http://2009.igem.org/wiki/index.php?title=Special:Contributions/Lydia&feed=atom&limit=50&target=Lydia&year=&month=2009.igem.org - User contributions [en]2024-03-28T18:30:38ZFrom 2009.igem.orgMediaWiki 1.16.5http://2009.igem.org/Team:UC_Davis/PhoR_ChvI1Team:UC Davis/PhoR ChvI12009-10-22T02:43:53Z<p>Lydia: </p>
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<div style="text-align: center;"><b style="color: rgb(0, 0, 0);">&nbsp;<br />
</b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
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style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><img<br />
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style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 78px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 81px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Parts"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a<br />
href="https://2009.igem.org/Team:UC_Davis/Contacts_References"><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"<br />
style="border: 0px solid ; width: 83px; height: 37px;"></a></big></big></big></big></span></b></div><br />
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style="width: 100%; height: 2px; background-color: rgb(255, 255, 255);"><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial;"><b><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"><big><big><span<br />
style="text-decoration: underline;"><small>PhoR--&gt;ChvI</small><br><br />
<br><br />
</span></big></big>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp;<br />
&nbsp;&nbsp;&nbsp; </span></b><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;">It<br />
has been hypothesized<br />
that due to the amino acid similarity of ChvI to PhoB, ChvI is able to<br />
act as a<br />
phosphate regulator in <i>E.coli</i> as a PhoB substitute (3).</span><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"><o:p></o:p></span></p><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial;"><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;">&nbsp;&nbsp;&nbsp;<br />
<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; </span>Since<br />
ChvI showed no abnormal activity<br />
inside <i>Agrobacterium tumefaciens</i> under phosphate<br />
limitation, it<br />
cannot serve as the primary phosphate regulator in <i>A. tumefaciens</i><br />
(3).<o:p></o:p></span></p><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial;"><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"><span<br />
style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
</span><i>Note: ChvI was not required for phosphate regulation<br />
in A. tumefaciens (3), but it might be involved in phosphate<br />
regulation in A. tumefaciens in some way (3). Therefore, the<br />
possibility of crosstalk still exists</i><o:p></o:p></span></p><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial;"><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;">&nbsp;&nbsp;<span<br />
style="">&nbsp;&nbsp; </span>&nbsp;Due to the structural similarity<br />
of<br />
ChvI and PhoB, the possibility of crosstalk exists (3).<o:p></o:p></span></p><br />
<p class="MsoNormal"><i><span<br />
style="font-size: 13pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;">&nbsp;&nbsp;&nbsp;<br />
Note: Since<br />
we are going to use E. coli as our model chassis for this<br />
project; we<br />
had to make sure that our proteins will keep their characteristics of<br />
our<br />
interest after being placed inside the E. coli.</span></i><span<br />
style="font-size: 13pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"><br />
<i>For example, VirG gene (promoter 1) after being introduced<br />
into E. coli cells, started to behave as if it were a member of<br />
phosphate<br />
regulon (note: PhoB function was necessary for virG induction by<br />
phosphate<br />
starvation)(6).</i> <i>Studies have suggested that the reason for this<br />
behavior<br />
was due to the fact that virG promoter 1 has two hexamer blocks which<br />
have the<br />
same positions as the pho genes (6).</i> <o:p></o:p></span></p><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial;"><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"><span<br />
style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
</span>Studies have shown that ChvI probably activated Alkaline<br />
Phosphatase</span><em><span<br />
style="font-size: 13pt; font-family: &quot;Calibri&quot;,&quot;sans-serif&quot;;"> </span></em><em><span<br />
style="font-size: 13pt;">(AP)</span></em><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"><br />
activity by activating transcription of PhoA (PhoA also gets activated<br />
by PhoB)<br />
(3). It was not proven that ChvI activates other PhoB-dependent<br />
promoters or<br />
recognizes the same DNA sequence as PhoB (3). However, there has been<br />
speculation<br />
that since PhoR mediates PhoB phosphorylation and dephosphorylation, it<br />
may do<br />
the same to ChvI(3).<o:p></o:p></span></p><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial;"><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
Therefore, the best option would be to find a promoter<br />
from <i>A. tumefaciens</i> that gets activated by ChvG/ChvI, a<br />
two-component<br />
regulatory system only; and does not interact with any <i>E. coli</i><br />
system.</span><b><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"><br><br />
</span></b></p><br />
<hr style="width: 100%; height: 2px;"><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial;"><b><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"><o:p></o:p></span></b></p><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_Davis/wiring_pH_sensorTeam:UC Davis/wiring pH sensor2009-10-22T02:37:56Z<p>Lydia: </p>
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<title></title><br />
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<div style="text-align: center;"><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC3.png"<br />
style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/d/d1/UCDAVIS_PIC4.png"<br />
style="border: 0px solid ; width: 82px; height: 36px;"></big></big></big></big></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 78px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 81px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Parts"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a<br />
href="https://2009.igem.org/Team:UC_Davis/Contact1"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"<br />
style="border: 0px solid ; width: 83px; height: 37px;"></a></big></big></big></big></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span><br><br />
</b><br />
<hr style="width: 100%; height: 2px;"><br />
<div style="text-align: left;"><big><span<br />
style="font-weight: bold; text-decoration: underline;">Wiring the pH<br />
sensor into our system: <br><br />
<br><br />
Th pH sensor: </span></big><br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; ChvG/ChvI is a<br />
two-component system and a suspected pH sensor. ChvG is the histidine kinase and ChvI is its<br />
cognate response regulator(2). Together they are believed to sense<br />
extracellular pH between 5.5 to 7 and relay this signal to regulate<br />
the expression of key virulence-related genes(2).<br><br />
In our design, we used this two-component system from <i>Agrobacterium<br />
tumefaciens</i> to regulate the expression of select target genes.<br><br />
<hr style="width: 100%; height: 2px;"><big><span<br />
style="text-decoration: underline; font-weight: bold;">Possibility of<br />
Cross-Talk:</span></big><br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Bacteria have<br />
numerous related two-component systems based on kinase/response-regulation(5). For instance, the <i>A. tumefaciens</i><br />
genome is known to contain at least 25 two-component pathways<br />
(2). Therefore, there are possibilities of cross-talk between some genes<br />
present in <i>E. coli</i> that have the same mechanism as the ChvI/ChvG<br />
system. In fact, there is evidence supporting that ChvI can complement<br />
a PhoB deletion in <i>E. coli</i>. (3) Therefore it is highly likely<br />
that there is a possibility of cross-talk between our new sensory<br />
systems elements, present in <i>E. coli</i>.<br />
Many protein-protein interactions are not fully understood (5), which<br />
makes this project more challenging but very interesting.<br />
<br><br />
<hr style="width: 100%; height: 2px;"><br />
<div style="text-align: center;"><span style="font-style: italic;"></span><br><br />
<span style="font-style: italic;"></span></div><br />
<span style="font-style: italic;">Click on the specific part or<br />
interaction for more information.<br><br />
Parts: <a href="https://2009.igem.org/Team:UC_Davis/Project1/ChvG.html">ChvG</a>,<a<br />
href="https://2009.igem.org/Team:UC_Davis/ChvI1"> ChvI</a>,<a<br />
href="https://2009.igem.org/Team:UC_Davis/PhoR1"> PhoR</a>,<a<br />
href="https://2009.igem.org/Team:UC_Davis/PhoB1"> PhoB</a><br><br />
Interactions: <a href="https://2009.igem.org/Team:UC_Davis/ChvG_ChvI">ChvG---&gt;<br />
ChvI</a> , <a href="https://2009.igem.org/Team:UC_Davis/PhoR_ChvI1">PhoR---&gt;<br />
ChvI </a>, <a href="https://2009.igem.org/Team:UC_Davis/PhoR_PhoB">PhoR---&gt;PhoB</a><br />
,<a href="https://2009.igem.org/Team:UC_Davis/ChvI_PhoA">ChvI---&gt;PhoA<br><br />
</a><br><br />
</span><big><span style="font-weight: bold; text-decoration: underline;"><br />
</span></big><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; text-indent: 0.5in; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial;"><b><span<br />
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"><o:p></o:p></span></b></p><br />
</div><br />
</div><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_Davis/ChvG_ChvITeam:UC Davis/ChvG ChvI2009-10-22T02:32:04Z<p>Lydia: </p>
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<head><br />
<meta content="text/html;charset=ISO-8859-1" http-equiv="Content-Type"><br />
<title>chvG_chvI</title><br />
</head><br />
<body style="background-color: rgb(255, 255, 255); color: rgb(0, 0, 0);"<br />
alink="#ee0000" link="#0000ee" vlink="#551a8b"><br />
<div style="text-align: center;"><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC3.png"<br />
style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/d/d1/UCDAVIS_PIC4.png"<br />
style="border: 0px solid ; width: 82px; height: 36px;"></big></big></big></big></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 78px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 81px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Parts"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a<br />
href="https://2009.igem.org/Team:UC_Davis/Contacts_References"><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"<br />
style="border: 0px solid ; width: 83px; height: 37px;"></a></big></big></big></big></span></b></div><br />
<hr<br />
style="width: 100%; height: 2px; background-color: rgb(255, 255, 255); color: rgb(0, 0, 0);"><b><span<br />
style="font-size: 14pt; line-height: 115%;"><span style=""></span></span></b><b><u><span<br />
style="font-size: 16pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">ChvG<br />
---&gt; ChvI<o:p></o:p></span></u></b><br />
<p class="MsoNormal"<br />
style="margin-bottom: 0.0001pt; line-height: normal;"><b><span<br />
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"><span<br />
style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; </span><o:p></o:p><span<br />
style=""></span></span></b><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;">ChvG<br />
and ChvI is the sensor kinase and cognate response regulator,<br />
respectively, of<br />
the two-component regulatory system in <i>Agrobacterium tumefaciens</i> thought to<br />
respond to<br />
extracellular pH changes(2).</span><b><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"><br><br />
</span></b></p><br />
<hr style="width: 100%; height: 2px;"><br />
<p class="MsoNormal"<br />
style="margin-bottom: 0.0001pt; line-height: normal;"><b><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"><br><br />
</span></b></p><br />
<p class="MsoNormal"<br />
style="margin-bottom: 0.0001pt; line-height: normal;"><br><br />
<b><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"></span></b><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span></p><br />
<b><span style="font-size: 14pt; line-height: 115%;"><span style=""><big><big><br />
<br><br />
</big></big></span></span></b><br><br />
<p style="background-color: rgb(255, 255, 255);" class="MsoNormal"><span<br />
style=""></span><b><span style="font-size: 12pt; line-height: 115%;"><o:p></o:p></span></b></p><br />
<p style="background-color: rgb(255, 255, 255);" class="MsoNormal"><b><span<br />
style="font-size: 12pt; line-height: 115%;"> <o:p></o:p></span></b></p><br />
<big style="color: rgb(0, 0, 0);"><big><big><big<br />
style="font-weight: bold;"><span style=""></span></big><big<br />
style="font-weight: bold;"><span style=""><small><span<br />
