meetup. Teams from MIT, BU, McGill, and Cold Spring Harbor presented their progress to each other and discussed the future of Synthetic Biology.

08.04.07

Brown iGEM presents summer research at the Brown University UTRA (Undergraduate Research) Symposium

07.30.07

Deepa and Tito gave a presentation to Brown Summer students and discussed the applications of synthetic biology

07.15.07

Brown iGEM presents June Update to faculty and friends of iGEM

07.03.07

The iGEM Team met up with Jody Hall's Biotechnology class on July 3rd. Afterwards we toured the lab and discussed the ethics of synthetic biology.

06.15.07

Brown iGEM presents June Update to Pfizer Team and Brown community

06.07.07

$25,000 grant from Pfizer for equipment purchases

06.01.07

Labnet sponsors Brown iGEM with vortex mixers, a microcentrifuge, pipettes, and autopipettes.

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