style="text-decoration: underline;"></span></small></span></big></big></big></big><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_Davis/ChvI1Team:UC Davis/ChvI12009-10-22T02:30:56Z<p>Lydia: </p>
<hr />
<div><html><br />
<head><br />
<meta content="text/html;charset=ISO-8859-1" http-equiv="Content-Type"><br />
<title>ChvI</title><br />
</head><br />
<body style="background-color: rgb(255, 255, 255); color: rgb(0, 0, 0);"<br />
alink="#ee0000" link="#0000ee" vlink="#551a8b"><br />
<div style="text-align: center;"><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC3.png"<br />
style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/d/d1/UCDAVIS_PIC4.png"<br />
style="border: 0px solid ; width: 82px; height: 36px;"></big></big></big></big></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 78px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 81px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Parts"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a<br />
href="https://2009.igem.org/Team:UC_Davis/Contacts_References"><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"<br />
style="border: 0px solid ; width: 83px; height: 37px;"></a></big></big></big></big></span></b></div><br />
<hr<br />
style="width: 100%; height: 2px; color: rgb(255, 255, 0); background-color: rgb(255, 255, 255);"><br />
<div<br />
style="text-align: center; color: rgb(255, 255, 0); background-color: rgb(255, 255, 255);"><br />
<p style="color: rgb(0, 0, 0); text-align: left;" class="MsoNormal"><b><u><span<br />
style="font-size: 18pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">ChvI:<o:p></o:p></span></u></b></p><br />
<p class="MsoNormal"<br />
style="line-height: normal; color: rgb(0, 0, 0); text-align: left;"><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
</span></span><b><span<br />
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="">&nbsp;</span></span></b><span<br />
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><small>ChvI<br />
is the response regulator of the<br />
ChvG/ChvI two-component regulatory system found in <i>Agrobacterium tumefaciens</i><br />
(4).<br><br />
<small><span style="font-style: italic;">For more information go to: <a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K265004">ChvI</a></span></small><br><br />
</small></span></p><br />
<hr style="width: 100%; height: 2px;"><br />
<p class="MsoNormal"<br />
style="line-height: normal; color: rgb(0, 0, 0); text-align: left;"><span<br />
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span></p><br />
<big><big><big><big><span<br />
style="text-decoration: underline; font-weight: bold;"><span<br />
style="color: rgb(0, 0, 0);"></span></span></big></big></big></big><big><big><big><big><span<br />
style="text-decoration: underline; font-weight: bold;"><br><br />
</span></big></big></big></big></div><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_Davis/Project1/ChvG.htmlTeam:UC Davis/Project1/ChvG.html2009-10-22T02:29:14Z<p>Lydia: </p>
<hr />
<div><html><br />
<head><br />
<meta content="text/html;charset=ISO-8859-1" http-equiv="Content-Type"><br />
<title>ChvG</title><br />
</head><br />
<body style="background-color: rgb(255, 255, 255); color: rgb(0, 0, 0);"<br />
alink="#ee0000" link="#0000ee" vlink="#551a8b"><br />
<div style="text-align: center;"><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC3.png"<br />
style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/d/d1/UCDAVIS_PIC4.png"<br />
style="border: 0px solid ; width: 82px; height: 36px;"></big></big></big></big></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 78px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 81px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Parts"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a<br />
href="https://2009.igem.org/Team:UC_Davis/Contacts_References"><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"<br />
style="border: 0px solid ; width: 83px; height: 37px;"></a><br><br />
</big></big></big></big></span></b><br />
<hr style="width: 100%; height: 2px;"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b></div><br />
<small> </small><big><big><big><big><span<br />
style="text-decoration: underline; font-weight: bold;"><small>ChvG</small></span></big></big></big></big><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"><br><br />
<br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; &nbsp;&nbsp; ChvG is the sensor<br />
kinase of a two-component regulatory system found in<br />
<i>Agrobacterium tumefaciens</i> (2).<o:p></o:p></span><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial;"><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"><span<br />
style="">&nbsp;&nbsp;&nbsp; &nbsp; &nbsp; &nbsp;&nbsp; </span>Acidic<br />
conditions play an important role in <i>Agrobacterium</i>-plant interactions<br />
(1).</span><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">Numerous<br />
studies<br />
have investigated the possible connection between the physiological<br />
response to<br />
acidic conditions and the activity of ChvG and ChvI<span<br />
style="color: black;">.<br />
Studies have shown that </span>ChvG/ChvI two-component systems<br />
specifically get<br />
induced in an acidic environment(1).<span style="color: black;"><o:p></o:p></span></span></p><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial;"><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"><span<br />
style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
</span>As<br />
a result, there is growing consensus that the ChvG/ChvI two-component<br />
regulatory system may be the primary global pH sensor in <i>A.tumefaciens</i> and as such, serves in coordinating the expression of<br />
acid-induced genes (2).<br><br />
<span style="font-style: italic;">More information :<a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K265005">ChvG</a></span><br><br />
</span></p><br />
<hr style="width: 100%; height: 2px;"><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial;"><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"><o:p></o:p></span></p><br />
<br><br />
<div style="text-align: center;"><big><big><big><big><span<br />
style="text-decoration: underline; font-weight: bold;"></span></big></big></big></big><br><br />
<big><big><big><big><span<br />
style="text-decoration: underline; font-weight: bold;"></span></big></big></big></big></div><br />
<big><big><big><big><span<br />
style="text-decoration: underline; font-weight: bold;"><br><br />
<br><br />
<br><br />
</span></big></big></big></big><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_Davis/Project1/Finding_ph_sensorTeam:UC Davis/Project1/Finding ph sensor2009-10-22T02:26:01Z<p>Lydia: </p>
<hr />
<div><html><br />
<head><br />
<title></title><br />
</head><br />
<body><br />
<div style="text-align: center;"><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC3.png"<br />
style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/d/d1/UCDAVIS_PIC4.png"<br />
style="border: 0px solid ; width: 82px; height: 36px;"></big></big></big></big></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 78px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 81px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Parts"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a<br />
href="https://2009.igem.org/Team:UC_Davis/Contacts_References"><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"<br />
style="border: 0px solid ; width: 83px; height: 37px;"></a></big></big></big></big></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span><br><br />
</b><br />
<hr style="width: 100%; height: 2px;"><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial; text-align: left; text-decoration: underline;"><big><big><b><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;">Finding<br />
an appropriate biological pH sensor:</span></b></big></big></p><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial; text-align: left;"><b><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;">Why<br />
is there a need for a pH sensor?<o:p></o:p></span></b></p><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial; text-align: left;"><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;We<br />
would like our probiotic organism to confine itself to the stomach.<br />
Since low<br />
pH is a hallmark of the stomach, it seems reasonable to assume that<br />
linking the<br />
production of a suicide gene to a dramatic increase in pH (to values<br />
found in<br />
the intestine) should allow us to sequester our organism to the<br />
stomach. There<br />
are no pH sensors available on the registry of biological parts. So, we<br />
will<br />
engineer a device to sense extracellular pH changes and design it so<br />
that it<br />
can be used to regulate the expression of any target gene in<br />
confinement by pH<br />
levels, and then add it to the registry!<o:p></o:p></span></p><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial; text-align: left;"><b><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;">Finding<br />
an appropriate biological pH sensor:</span></b><b><span<br />
style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span></b></p><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial; text-align: left;"><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
We have decided to design our pH sensor by adopting one of the pH<br />
sensors of <i>Agrobacterium<br />
tumefaciens</i>. The ChvG/ChvI two-component system of <i>A.tumefaciens</i><br />
is<br />
suspected to sense extracellular pH changes and relay this<br />
environmental<br />
signal to numerous virulence-related promoters(2). <o:p></o:p></span></p><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial; text-align: left;"><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
<i>For more information regarding </i></span><span<br />
style="font-style: italic;">ChvG/ChvI two component system</span><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"><i>,<br />
go to <a href="https://2009.igem.org/Team:UC_Davis/Project1/ChvG.html">ChvG</a>,<br />
<a href="https://2009.igem.org/Team:UC_Davis/ChvI1">ChvI</a>, <a<br />
href="https://2009.igem.org/Team:UC_Davis/ChvG_ChvI">ChvG---&gt;ChvI</a><br><br />
</i></span></p><br />
<hr style="width: 100%; height: 2px;"><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial; text-align: left;"><br><br />
<span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"></span><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><a<br />
href="https://2009.igem.org/Team:UC_Davis/ChvG_ChvI"><i><span<br />
style="color: blue;"></span></i></a><o:p></o:p></span></p><br />
<p class="MsoNormal"<br />
style="background: white none repeat scroll 0% 50%; line-height: normal; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial; text-align: left;"><i><u><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: blue;"></span></u></i><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;"><o:p></o:p></span></p><br />
</div><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_Davis/assay_for_measuring_protein_localization_and_activityTeam:UC Davis/assay for measuring protein localization and activity2009-10-22T02:23:59Z<p>Lydia: </p>
<hr />
<div><html><br />
<head><br />
<title></title><br />
</head><br />
<body><br />
<div style="text-align: center;"><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC3.png"<br />
style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/d/d1/UCDAVIS_PIC4.png"<br />
style="border: 0px solid ; width: 82px; height: 36px;"></big></big></big></big></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 78px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 81px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Parts"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a<br />
href="https://2009.igem.org/Team:UC_Davis/Contact1"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"<br />
style="border: 0px solid ; width: 83px; height: 37px;"></a></big></big></big></big></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><br><br />
</div><br />
<hr style="width: 100%; height: 2px;"><br />
<div style="text-align: center;"><span<br />
style="font-weight: bold; text-decoration: underline;"><big>Assay for<br />
measuring protein localization and activity:</big><br><br />
</span><br />
<div style="text-align: left;">&nbsp;&nbsp;&nbsp; &nbsp;&nbsp; <br><br />
&nbsp;&nbsp;&nbsp;&nbsp; We are using western blotting as our assay to<br />
measure protein localization and activity, where we have tagged<br />
all of our constructs with a 6xHIS tag.&nbsp; This allows us to compare<br />
basal GFP or Luciferase activity inside (basal) and outside of the cell<br />
(medium).&nbsp; <br><br />
</div><br />
<div style="text-align: left;">&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; We<br />
may also use fluorescence as an assay by comparing the<br />
brightness of the cell when it keeps it inside the cell, and when it<br />
secretes it into the medium.&nbsp; Both of our test genes, GFP and<br />
Luciferase, are fluorescence proteins.<br><br />
<hr style="width: 100%; height: 2px;"><span<br />
style="font-weight: bold; text-decoration: underline;"></span><br><br />
</div><br />
<div style="text-align: left;"><span<br />
style="font-weight: bold; text-decoration: underline;"></span></div><br />
<span style="font-weight: bold; text-decoration: underline;"><br><br />
</span></div><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_Davis/PartsTeam:UC Davis/Parts2009-10-22T02:21:51Z<p>Lydia: </p>
<hr />
<div><html><br />
<head><br />
<meta content="text/html;charset=ISO-8859-1" http-equiv="Content-Type"><br />
<title>PARTS1</title><br />
</head><br />
<body><br />
<div style="text-align: center;"><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC3.png"<br />
style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/d/d1/UCDAVIS_PIC4.png"<br />
style="border: 0px solid ; width: 82px; height: 36px;"></big></big></big></big></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 78px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 81px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Parts"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a<br />
href="https://2009.igem.org/Team:UC_Davis/Contacts_References"><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"<br />
style="border: 0px solid ; width: 83px; height: 37px;"></a></big></big></big></big></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><small><br />
</small></big></big></big></big></span></b><br />
<hr style="width: 100%; height: 2px;"><br />
<div style="text-align: left;"><big><big><b<br />
style="color: rgb(0, 0, 0); text-decoration: underline;">Parts:</b></big></big><b<br />
style="color: rgb(0, 0, 0);">&nbsp;&nbsp;&nbsp; <br><br />
Parts related to<br />
secretion:&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
&nbsp; &nbsp; &nbsp;&nbsp;&nbsp; Parts related to pH<br />
sensor:<br><br />
</b><br />
<table<br />
style="text-align: left; width: 100%; background-color: rgb(255, 255, 0);"<br />
border="2" cellpadding="2" cellspacing="2"><br />
<tbody><br />
<tr><br />
<td style="vertical-align: top;">Proteins:<br><br />
</td><br />
<td style="vertical-align: top;">Promoters:<br><br />
</td><br />
<td style="vertical-align: top;">Others:<br><br />
</td><br />
<td style="vertical-align: top;">&nbsp;New parts:<br><br />
</td><br />
<td style="vertical-align: top;">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
Promoters:<br><br />
</td><br />
<td style="vertical-align: top;">Proteins:</td><br />
</tr><br />
<tr><br />
<td style="vertical-align: top;"><br />
<ul><br />
<li><a href="#INPNC">INPNC</a></li><br />
<li><a href="#OmpA">OmpA</a></li><br />
<li><a href="#Luciferase">Luciferase</a></li><br />
<li><a href="#GFP">GFP</a></li><br />
</ul><br />
</td><br />
<td style="vertical-align: top;"><br />
<ul><br />
<li><a href="#LacI">LacI</a></li><br />
</ul><br />
</td><br />
<td style="vertical-align: top;"><br />
<ul><br />
<li><a href="#SS">SS:signal sequence</a></li><br />
<li><a href="#His">6-His-Tag</a></li><br />
<li><a href="#RBS">RBS</a></li><br />
<li><a href="#Terminator">Terminator</a></li><br />
</ul><br />
</td><br />
<td style="vertical-align: top;"><br />
<ul><br />
<li><a href="#INPNCSS">INPNC + SS<br><br />
</a></li><br />
<li><a href="#OmpAss">OmpA + SS<br><br />
</a></li><br />
<li><a href="#INPNC">INPNC </a><br><br />
</li><br />
<li><a href="#SS">SS</a><br><br />
</li><br />
</ul><br />
</td><br />
<td style="vertical-align: top;"><br />
<ul><br />
<li><a href="#ChvI_promoter">ChvI promoter</a></li><br />
<li><a href="#katA">KatA promoter</a></li><br />
<li><a href="#aopB">AopB promoter</a></li><br />
<li><a href="#PhoA">PhoA<br />
promoter</a>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
&nbsp;&nbsp; </li><br />
<li>I<a href="#impA">mpA promoter</a></li><br />
</ul><br />
</td><br />
<td style="vertical-align: top;"><br />
<ul><br />
<li><a href="https://2009.igem.org/Team:UC_Davis/ChvI1">ChvI</a></li><br />
<li><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project1/ChvG.html">ChvG</a></li><br />
</ul><br />
</td><br />
</tr><br />
</tbody><br />
</table><br />
<div style="text-align: center;"><b style="color: rgb(0, 0, 0);">&nbsp;<br />
<img style="width: 493px; height: 242px;" alt=""<br />
src="https://static.igem.org/mediawiki/2009/5/5e/UCDAVIS_picture1.jpg"></b><br><br />
</div><br />
<hr style="width: 100%; height: 2px;"><br />
<p class="MsoNormal"<br />
style="line-height: normal; font-weight: bold; text-align: center; text-decoration: underline;"><big><big>New<br />
parts:</big></big>&nbsp;</p><br />
<span style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span><br />
<p class="freeform0" style="margin-bottom: 6pt; line-height: 19pt;"><a<br />
name="INPNC"></a><b>INPNC:</b> The ice-nucleation protein (INP) from <i>Pseudomonas<br />
syringae</i> is used by its natural host to nucleate ice formation and<br />
is implicated in<i> P.syringae</i>-associated pathogenesis<i>.&nbsp; </i>INP<br />
and<br />
a truncated derivative lacking the central domain (INPNC) have been<br />
used<br />
extensively for displaying proteins on the surface of <i>E. coli (7)</i>.&nbsp;<br />
For instance, AldO and PhaZ1 have been successfully displayed on the<br />
surface of<br />
<i>E.coli </i>using INPNC (7, 15). <br><br />
<u1:p></u1:p>Park <i>et al.</i> have shown that when INPNC is fused to<br />
the <i>phaZ1</i><br />
gene and its signal sequence, it can serve as a suitable surface<br />
delivery<br />
and secretion device of the otherwise toxic <i>phaZ1</i> gene product(15).&nbsp;<br />
<u1:p></u1:p>This part was synthesized by Mr. Gene (Regensburg,<br />
Germany) with<br />
codon optimization and subsequently transferred into vector (<span<br />
style="background: rgb(255, 255, 51) none repeat scroll 0% 50%; color: black; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial;"></span><br />
<a href="http://partsregistry.org/Part:pSB1AK3">pSB1AK3</a>). As it is<br />
expected that this part will be used in the context of the<br />
fusion<br />
protein, the prefix and suffix for this part are consistent with the <i>BBF<br />
RCF-12</i> standard.&nbsp; <br><br />
<u1:p></u1:p>We have proposed to build and test a general protein<br />
secretion<br />
system modeled after that developed by Park <i>et al. </i>in which a<br />
fusion of<br />
INPNC and the signal sequence from the <i>phaZ1</i> gene are used to<br />
secrete<br />
any target protein.&nbsp; <br><br />
<u1:p></u1:p><i>We have modified this protein to be consistent with the BBF<br />
RFC-12<br />
Standard. We have submitted this part to the parts registry as part </i><a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K265008"><i><span<br />
style="color: rgb(0, 0, 153);">BBa_K265008</span></i></a><i>.<br><br />
</i></p><br />
<p class="freeform0" style="margin-bottom: 6pt; line-height: 19pt;"><br><br />
<i><u1:p></u1:p></i><o:p></o:p></p><br />
<p class="freeform0" style="margin-bottom: 6pt; line-height: 19pt;"><a<br />
name="SS"></a><b><u1:p></u1:p>SS:</b> The<br />
signal sequence (SS) for the <i>phaZ1 </i>gene product of <i>Paucimonas<br />
lemoignei</i>, a polyhydroxybutyrate depolymerase (15).&nbsp; In the<br />
native<br />
protein the signal sequence is cleaved between residues Ala37 and<br />
Leu38.&nbsp;<br />
Park <i>et al. </i>have showed that the fusion of the complete <i>phaZ1<br />
</i>gene<br />
(including SS) and a truncated ice nucleation protein from <i>Pseudomonas<br />
syringae</i> (<a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K265008"><i><span<br />
style="color: rgb(0, 0, 153);">BBa_K265008</span></i></a>), could lead<br />
to stable<br />
expression and secretion of the <i>phaZ1</i> gene product.&nbsp; <br><br />
<u1:p></u1:p>We propose that the signal sequence might be generally<br />
useful as a<br />
cleavage tag in secretion systems that include a membrane anchor<br />
component,<br />
such as INPNC (<a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K265008"><i><span<br />
style="color: rgb(0, 0, 153);">BBa_K265008</span></i></a>) or OmpA (<i><a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K103006"><span<br />
style="color: blue;">BBa_K103006</span></a>).<span<br />
style="color: rgb(0, 41, 57);"> </span></i>The<br />
proposed constructs would consists of a membrane anchor (INPNC or OmpA)<br />
followed by the cleavable signal sequence and finally a target protein<br />
marked<br />
for secretion.&nbsp; <br><br />
<u1:p></u1:p><i>Since we expect that this part will be used in the<br />
context of a<br />
fusion protein, we have modified this protein to be consistent with BBF<br />
RFC-12<br />
Standard. We have submitted this part to the part registry as part </i><a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K265002"><i><span<br />
style="color: rgb(0, 0, 153);">BBa_K265002</span></i></a>.<br><br />
<br><br />
</p><br />
<u1:p></u1:p><br />
<p class="freeform0" style="margin-bottom: 6pt; line-height: 19pt;"><a<br />
name="INPNCSS"></a><b>INPNC+SS:</b> Park <i>et al. </i>have showed<br />
that the<br />
fusion of the complete <i>phaZ1 </i>gene (including SS) and a<br />
truncated ice<br />
nucleation protein from <i>Pseudomonas syringae</i> (<a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K265008"><i><span<br />
style="color: rgb(0, 0, 153);">BBa_K265008</span></i></a>), could lead<br />
to stable<br />
expression and secretion of the <i>phaZ1</i> gene product (15).<br><br />
<u1:p></u1:p>We propose that this system might be generally useful for<br />
the<br />
secretion of other target proteins in <i>E. coli</i> and have<br />
therefore created<br />
a fusion of parts <a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K265008"><i><span<br />
style="color: rgb(0, 0, 153);">BBa_K265008</span></i></a> (INPNC) and <a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K265002"><i><span<br />
style="color: rgb(0, 0, 153);">BBa_K265002</span></i></a> (SS) which<br />
is compatible with<br />
the <i>BBF RFC-12 Standard. <u1:p></u1:p></i><br><br />
During the construction of this part, two silent mutations were<br />
introduced in<br />
the coding region of INPNC (T324A and A348T) that differ from those in<br />
part <a href="http://partsregistry.org/wiki/index.php/Part:BBa_K265008"><i><span<br />
style="color: rgb(0, 0, 153);">BBa_K265008</span></i></a>.&nbsp; <br><br />
<u1:p></u1:p><i>We have submitted this part to the part registry in the<br />
BBF<br />
RFC-12 Standard </i>as part<i> </i><a<br />
href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K265009"><i><span<br />
style="color: rgb(0, 0, 153);">BBa_K265009</span></i></a>.<br><br />
<br><br />
</p><br />
<u1:p></u1:p><br />
<p class="freeform0" style="margin-bottom: 6pt; line-height: 19pt;"><a<br />
name="OmpAss"></a><b>OmpA+SS:</b> Since OmpA is believed to function<br />
similarly<br />
to INPNC and Park <i>et al. </i>have showed that the fusion of the<br />
complete <i>phaZ1<br />
</i>gene (including SS) and a truncated ice nucleation protein from <i>Pseudomonas<br />
syringae</i> (<a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K265008"><i><span<br />
style="color: rgb(0, 0, 153);">BBa_K265008</span></i></a>), could lead<br />
to stable<br />
expression and secretion of the <i>phaZ1</i> gene product (15), we<br />
have decided<br />
to test and see if OmpA's ability to secret increases when it is used<br />
with a<br />
signal sequence.<br><br />
<u1:p></u1:p><i>We have modified this protein to be consistent with BBF<br />
RFC-12<br />
Standard and have submitted this part to the part registry, </i><a<br />
href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K265011"><i><span<br />
style="color: rgb(0, 0, 153);">BBa_K265011</span></i><u><span<br />
style="color: blue;">.</span></u></a></p><br />
<u1:p></u1:p><br />
<div class="MsoNormal" style="text-align: center;" align="center"><br />
<hr align="center" size="2" width="100%"></div><br />
<p class="MsoNormal"><a name="OmpA"></a><b>OmpA</b>: OmpA is one of the<br />
proteins on<br />
the outer membrane of <i>E. coli</i> (13),it is used as a displaying<br />
fusion<br />
protein on the cell surface . This part has already been documented on<br />
the parts<br />
registry; however, it has not been tested as a compnent of secretion<br />
system<br />
(via fusion with a target protein linked with a cleavable signal<br />
sequence) <i><br><br />
<u1:p></u1:p>We have modified this protein to be consistent with BBF<br />
RFC-12<br />
Standard.<br><br />
Note: “It has remained essentially unknown how proteins of E. coli<br />
outer<br />
membrane are sorted and incorporated into this membrane” (10)</i> <i><br><br />
For more information go to:<a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K103006"><u><span<br />
style="color: blue;"> BBa_K103006</span></u></a></i> </p><br />
<u1:p></u1:p><br />
<div class="MsoNormal" style="text-align: center;" align="center"><br />
<hr align="center" size="2" width="100%"></div><br />
<p class="MsoNormal" style=""><a name="RBS"></a><b>RBS</b>:&nbsp;<br />
Ribosome Binding site number 32 (BBa_J61132)<br />
from the registry is being used in our secretion system. <br><br />
<i>For more information go to:</i> <a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_J61132"><i><u><span<br />
style="color: blue;">BBa_J61132</span></u></i></a></p><br />
<u1:p></u1:p><br />
<div class="MsoNormal" style="text-align: center;" align="center"><br />
<hr align="center" size="2" width="100%"></div><br />
<p class="MsoNormal" style=""><a name="Terminator"></a><b>Terminator</b>:<br />
We are using BBa_B0015, a double<br />
terminator, as our terminator in both our secretion and pH system.<br><br />
<i>For more information go to:</i> <a<br />
href="http://partsregistry.org/wiki/index.php?title=Part:BBa_B0015"><i><u><span<br />
style="color: blue;">BBa_B0015</span></u></i></a> </p><br />
<u1:p></u1:p><br />
<div class="MsoNormal" style="text-align: center;" align="center"><br />
<hr align="center" size="2" width="100%"></div><br />
<p class="MsoNormal" style=""><a name="GFP"></a><b>GFP</b> <b>(Green<br />
Fluorescent Protein)</b>: Mutant of GFP<br />
known to be very stable (superfolder), which will let this protein fold<br />
quickly<br />
so we can use either a fluorescent reader or UV light to detect it.<br />
Therefore<br />
it has been used as a reporter in our secretion system. It also serves<br />
as a<br />
small protein in testing our secretion system.<br><br />
<i>For more informaiton go to: </i><a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K265003"><i><u><span<br />
style="color: blue;">BBa_K265003</span></u></i></a><i>&nbsp;</i></p><br />
<u1:p></u1:p><br />
<div class="MsoNormal" style="text-align: center;" align="center"><br />
<hr align="center" size="2" width="100%"></div><br />
<p class="MsoNormal" style=""><a name="Luciferase"></a><b>Luciferase</b>:<br />
Luciferase is a firefly protein that<br />
also fluoresces, so it serves as a reporter as well as a testable large<br />
protein.<br><br />
<i>For more information go to: <a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_I712019"><u><span<br />
style="color: blue;">BBa_1712019</span></u></a></i> </p><br />
<u1:p></u1:p><br />
<div class="MsoNormal" style="text-align: center;" align="center"><br />
<hr align="center" size="2" width="100%"></div><br />
<p class="MsoNormal" style=""><a name="LacI"></a><b>LacI</b>: An<br />
inducible promoter that was found in the part<br />
registry.<br><br />
<i>For more information go to:<a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_R0010"><u><span<br />
style="color: blue;"> BBa_R0010</span></u></a><br style=""><br />
<!--[endif]--></i></p><br />
<u1:p></u1:p><br />
<div class="MsoNormal" style="text-align: center;" align="center"><br />
<hr align="center" size="2" width="100%"></div><br />
<p class="MsoNormal" style=""><a name="His"></a><b>6-His Tag</b>: The<br />
6-Histidine Tag serves as a tag for Western<br />
Blotting if our fluorescent reporters are not expressed as highly as we<br />
would like. <br><br />
<i>Note: We are using this tag as an additional method for assay beside fluorescence of GFP and Luciferase.</i> </p><div class="MsoNormal" style="text-align: center;" align="center"><br />
<hr align="center" size="2" width="100%"></div><br />
<br><br />
more information go to: </i><a<br />
href="http://partsregistry.org/cgi/partsdb/pgroup.cgi?pgroup=iGEM2009&amp;group=UC_Davis"><i><u><span<br />
style="color: blue;">UCDAVIS_Parts</span></u></i></a> </p><br />
<u1:p></u1:p><br />
<hr style="width: 100%; height: 2px;"><small style="font-weight: bold;"><span<br />
style="font-size: 18pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></small></div><br />
</div><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_Davis/ProjectTeam:UC Davis/Project2009-10-22T02:07:17Z<p>Lydia: </p>
<hr />
<div><html><br />
<head><br />
<meta content="text/html;charset=ISO-8859-1" http-equiv="Content-Type"><br />
<title>lol</title><br />
</head><br />
<body><br />
<div style="text-align: center;"><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span>&nbsp;<br />
</b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC3.png"<br />
style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/d/d1/UCDAVIS_PIC4.png"<br />
style="border: 0px solid ; width: 82px; height: 36px;"></big></big></big></big></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 78px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 81px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Parts"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a<br />
href="https://2009.igem.org/Team:UC_Davis/Contacts_References"><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"<br />
style="border: 0px solid ; width: 83px; height: 37px;"></a></big></big></big></big></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><small><br />
</small></big></big></big></big></span></b><b<br />
style="color: rgb(0, 0, 0);"> </b><br />
<hr style="width: 100%; height: 2px;"><br />
<div style="text-align: left;"><b style="color: rgb(0, 0, 0);"><big<br />
style="text-decoration: underline;"><big>Project:</big></big><br><br />
</b><br />
<div style="text-align: center;"><b style="color: rgb(0, 0, 0);"><a<br />
href="#celiac_disease">Celiac<br />
Disease</a>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
<a href="#current_treatment">Current<br />
treatment</a>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<a<br />
href="#our_approach"><br />
Our Approach</a></b><br><br />
</div><br />
<small><u><span<br />
style="font-size: 20pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="font-weight: bold;"><a name="celiac_disease"></a></span></span></u></small><span<br />
style=""><big><big style="text-decoration: underline;"><span<br />
style="font-weight: bold;">What is Celiac Disease?</span></big></big><o:p></o:p></span><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Celiac Disease is an autoimmune<br />
disorder that occurs inside the small<br />
intestine. When the body cannot properly digest gliadin (a component of gluten),<br />
this leads to an immune response on the surface of the small intestine<br />
(14).<br />
Different people have varying degrees of immune<br />
response<br />
such as bloating, diarrhea, and weight loss (14). An estimated one out of 133 Americans currently suffer from this<br />
disease (11). Considering the estimated 3.08 million people in America, this isn't a minor issue. In fact, it was one of our teammate's friends suffering from celiac who initially sparked our interest in designing this project. <o:p></o:p><span style=""><small><span<br />
style="font-style: italic;"><br><br />
</span></small></span></p><br />
<p><span style=""><big style="font-weight: bold;"><span<br />
style="font-style: italic;">What happens in the normal small<br />
intestine?</span></big><o:p></o:p></span><span<br />
style="font-weight: bold;"></span><br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; <br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; The inside surface of the small intestine is covered with small microvilli, constituting the brush border membrane. In the normal small intestine, the brush border membrane is where polysaccharides, proteins, and fat droplets are digested into smaller parts that then get absorbed into the bloodstream. <br><br />
</p><br />
<p><o:p></o:p><span style=""><big><span<br />
style="font-weight: bold; font-style: italic;">What happens in the small intestine of people with celiac<br />
disease?</span></big><o:p></o:p></span></p><br />
<span style="font-weight: bold;"></span><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; The brush border characteristic to people with celiac disease allows large molecules such as gliadin to go straight<br />
into the<br />
blood stream without directing them through the transcellular route.<br />
Once<br />
gliadin passes through, Antigen Presenting Cells (APCs) recognize<br />
gliadin as a<br />
foreign object and attack. This immune response results in the inflammation of<br />
the surface of the intestine with loss of the normal cells required for<br />
absorption of sugars, protein, and fat from the diet. <o:p></o:p></p><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; In addition to the array of symtpoms illustrated above, the immune response may also damage intestinal villi, which are important<br />
for<br />
absorbing nutrients. The destruction of villi eventually leads to a<br />
lack of<br />
absorption of different nutrients, which can lead to malnutrition (14).<br />
Some<br />
people affected with this disorder can also suffer from autoimmune<br />
thyroid<br />
disease, autoimmune liver disease, and rheumatoid arthritis (diseases<br />
in<br />
which body immune system attacks healthy cells/tissues) (14). <o:p></o:p></p><br />
<hr style="width: 100%; height: 2px;"><br />
<p><u><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><span<br />
style="font-weight: bold;"><a name="current_treatment"></a>Current<br />
treatments:</span></big></big><o:p></o:p></span></u><br><br />
<b><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span><br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Currently, no cure has been found<br />
for this illness. The only way to avoid<br />
the symptoms of celiac disease is by adopting a gluten-free diet<br />
(14, 12,<br />
and 11). However, you are highly likely to find gluten in your everyday<br />
diet (in foods such as grains), and<br />
people with celiac disease must purchase gluten-free substances from<br />
specialized grocery stores. "Oral supplementation with prolyl<br />
oligopeptidases that can digest and detoxify gluten has therefore been<br />
proposed<br />
as a potential therapeutic approach."(8) However, enzymes studied<br />
earlier<br />
were not able to degrade gluten inside the stomach (before it reaches the small<br />
intestine), because they were "irreversibly inactivated by pepsin and<br />
acidic pH, both present in the stomach."(8) <o:p></o:p></p><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Nevertheless, over the past<br />
years, researchers have discovered an enzyme--a newly-identified prolyl endoprotease--from <i>Aspergillus niger</i>, <br />
that was<br />
observed to "work optimally at 4-5pH and remains stable at 2pH"(8).<br />
Perhaps this enzyme will lead us to an alternative treatment for this<br />
disorder(8). Studies have shown that prolyl endoprotease from <i>A.niger</i> is able to "degrade gluten in vitro and under conditions<br />
similar<br />
to the ones present in the gastrointestinal tract." (8)&nbsp;; However, due<br />
to<br />
licensing restrictions, we have opted not to work with this protein. <o:p></o:p></p><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Recently (year 2007), a study<br />
has suggested an alternative approach by<br />
combining a glutamine-specific endoprotease (EP-B2 from barley) and a<br />
prolyl<br />
endopeptidase (SC PEP from <span style="font-style: italic;">Sphingomonas<br />
capsulata</span>) with gastric<br />
activity and<br />
complementary substrate specificity(12). Using this method, there is a possibility of<br />
increasing the<br />
safe threshold of ingested gluten(12). One of the advantages of this<br />
“combination product is that both enzymes are active and stable in<br />
stomach and<br />
can therefore be administered as lyophilized powders or simple capsules<br />
or<br />
tablets”(12). <o:p></o:p><br><br />
<o:p></o:p></p><br />
<p class="MsoNormal" style="line-height: normal;"><i><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></i><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span></p><br />
<hr style="width: 100%; height: 2px;"><b><u><span<br />
style="font-size: 20pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><a<br />
name="our_approach"></a>Our<br />
approach:<o:p></o:p></span></u></b><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Gliadin induces an immune<br />
system response when it is absorbed into<br />
the blood stream through the small intestine. We plan to take advantage<br />
of this by breaking down gliadin in the stomach before it reaches the<br />
small intestine. We have developed a secretion system to release an<br />
enzyme that can break down gliadin. Also, in order to prevent our<br />
delivery system from taking residence anywhere outside of the stomach,<br />
we plan to incorporate the difference in pH between the stomach and<br />
small intestine with a pH-inducible apoptosis system. By adopting these<br />
two systems, we will create a pH-moderated secretion system.<br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Unlike other suggested treatments<br />
requiring the consumption of capsules or tablets, our method of using <i>E.<br />
coli</i> as our delivery system is more cost- and time-efficient.<br />
</p><br />
<hr style="width: 100%; height: 2px;"><br />
<p class="MsoNormal"><span style="font-weight: bold;">Our project is<br />
split into two parts:</span><br><br />
1. Induced Secretion<br><br />
</p><br />
<ul><br />
<li><a href="https://2009.igem.org/Team:UC_Davis/Adding_secretion">Adding<br />
secretion</a><span style="text-decoration: underline;"></span></li><br />
<li><span style="text-decoration: underline;"></span><a<br />
href="https://2009.igem.org/Team:UC_Davis/Gene_sequence_for_secretion_system">Gene<br />
sequence for secretion system</a><span<br />
style="text-decoration: underline;"></span></li><br />
<li><span style="text-decoration: underline;"></span><a<br />
href="https://2009.igem.org/Team:UC_Davis/assay_for_measuring_protein_localization_and_activity">Assay<br />
for measuring protein localization and activity</a></li><br />
</ul><br />
<a<br />
href="https://2009.igem.org/Team:UC_Davis/assay_for_measuring_protein_localization_and_activity"></a><br />
<ul><br />
</ul><br />
2. Sensing pH&nbsp; and inducing cell death<br><br />
<ul><br />
<li><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project1/Finding_ph_sensor">Finding<br />
an appropriate biological pH sensor</a><br />
</li><br />
<li><a href="https://2009.igem.org/Team:UC_Davis/wiring_pH_sensor">Wiring<br />
the pH sensor into our system</a><br />
</li><br />
</ul><br />
<hr style="width: 100%; height: 2px;"><br />
<p class="MsoNormal"><span<br />
style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="color: black;"></span></span><b><span<br />
style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="color: black;"><o:p></o:p></span></span></b></p><br />
</div><br />
<p class="MsoListParagraphCxSpLast"<br />
style="margin: 0in 0in 0.0001pt 67.5pt; background: white none repeat scroll 0% 50%; text-indent: -0.25in; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial; text-align: left;"><small><span<br />
style="font-size: 18pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><br />
</span></small><!--[endif]--><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span></p><br />
</div><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_Davis/ProjectTeam:UC Davis/Project2009-10-22T02:03:13Z<p>Lydia: </p>
<hr />
<div><html><br />
<head><br />
<meta content="text/html;charset=ISO-8859-1" http-equiv="Content-Type"><br />
<title>lol</title><br />
</head><br />
<body><br />
<div style="text-align: center;"><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span>&nbsp;<br />
</b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC3.png"<br />
style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/d/d1/UCDAVIS_PIC4.png"<br />
style="border: 0px solid ; width: 82px; height: 36px;"></big></big></big></big></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 78px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 81px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Parts"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a<br />
href="https://2009.igem.org/Team:UC_Davis/Contacts_References"><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"<br />
style="border: 0px solid ; width: 83px; height: 37px;"></a></big></big></big></big></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><small><br />
</small></big></big></big></big></span></b><b<br />
style="color: rgb(0, 0, 0);"> </b><br />
<hr style="width: 100%; height: 2px;"><br />
<div style="text-align: left;"><b style="color: rgb(0, 0, 0);"><big<br />
style="text-decoration: underline;"><big>Project:</big></big><br><br />
</b><br />
<div style="text-align: center;"><b style="color: rgb(0, 0, 0);"><a<br />
href="#celiac_disease">Celiac<br />
Disease</a>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
<a href="#current_treatment">Current<br />
treatment</a>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<a<br />
href="#our_approach"><br />
Our Approach</a></b><br><br />
</div><br />
<small><u><span<br />
style="font-size: 20pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="font-weight: bold;"><a name="celiac_disease"></a></span></span></u></small><span<br />
style=""><big><big style="text-decoration: underline;"><span<br />
style="font-weight: bold;">What is Celiac Disease?</span></big></big><o:p></o:p></span><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Celiac Disease is an autoimmune<br />
disorder that occurs inside the small<br />
intestine. When the body cannot properly digest gliadin (a component of gluten),<br />
this leads to an immune response on the surface of the small intestine<br />
(14).<br />
Different people have varying degrees of immune<br />
response<br />
such as bloating, diarrhea, and weight loss (14). An estimated one out of 133 Americans currently suffer from this<br />
disease (11). Considering the estimated 3.08 million people in America, this isn't a minor issue. In fact, it was one of our teammate's friends suffering from celiac who initially sparked our interest in designing this project. <o:p></o:p><span style=""><small><span<br />
style="font-style: italic;"><br><br />
</span></small></span></p><br />
<p><span style=""><big style="font-weight: bold;"><span<br />
style="font-style: italic;">What happens in the normal small<br />
intestine?</span></big><o:p></o:p></span><span<br />
style="font-weight: bold;"></span><br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; <br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; The inside surface of the small intestine is covered with small microvilli, constituting the brush border membrane. In the normal small intestine, the brush border membrane is where polysaccharides, proteins, and fat droplets are digested into smaller parts that then get absorbed into the bloodstream. <br><br />
</p><br />
<p><o:p></o:p><span style=""><big><span<br />
style="font-weight: bold; font-style: italic;">What happens in the small intestine of people with celiac<br />
disease?</span></big><o:p></o:p></span></p><br />
<span style="font-weight: bold;"></span><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; The brush border characteristic to people with celiac disease allows large molecules such as gliadin to go straight<br />
into the<br />
blood stream without directing them through the transcellular route.<br />
Once<br />
gliadin passes through, Antigen Presenting Cells (APCs) recognize<br />
gliadin as a<br />
foreign object and attack. This immune response results in the inflammation of<br />
the surface of the intestine with loss of the normal cells required for<br />
absorption of sugars, protein, and fat from the diet. <o:p></o:p></p><br />
<p>In addition to the array of symtpoms illustrated above, the immune response may also damage intestinal villi, which are important<br />
for<br />
absorbing nutrients. The destruction of villi eventually leads to a<br />
lack of<br />
absorption of different nutrients, which can lead to malnutrition (14).<br />
Some<br />
people affected with this disorder can also suffer from autoimmune<br />
thyroid<br />
disease, autoimmune liver disease, and rheumatoid arthritis (<i>diseases<br />
in<br />
which body immune system attacks healthy cells/tissues</i>) (14). <o:p></o:p></p><br />
<hr style="width: 100%; height: 2px;"><br />
<p><u><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><span<br />
style="font-weight: bold;"><a name="current_treatment"></a>Current<br />
treatments:</span></big></big><o:p></o:p></span></u><br><br />
<b><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span><br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Currently, no cure has been found<br />
for this illness. The only way to avoid<br />
the symptoms of celiac disease is by adopting a gluten-free diet<br />
(14, 12,<br />
and 11). However, you are highly likely to find gluten in your everyday<br />
diet (in foods such as grains), and<br />
people with celiac disease must purchase gluten-free substances from<br />
specialized grocery stores. "Oral supplementation with prolyl<br />
oligopeptidases that can digest and detoxify gluten has therefore been<br />
proposed<br />
as a potential therapeutic approach."(8) However, enzymes studied<br />
earlier<br />
were not able to degrade gluten inside the stomach (before it reaches the small<br />
intestine), because they were "irreversibly inactivated by pepsin and<br />
acidic pH, both present in the stomach."(8) <o:p></o:p></p><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Nevertheless, over the past<br />
years, researchers have discovered an enzyme--a newly-identified prolyl endoprotease--from <i>Aspergillus niger</i>, <br />
that was<br />
observed to "work optimally at 4-5pH and remains stable at 2pH"(8).<br />
Perhaps this enzyme will lead us to an alternative treatment for this<br />
disorder(8). Studies have shown that prolyl endoprotease from <i>A.niger</i> is able to "degrade gluten in vitro and under conditions<br />
similar<br />
to the ones present in the gastrointestinal tract." (8)&nbsp;; However, due<br />
to<br />
licensing restrictions, we have opted not to work with this protein. <o:p></o:p></p><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Recently (year 2007), a study<br />
has suggested an alternative approach by<br />
combining a glutamine-specific endoprotease (EP-B2 from barley) and a<br />
prolyl<br />
endopeptidase (SC PEP from <span style="font-style: italic;">Sphingomonas<br />
capsulata</span>) with gastric<br />
activity and<br />
complementary substrate specificity(12). Using this method, there is a possibility of<br />
increasing the<br />
safe threshold of ingested gluten(12). One of the advantages of this<br />
“combination product is that both enzymes are active and stable in<br />
stomach and<br />
can therefore be administered as lyophilized powders or simple capsules<br />
or<br />
tablets”(12). <o:p></o:p><br><br />
<o:p></o:p></p><br />
<p class="MsoNormal" style="line-height: normal;"><i><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></i><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span></p><br />
<hr style="width: 100%; height: 2px;"><b><u><span<br />
style="font-size: 20pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><a<br />
name="our_approach"></a>Our<br />
approach:<o:p></o:p></span></u></b><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Gliadin induces an immune<br />
system response when it is absorbed into<br />
the blood stream through the small intestine. We plan to take advantage<br />
of this by breaking down gliadin in the stomach before it reaches the<br />
small intestine. We have developed a secretion system to release an<br />
enzyme that can break down gliadin. Also, in order to prevent our<br />
delivery system from taking residence anywhere outside of the stomach,<br />
we plan to incorporate the difference in pH between the stomach and<br />
small intestine with a pH-inducible apoptosis system. By adopting these<br />
two systems, we will create a pH-moderated secretion system.<br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Unlike other suggested treatments<br />
requiring the consumption of capsules or tablets, our method of using <i>E.<br />
coli</i> as our delivery system is more cost- and time-efficient.<br />
</p><br />
<hr style="width: 100%; height: 2px;"><br />
<p class="MsoNormal"><span style="font-weight: bold;">Our project is<br />
split into two parts:</span><br><br />
1.Induced Secretion<br><br />
</p><br />
<ul><br />
<li><a href="https://2009.igem.org/Team:UC_Davis/Adding_secretion">Adding<br />
secretion</a><span style="text-decoration: underline;"></span></li><br />
<li><span style="text-decoration: underline;"></span><a<br />
href="https://2009.igem.org/Team:UC_Davis/Gene_sequence_for_secretion_system">Gene<br />
sequence for secretion system</a><span<br />
style="text-decoration: underline;"></span></li><br />
<li><span style="text-decoration: underline;"></span><a<br />
href="https://2009.igem.org/Team:UC_Davis/assay_for_measuring_protein_localization_and_activity">Assay<br />
for measuring protein localization and activity</a></li><br />
</ul><br />
<a<br />
href="https://2009.igem.org/Team:UC_Davis/assay_for_measuring_protein_localization_and_activity"></a><br />
<ul><br />
</ul><br />
2. Sensing pH&nbsp; and inducing cell death<br><br />
<ul><br />
<li><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project1/Finding_ph_sensor">Finding<br />
an appropriate biological pH sensor</a><br />
</li><br />
<li><a href="https://2009.igem.org/Team:UC_Davis/wiring_pH_sensor">Wiring<br />
the pH sensor into our system</a><br />
</li><br />
</ul><br />
<hr style="width: 100%; height: 2px;"><br />
<p class="MsoNormal"><span<br />
style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="color: black;"></span></span><b><span<br />
style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="color: black;"><o:p></o:p></span></span></b></p><br />
</div><br />
<p class="MsoListParagraphCxSpLast"<br />
style="margin: 0in 0in 0.0001pt 67.5pt; background: white none repeat scroll 0% 50%; text-indent: -0.25in; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial; text-align: left;"><small><span<br />
style="font-size: 18pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><br />
</span></small><!--[endif]--><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span></p><br />
</div><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_Davis/ProjectTeam:UC Davis/Project2009-10-21T23:54:24Z<p>Lydia: </p>
<hr />
<div><html><br />
<head><br />
<meta content="text/html;charset=ISO-8859-1" http-equiv="Content-Type"><br />
<title>lol</title><br />
</head><br />
<body><br />
<div style="text-align: center;"><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span>&nbsp;<br />
</b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC3.png"<br />
style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/d/d1/UCDAVIS_PIC4.png"<br />
style="border: 0px solid ; width: 82px; height: 36px;"></big></big></big></big></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 78px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 81px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Parts"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a<br />
href="https://2009.igem.org/Team:UC_Davis/Contacts_References"><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"<br />
style="border: 0px solid ; width: 83px; height: 37px;"></a></big></big></big></big></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><small><br />
</small></big></big></big></big></span></b><b<br />
style="color: rgb(0, 0, 0);"> </b><br />
<hr style="width: 100%; height: 2px;"><br />
<div style="text-align: left;"><b style="color: rgb(0, 0, 0);"><big<br />
style="text-decoration: underline;"><big>Project:</big></big><br><br />
</b><br />
<div style="text-align: center;"><b style="color: rgb(0, 0, 0);"><a<br />
href="#celiac_disease">Celiac<br />
Disease</a>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
<a href="#current_treatment">Current<br />
treatment</a>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<a<br />
href="#our_approach"><br />
Our Approach</a></b><br><br />
</div><br />
<small><u><span<br />
style="font-size: 20pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="font-weight: bold;"><a name="celiac_disease"></a></span></span></u></small><span<br />
style=""><big><big style="text-decoration: underline;"><span<br />
style="font-weight: bold;">What is Celiac Disease?</span></big></big><o:p></o:p></span><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Celiac Disease is an autoimmune<br />
disorder that occurs inside the small<br />
intestine. When the body cannot properly digest gliadin (a component of gluten),<br />
this leads to an immune response on the surface of the small intestine<br />
(14).<br />
Different people have varying degrees of immune<br />
response<br />
such as bloating, diarrhea, and weight loss (14). An estimated one out of 133 Americans currently suffer from this<br />
disease (11). Considering the estimated 3.08 million people in America, this isn't a minor issue. In fact, it was one of our teammate's friends suffering from celiac who initially sparked our interest in designing this project. <o:p></o:p><span style=""><small><span<br />
style="font-style: italic;"><br><br />
</span></small></span></p><br />
<p><span style=""><big style="font-weight: bold;"><span<br />
style="font-style: italic;">What happens in the normal small<br />
intestine?</span></big><o:p></o:p></span><span<br />
style="font-weight: bold;"></span><br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; <br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; The inside surface of the small intestine is covered with small microvilli, constituting the brush border membrane. In the normal small intestine, the brush border membrane is where polysaccharides, proteins, and fat droplets are digested into smaller parts that then get absorbed into the bloodstream. <br><br />
</p><br />
<p><o:p></o:p><span style=""><big><span<br />
style="font-weight: bold; font-style: italic;">What happens in Celiac<br />
disease small<br />
intestine?</span></big><o:p></o:p></span></p><br />
<span style="font-weight: bold;"></span><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; The brush border membrane in<br />
the small intestine of people suffering from<br />
Celiac Disease allows large molecules such as gliadin to go straight<br />
into the<br />
blood stream without directing them through the transcellular route.<br />
Once<br />
gliadin passes through, Antigen Presenting Cells (APCs) recognize<br />
gliadin as a<br />
foreign object and will attack. This immune response results in<br />
inflammation of<br />
the surface of the intestine with loss of the normal cells required for<br />
absorption of sugars, protein, and fat from the diet. <o:p></o:p></p><br />
<p>The immune response not only causes abdominal pain, bloating,<br />
diarrhea and<br />
weight loss it may also damage intestinal villi, which are important<br />
for<br />
absorbing nutrients. The destruction of villi eventually leads to a<br />
lack of<br />
absorption of different nutrients, which can lead to malnutrition (14).<br />
Some<br />
people affected with this disorder can also suffer from autoimmune<br />
thyroid<br />
disease, autoimmune liver disease, and rheumatoid arthritis (<i>diseases<br />
in<br />
which body immune system attacks healthy cells/tissues</i>) (14). <o:p></o:p></p><br />
<hr style="width: 100%; height: 2px;"><br />
<p><u><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><span<br />
style="font-weight: bold;"><a name="current_treatment"></a>Current<br />
treatments:</span></big></big><o:p></o:p></span></u><br><br />
<b><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span><br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Currently, no cure has been found<br />
for this illness. The only way to avoid<br />
some of celiac disease’s symptoms is by adopting a gluten free diet<br />
(14, 12,<br />
and 11). However, you are more likely to find gluten in your everyday<br />
diet, and<br />
people with celiac disease must purchase gluten-free substances from<br />
specialized grocery stores. "Oral supplementation with prolyl<br />
oligopeptidases that can digest and detoxify gluten has therefore been<br />
proposed<br />
as a potential therapeutic approach."(8). However, enzymes studied<br />
earlier<br />
were not able to degrade gluten inside stomach (before it reaches small<br />
intestine) because they were "irreversibly inactivated by pepsin and<br />
acidic pH, both present in the stomach."(8) <o:p></o:p></p><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Nevertheless, over the past<br />
years, researchers have discovered an enzyme<br />
from <i>Aspergillus niger</i>, a newly identified prolyl endoprotease,<br />
that was<br />
observed to "work optimally at 4-5pH and remains stable at 2pH"(8)<br />
and maybe this enzyme will lead us to an alternative treatment for this<br />
disorder (8). Studies have shown that prolyl-endoprotease from <i>Aspergillus<br />
niger</i> is able to "degrade gluten in vitro and under conditions<br />
similar<br />
to the ones present in the gastrointestinal tract." (8)&nbsp;; but due<br />
to<br />
licensing restrictions we have opted not to work with this protein. <o:p></o:p></p><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Recently (year 2007), a study<br />
has suggested an alternative approach by<br />
combining a glutamine-specific endoprotease (EP-B2 from barley) and a<br />
prolyl<br />
endopeptidase (SC PEP from <span style="font-style: italic;">Sphingomonas<br />
capsulata</span>); with gastric<br />
activity and<br />
complementary substrate specificity there is a possibility of<br />
increasing the<br />
safe threshold of ingested gluten (12). One of the advantages of this<br />
“combination product is that both enzymes are active and stable in<br />
stomach and<br />
can therefore be administered as lyophilized powders or simple capsules<br />
or<br />
tablets” (12). <o:p></o:p><br><br />
<o:p></o:p></p><br />
<p class="MsoNormal" style="line-height: normal;"><i><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></i><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span></p><br />
<hr style="width: 100%; height: 2px;"><b><u><span<br />
style="font-size: 20pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><a<br />
name="our_approach"></a>Our<br />
approach:<o:p></o:p></span></u></b><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Gliadin induces an immune<br />
system response when it is absorbed into<br />
the blood stream through the small intestine. We plan to take advantage<br />
of this by breaking down gliadin in the stomach before it reaches the<br />
small intestine. We have developed a secretion system to release an<br />
enzyme that can break down gliadin. Also, in order to prevent our<br />
delivery system from taking residence anywhere outside of the stomach,<br />
we plan to incorporate the difference in pH between the stomach and<br />
small intestine with a pH-inducible apoptosis system. By adopting these<br />
two systems, we will create a pH-moderated secretion system.<br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Unlike other suggested treatments<br />
requiring the consumption of capsules or tablets, our method of using <i>E.<br />
coli</i> as our delivery system is more cost- and time-efficient.<br />
</p><br />
<hr style="width: 100%; height: 2px;"><br />
<p class="MsoNormal"><span style="font-weight: bold;">Our project is<br />
split into two parts:</span><br><br />
1.Induced Secretion<br><br />
</p><br />
<ul><br />
<li><a href="https://2009.igem.org/Team:UC_Davis/Adding_secretion">Adding<br />
secretion</a><span style="text-decoration: underline;"></span></li><br />
<li><span style="text-decoration: underline;"></span><a<br />
href="https://2009.igem.org/Team:UC_Davis/Gene_sequence_for_secretion_system">Gene<br />
sequence for secretion system</a><span<br />
style="text-decoration: underline;"></span></li><br />
<li><span style="text-decoration: underline;"></span><a<br />
href="https://2009.igem.org/Team:UC_Davis/assay_for_measuring_protein_localization_and_activity">Assay<br />
for measuring protein localization and activity</a></li><br />
</ul><br />
<a<br />
href="https://2009.igem.org/Team:UC_Davis/assay_for_measuring_protein_localization_and_activity"></a><br />
<ul><br />
</ul><br />
2. Sensing pH&nbsp; and inducing cell death<br><br />
<ul><br />
<li><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project1/Finding_ph_sensor">Finding<br />
an appropriate biological pH sensor</a><br />
</li><br />
<li><a href="https://2009.igem.org/Team:UC_Davis/wiring_pH_sensor">Wiring<br />
the pH sensor into our system</a><br />
</li><br />
</ul><br />
<hr style="width: 100%; height: 2px;"><br />
<p class="MsoNormal"><span<br />
style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="color: black;"></span></span><b><span<br />
style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="color: black;"><o:p></o:p></span></span></b></p><br />
</div><br />
<p class="MsoListParagraphCxSpLast"<br />
style="margin: 0in 0in 0.0001pt 67.5pt; background: white none repeat scroll 0% 50%; text-indent: -0.25in; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial; text-align: left;"><small><span<br />
style="font-size: 18pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><br />
</span></small><!--[endif]--><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span></p><br />
</div><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_Davis/ProjectTeam:UC Davis/Project2009-10-21T18:46:39Z<p>Lydia: </p>
<hr />
<div><html><br />
<head><br />
<meta content="text/html;charset=ISO-8859-1" http-equiv="Content-Type"><br />
<title>lol</title><br />
</head><br />
<body><br />
<div style="text-align: center;"><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span>&nbsp;<br />
</b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC3.png"<br />
style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/d/d1/UCDAVIS_PIC4.png"<br />
style="border: 0px solid ; width: 82px; height: 36px;"></big></big></big></big></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 78px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 81px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Parts"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a<br />
href="https://2009.igem.org/Team:UC_Davis/Contacts_References"><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"<br />
style="border: 0px solid ; width: 83px; height: 37px;"></a></big></big></big></big></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><small><br />
</small></big></big></big></big></span></b><b<br />
style="color: rgb(0, 0, 0);"> </b><br />
<hr style="width: 100%; height: 2px;"><br />
<div style="text-align: left;"><b style="color: rgb(0, 0, 0);"><big<br />
style="text-decoration: underline;"><big>Project:</big></big><br><br />
</b><br />
<div style="text-align: center;"><b style="color: rgb(0, 0, 0);"><a<br />
href="#celiac_disease">Celiac<br />
Disease</a>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
<a href="#current_treatment">Current<br />
treatment</a>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<a<br />
href="#our_approach"><br />
Our Approach</a></b><br><br />
</div><br />
<small><u><span<br />
style="font-size: 20pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="font-weight: bold;"><a name="celiac_disease"></a></span></span></u></small><span<br />
style=""><big><big style="text-decoration: underline;"><span<br />
style="font-weight: bold;">What is Celiac Disease?</span></big></big><o:p></o:p></span><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Celiac Disease is an autoimmune<br />
disorder that occurs inside the small<br />
intestine. When the body cannot digest gliadin (a component of gluten)<br />
properly,<br />
this leads to an immune response in the surface of the small intestine<br />
(14).<br />
Furthermore, different people have different reactions to this immune<br />
response<br />
such as bloating, diarrhea, and weight loss (14). It has been estimated<br />
that<br />
about one out of 133 Americans are currently suffering from this<br />
disease (11);<br />
including one of our teammate's friends, and that was what sparked our<br />
interest<br />
in designing this project. <o:p></o:p><span style=""><small><span<br />
style="font-style: italic;"><br><br />
</span></small></span></p><br />
<p><span style=""><big style="font-weight: bold;"><span<br />
style="font-style: italic;">What happens in the normal small<br />
intestine?</span></big><o:p></o:p></span><span<br />
style="font-weight: bold;"></span><br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; <br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; In the normal small intestine,<br />
the brush border membrane on the surface<br />
digests starches and other sugars, proteins, and fat droplets and lets<br />
their<br />
breakdown products pass into the bloodstream. <br><br />
</p><br />
<p><o:p></o:p><span style=""><big><span<br />
style="font-weight: bold; font-style: italic;">What happens in Celiac<br />
disease small<br />
intestine?</span></big><o:p></o:p></span></p><br />
<span style="font-weight: bold;"></span><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; The brush border membrane in<br />
the small intestine of people suffering from<br />
Celiac Disease allows large molecules such as gliadin to go straight<br />
into the<br />
blood stream without directing them through the transcellular route.<br />
Once<br />
gliadin passes through, Antigen Presenting Cells (APCs) recognize<br />
gliadin as a<br />
foreign object and will attack. This immune response results in<br />
inflammation of<br />
the surface of the intestine with loss of the normal cells required for<br />
absorption of sugars, protein, and fat from the diet. <o:p></o:p></p><br />
<p>The immune response not only causes abdominal pain, bloating,<br />
diarrhea and<br />
weight loss it may also damage intestinal villi, which are important<br />
for<br />
absorbing nutrients. The destruction of villi eventually leads to a<br />
lack of<br />
absorption of different nutrients, which can lead to malnutrition (14).<br />
Some<br />
people affected with this disorder can also suffer from autoimmune<br />
thyroid<br />
disease, autoimmune liver disease, and rheumatoid arthritis (<i>diseases<br />
in<br />
which body immune system attacks healthy cells/tissues</i>) (14). <o:p></o:p></p><br />
<hr style="width: 100%; height: 2px;"><br />
<p><u><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><span<br />
style="font-weight: bold;"><a name="current_treatment"></a>Current<br />
treatments:</span></big></big><o:p></o:p></span></u><br><br />
<b><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span><br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Currently, no cure has been found<br />
for this illness. The only way to avoid<br />
some of celiac disease’s symptoms is by adopting a gluten free diet<br />
(14, 12,<br />
and 11). However, you are more likely to find gluten in your everyday<br />
diet, and<br />
people with celiac disease must purchase gluten-free substances from<br />
specialized grocery stores. "Oral supplementation with prolyl<br />
oligopeptidases that can digest and detoxify gluten has therefore been<br />
proposed<br />
as a potential therapeutic approach."(8). However, enzymes studied<br />
earlier<br />
were not able to degrade gluten inside stomach (before it reaches small<br />
intestine) because they were "irreversibly inactivated by pepsin and<br />
acidic pH, both present in the stomach."(8) <o:p></o:p></p><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Nevertheless, over the past<br />
years, researchers have discovered an enzyme<br />
from <i>Aspergillus niger</i>, a newly identified prolyl endoprotease,<br />
that was<br />
observed to "work optimally at 4-5pH and remains stable at 2pH"(8)<br />
and maybe this enzyme will lead us to an alternative treatment for this<br />
disorder (8). Studies have shown that prolyl-endoprotease from <i>Aspergillus<br />
niger</i> is able to "degrade gluten in vitro and under conditions<br />
similar<br />
to the ones present in the gastrointestinal tract." (8)&nbsp;; but due<br />
to<br />
licensing restrictions we have opted not to work with this protein. <o:p></o:p></p><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Recently (year 2007), a study<br />
has suggested an alternative approach by<br />
combining a glutamine-specific endoprotease (EP-B2 from barley) and a<br />
prolyl<br />
endopeptidase (SC PEP from <span style="font-style: italic;">Sphingomonas<br />
capsulata</span>); with gastric<br />
activity and<br />
complementary substrate specificity there is a possibility of<br />
increasing the<br />
safe threshold of ingested gluten (12). One of the advantages of this<br />
“combination product is that both enzymes are active and stable in<br />
stomach and<br />
can therefore be administered as lyophilized powders or simple capsules<br />
or<br />
tablets” (12). <o:p></o:p><br><br />
<o:p></o:p></p><br />
<p class="MsoNormal" style="line-height: normal;"><i><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></i><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span></p><br />
<hr style="width: 100%; height: 2px;"><b><u><span<br />
style="font-size: 20pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><a<br />
name="our_approach"></a>Our<br />
approach:<o:p></o:p></span></u></b><br />
<p>&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Gliadin induces an immune<br />
system response when it is absorbed into<br />
the blood stream through the small intestine. We plan to take advantage<br />
of this by breaking down gliadin in the stomach before it reaches the<br />
small intestine. We have developed a secretion system to release an<br />
enzyme that can break down gliadin. Also, in order to prevent our<br />
delivery system from taking residence anywhere outside of the stomach,<br />
we plan to incorporate the difference in pH between the stomach and<br />
small intestine with a pH-inducible apoptosis system. By adopting these<br />
two systems, we will create a pH-moderated secretion system.<br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; Unlike other suggested treatments<br />
requiring the consumption of capsules or tablets, our method of using <i>E.<br />
coli</i> as our delivery system is more cost- and time-efficient.<br />
</p><br />
<hr style="width: 100%; height: 2px;"><br />
<p class="MsoNormal"><span style="font-weight: bold;">Our project is<br />
split into two parts:</span><br><br />
1.Induced Secretion<br><br />
</p><br />
<ul><br />
<li><a href="https://2009.igem.org/Team:UC_Davis/Adding_secretion">Adding<br />
secretion</a><span style="text-decoration: underline;"></span></li><br />
<li><span style="text-decoration: underline;"></span><a<br />
href="https://2009.igem.org/Team:UC_Davis/Gene_sequence_for_secretion_system">Gene<br />
sequence for secretion system</a><span<br />
style="text-decoration: underline;"></span></li><br />
<li><span style="text-decoration: underline;"></span><a<br />
href="https://2009.igem.org/Team:UC_Davis/assay_for_measuring_protein_localization_and_activity">Assay<br />
for measuring protein localization and activity</a></li><br />
</ul><br />
<a<br />
href="https://2009.igem.org/Team:UC_Davis/assay_for_measuring_protein_localization_and_activity"></a><br />
<ul><br />
</ul><br />
2. Sensing pH&nbsp; and inducing cell death<br><br />
<ul><br />
<li><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project1/Finding_ph_sensor">Finding<br />
an appropriate biological pH sensor</a><br />
</li><br />
<li><a href="https://2009.igem.org/Team:UC_Davis/wiring_pH_sensor">Wiring<br />
the pH sensor into our system</a><br />
</li><br />
</ul><br />
<hr style="width: 100%; height: 2px;"><br />
<p class="MsoNormal"><span<br />
style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="color: black;"></span></span><b><span<br />
style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="color: black;"><o:p></o:p></span></span></b></p><br />
</div><br />
<p class="MsoListParagraphCxSpLast"<br />
style="margin: 0in 0in 0.0001pt 67.5pt; background: white none repeat scroll 0% 50%; text-indent: -0.25in; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial; text-align: left;"><small><span<br />
style="font-size: 18pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><br />
</span></small><!--[endif]--><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span></p><br />
</div><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_Davis/Adding_secretion/model_1Team:UC Davis/Adding secretion/model 12009-10-21T18:45:08Z<p>Lydia: </p>
<hr />
<div><html><br />
<head><br />
<meta content="text/html;charset=ISO-8859-1" http-equiv="Content-Type"><br />
<title>Model 1</title><br />
</head><br />
<body><br />
<div style="text-align: center;"><span<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="font-weight: bold;">=</span></span></span><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC3.png"<br />
style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/d/d1/UCDAVIS_PIC4.png"<br />
style="border: 0px solid ; width: 82px; height: 36px;"></big></big></big></big></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 78px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 81px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Parts"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 78px; height: 37px;"></a></big></big></big></big></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b></div><br />
<hr style="width: 100%; height: 2px;"><big><big><big><span<br />
style="text-decoration: underline; font-weight: bold;"><small>Secretion<br />
Model<br />
1:<br><br />
<br><br />
</small></span></big></big></big><big><span style="font-style: italic;">Click<br />
on an<br />
individual part for more information.<br><br />
</span></big><br />
<div style="text-align: center;"><big><big><big><span<br />
style="font-weight: bold;"><small><span<br />
style="text-decoration: underline; background-color: rgb(255, 255, 0);"><a<br />
href="#LacI"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/4/4e/UCDAVIS_PROMOTER_1.png"<br />
style="border: 0px solid ; width: 242px; height: 58px; background-color: rgb(255, 255, 255);"></a><a<br />
href="#RBS"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/1/15/UCDAVIS_RBS1.png"<br />
style="border: 0px solid ; width: 89px; height: 81px; background-color: rgb(255, 255, 255);"></a><a<br />
href="#INPNC"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/9/9c/UCDAVIS_ompA.png"<br />
style="border: 0px solid ; width: 106px; height: 59px; background-color: rgb(255, 255, 255);"></a><a<br />
href="#SS"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/9/9d/UCDAVIS_SS1.png"<br />
style="border: 0px solid ; width: 94px; height: 69px; background-color: rgb(255, 255, 255);"></a><img<br />
style="width: 106px; height: 59px; background-color: rgb(255, 255, 255);"<br />
alt="" src="https://static.igem.org/mediawiki/2009/0/0c/UCDAVIS_GENE1.png"><a<br />
href="#Tag"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/c/c0/UCDAVIS_6HIS.png"<br />
style="border: 0px solid ; width: 134px; height: 66px; background-color: rgb(255, 255, 255);"></a><a<br />
href="#Terminator"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/c/ca/UCDAVIS_STOP.png"<br />
style="border: 0px solid ; width: 94px; height: 83px; background-color: rgb(255, 255, 255);"></a><br><br />
</span></small></span></big></big></big><big><big><big><span<br />
style="font-weight: bold;"><small><span<br />
style="text-decoration: underline; background-color: rgb(255, 255, 0);"><br><br />
</span>a.<a href="#LacI"><span style="text-decoration: underline;"><img<br />
alt=""<br />
src="https://static.igem.org/mediawiki/2009/4/4e/UCDAVIS_PROMOTER_1.png"<br />
style="border: 0px solid ; width: 200px; height: 52px;"></span></a><span<br />
style="text-decoration: underline; background-color: rgb(255, 255, 0);"><a<br />
href="#RBS"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/1/15/UCDAVIS_RBS1.png"<br />
style="border: 0px solid ; width: 69px; height: 63px; background-color: rgb(255, 255, 255);"></a><a<br />
href="#INPNC"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/9/9c/UCDAVIS_ompA.png"<br />
style="border: 0px solid ; width: 106px; height: 51px; background-color: rgb(255, 255, 255);"></a><a<br />
href="#SS"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/9/9d/UCDAVIS_SS1.png"<br />
style="border: 0px solid ; width: 73px; height: 65px; background-color: rgb(255, 255, 255);"></a><a<br />
href="#GFP"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/7/7d/UCDAVIS_GFP.png"<br />
style="border: 0px solid ; width: 83px; height: 55px; background-color: rgb(255, 255, 255);"></a><a<br />
href="#Tag"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/c/c0/UCDAVIS_6HIS.png"<br />
style="border: 0px solid ; width: 110px; height: 58px; background-color: rgb(255, 255, 255);"></a><a<br />
href="#Terminator"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/c/ca/UCDAVIS_STOP.png"<br />
style="border: 0px solid ; width: 78px; height: 71px; background-color: rgb(255, 255, 255);"></a></span></small></span></big></big></big><br><br />
<div style="text-align: center;"><big><big><big><span<br />
style="font-weight: bold;"><small>b.<span<br />
style="text-decoration: underline;"><a href="#LacI"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/4/4e/UCDAVIS_PROMOTER_1.png"<br />
style="border: 0px solid ; width: 195px; height: 52px;"></a><a<br />
href="#RBS"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/1/15/UCDAVIS_RBS1.png"<br />
style="border: 0px solid ; width: 68px; height: 63px;"></a><a<br />
href="#INPNC"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/9/9c/UCDAVIS_ompA.png"<br />
style="border: 0px solid ; width: 105px; height: 51px;"></a><a<br />
href="#SS"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/9/9d/UCDAVIS_SS1.png"<br />
style="border: 0px solid ; width: 73px; height: 65px;"></a><a<br />
href="#Luciferase"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/9/98/UCDAVIS_Luciferase.png"<br />
style="border: 0px solid ; width: 87px; height: 49px;"></a><a<br />
href="#Tag"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/c/c0/UCDAVIS_6HIS.png"<br />
style="border: 0px solid ; width: 110px; height: 58px;"></a><a<br />
href="#Terminator"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/c/ca/UCDAVIS_STOP.png"<br />
style="border: 0px solid ; width: 78px; height: 71px;"></a></span></small></span></big></big></big><big><big><big><span<br />
style="font-weight: bold;"><small><span<br />
style="text-decoration: underline; background-color: rgb(255, 255, 0);"></span></small></span></big></big></big></div><br />
</div><br />
<div style="text-align: center;"><big><span style="font-style: italic;"></span></big><br><br />
</div><br />
<big><big><big><span<br />
style="text-decoration: underline; font-weight: bold;"></span></big></big></big><br />
<hr style="width: 100%; height: 2px;"><br />
<p class="MsoNormal" style=""><b><span style=""><a name="INPNC"></a>INPNC:<br />
</span></b><span style="">The<br />
ice-nucleation protein (INP) from <i>Pseudomonas syringae</i> is used<br />
by its<br />
natural host to nucleate ice formation and is implicated in<i> P.syringae</i>-associated pathogenesis<i>.&nbsp; </i>INP, as well as a truncated derivative<br />
lacking<br />
the central domain (INPNC), have been used extensively for displaying<br />
proteins<br />
on the surface of <i>E. coli (7)</i>.&nbsp; For instance, AldO and<br />
PhaZ1 have<br />
been successfully displayed on the surface of <i>E. coli </i>using<br />
INPNC (7,<br />
15). <br><br />
<u1:p></u1:p>Park <i>et al.</i> have shown that INPNC, when fused to<br />
the <i>phaZ1</i><br />
gene, including its signal sequence, can serve as a suitable surface<br />
delivery<br />
and secretion device of the otherwise toxic <i>phaZ1</i> gene product<i><br />
</i>(15).&nbsp;<br />
<u1:p></u1:p>This part was synthesized by Mr. Gene (Regensburg,<br />
Germany) with<br />
codon optimization and subsequently transferred into vector (<span<br />
style="background: rgb(255, 255, 51) none repeat scroll 0%; color: black; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial;"><span<br />
style="-moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial; background-attachment: scroll;"> <i>pSB1AK3</i>).&nbsp;</span></span> As it is expected that this<br />
part will be used in the context of the fusion protein, the prefix and<br />
suffix<br />
for this part are consistent with the <i>BBF RCF-12</i><br />
standard.&nbsp; <br><br />
<u1:p></u1:p>We have proposed to build and test a general protein<br />
secretion<br />
system modeled after that developed by Park <i>et al. </i>in which a<br />
fusion of<br />
INPNC and the signal sequence from the <i>phaZ1</i> gene are used to<br />
secrete<br />
any target protein.&nbsp; <br><br />
<i><u1:p></u1:p>We have modified this protein to be consistent with the BBF<br />
RFC-12<br />
Standard. We have submitted this part to the parts registry as part </i><a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K265008"><i><span<br />
style="color: rgb(0, 0, 153);">BBa_K265008</span></i></a><i>.</i> <o:p></o:p></span></p><br />
<u2:p></u2:p><br />
<div class="MsoNormal" style="text-align: center;" align="center"><span<br />
style=""><br />
<hr align="center" size="2" width="100%"></span></div><br />
<p class="MsoNormal" style=""><b><span style=""><a name="OmpA"></a>OmpA</span></b><span<br />
style="">: OmpA<br />
is a protein found on the outer membrane of <i>E. coli</i> (13) and<br />
is used<br />
as a displaying fusion protein on the cell surface. This part has<br />
already been<br />
documented on the parts registry; however, it has not been tested as a<br />
component<br />
of a secretion system (via fusion with a target protein linked with a<br />
cleavable<br />
signal sequence). <i><br><br />
<u1:p></u1:p>We have modified this protein to be consistent with the BBF<br />
RFC-12 Standard.<br><br />
Note: “It has remained essentially unknown how proteins of E. coli<br />
outer<br />
membrane are sorted and incorporated into this membrane” (10)</i> <i><br><br />
For more information go to:<a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K103006"><br />
BBa_K103006</a></i><br />
<o:p></o:p></span></p><br />
<u2:p></u2:p><br />
<div class="MsoNormal" style="text-align: center;" align="center"><span<br />
style=""><br />
<hr align="center" size="2" width="100%"></span></div><br />
<p class="MsoNormal" style=""><b><span style=""><a name="RBS"></a>RBS</span></b><span<br />
style="">:&nbsp; Ribosome<br />
Binding site number 32 (BBa_J61132) from the registry is being used in<br />
our<br />
secretion system. <u2:p></u2:p><br><br />
<i>For more information go to:</i> <a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_J61132"><i>BBa_J61132</i></a><o:p></o:p></span></p><br />
<u2:p></u2:p><br />
<div class="MsoNormal" style="text-align: center;" align="center"><span<br />
style=""><br />
<hr align="center" size="2" width="100%"></span></div><br />
<p class="MsoNormal" style=""><b><span style=""><a name="Terminator"></a>Terminator</span></b><span<br />
style="">: We are<br />
using BBa_B0015, a double terminator, as our terminator in both our<br />
secretion<br />
and pH system.<u2:p></u2:p><br><br />
<i>For more information go to:</i> <a<br />
href="http://partsregistry.org/wiki/index.php?title=Part:BBa_B0015"><i>BBa_B0015</i></a><br />
<o:p></o:p></span></p><br />
<u2:p></u2:p><br />
<div class="MsoNormal" style="text-align: center;" align="center"><span<br />
style=""><br />
<hr align="center" size="2" width="100%"></span></div><br />
<p class="MsoNormal" style=""><b><span style=""><a name="GFP"></a>GFP</span></b><span<br />
style="">: We are<br />
using Green Fluorescent Protein as a reporter that also serves as a<br />
small<br />
protein in testing our secretion system.<br><br />
<i>For more informaiton go to: </i><a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K265003"><i>BBa_K265003</i></a><i>&nbsp;</i><u2:p></u2:p><o:p></o:p></span></p><br />
<div class="MsoNormal" style="text-align: center;" align="center"><span<br />
style=""><br />
<hr align="center" size="2" width="100%"></span></div><br />
<p class="MsoNormal" style=""><b><span style=""><a name="Luciferase"></a>Luciferase</span></b><span<br />
style="">: Luciferase<br />
is a firefly protein that also fluoresces, so it serves as a reporter<br />
as well<br />
as a testable large protein.<br><br />
<i>For more information go to: <a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_I712019">BBa_1712019</a></i><br />
<u2:p></u2:p><o:p></o:p></span></p><br />
<div class="MsoNormal" style="text-align: center;" align="center"><span<br />
style=""><br />
<hr align="center" size="2" width="100%"></span></div><br />
<p class="MsoNormal" style=""><b><span style=""><a name="LacI"></a>LacI</span></b><span<br />
style="">: This is an<br />
inducible promoter which was found in the part registry.<br><br />
<i>For more information go to:<a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_R0010"><br />
BBa_R0010</a><br style=""><br />
<!--[endif]--></i><o:p></o:p></span></p><br />
<u2:p></u2:p><br />
<div class="MsoNormal" style="text-align: center;" align="center"><span<br />
style=""><br />
<hr align="center" size="2" width="100%"></span></div><br />
<p class="MsoNormal" style=""><b><span style=""><a name="SS"></a>SS</span></b><span<br />
style="">: The<br />
signal sequence (SS) for the <i>phaZ1 </i>gene product of <i>Paucimonas<br />
lemoignei</i>, a polyhydroxybutyrate depolymerase (15).&nbsp; In the<br />
native<br />
protein the signal sequence is cleaved between residues Ala37 and<br />
Leu38.&nbsp;<br />
Park <i>et al. </i>have showed that the fusion of the complete <i>phaZ1<br />
</i>gene<br />
(including SS) and a truncated ice nucleation protein from <i>Pseudomonas<br />
syringae</i> (<a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K265008"><i><span<br />
style="color: rgb(0, 0, 153);">BBa_K265008</span></i></a>), could lead<br />
to stable<br />
expression and secretion of the <i>phaZ1</i> gene product.&nbsp; <br><br />
<u1:p></u1:p>We propose that the signal sequence might be generally<br />
useful as a<br />
cleavage tag in secretion systems that include a membrane anchor<br />
component,<br />
such as INPNC (<a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K265008"><i><span<br />
style="color: rgb(0, 0, 153);">BBa_K265008</span></i></a>) or OmpA (<i><a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K103006">BBa_K103006</a>).<span<br />
style="color: rgb(0, 41, 57);"> </span></i>The proposed constructs<br />
would consist of a<br />
membrane anchor (INPNC or OmpA) followed by the cleavable signal<br />
sequence and<br />
finally a target protein marked for secretion.&nbsp; <br><br />
<i><u1:p></u1:p>Since we expect that this part will be used in the<br />
context of a<br />
fusion protein, we have modified this protein to be consistent with the BBF<br />
RFC-12<br />
Standard. We have submitted this part to the part registry as part </i><a<br />
href="http://partsregistry.org/wiki/index.php/Part:BBa_K265002"><i><span<br />
style="color: rgb(0, 0, 153);">BBa_K265002</span></i></a>. <o:p></o:p></span></p><br />
<u2:p></u2:p><br />
<div class="MsoNormal" style="text-align: center;" align="center"><span<br />
style=""><br />
<hr align="center" size="2" width="100%"></span></div><br />
<p class="MsoNormal"><b><span style=""><a name="Tag"></a>6-His Tag</span></b><span<br />
style="">: The 6-Histidine Tag serves as a tag for<br />
Western Blotting if our fluorescent reporters are not expressed as<br />
highly as we<br />
would like. <u2:p></u2:p><br><br />
<i>Note: We are using this tag just in case the GFP or Luciferase<br />
do not<br />
work under a plate reader.</i> <o:p></o:p></span></p><br />
<div class="MsoNormal"<br />
style="margin-bottom: 0.0001pt; text-align: center; line-height: normal;"<br />
align="center"><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><br />
<hr align="center" size="2" width="100%"></span></div><br />
<b><i><span<br />
style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></i></b><big><big><big><span<br />
style="text-decoration: underline; font-weight: bold;"></span></big></big></big><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_Davis/Adding_secretionTeam:UC Davis/Adding secretion2009-10-21T18:27:03Z<p>Lydia: </p>
<hr />
<div><html><br />
<head><br />
<meta content="text/html;charset=ISO-8859-1" http-equiv="Content-Type"><br />
<title>Adding Secretion</title><br />
</head><br />
<body style="color: rgb(0, 0, 0); background-color: rgb(255, 255, 255);"<br />
alink="#ee0000" link="#0000ee" vlink="#551a8b"><br />
<div style="text-align: center;"><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC3.png"<br />
style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/d/d1/UCDAVIS_PIC4.png"<br />
style="border: 0px solid ; width: 82px; height: 36px;"></big></big></big></big></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 78px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 81px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Parts"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a<br />
href="https://2009.igem.org/Team:UC_Davis/Contacts_References"><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"<br />
style="border: 0px solid ; width: 83px; height: 37px;"></a></big></big></big></big></span></b></div><br />
<hr<br />
style="width: 100%; height: 2px; background-color: rgb(255, 255, 255);"><br />
<div style="text-align: left; background-color: rgb(255, 255, 255);"><big<br />
style="text-decoration: underline;"><big><span<br />
style="font-weight: bold;"><big><small>Adding Secretion:</small></big></span></big></big><br><br />
<div style="text-align: center;"><big style="font-weight: bold;"><a<br />
href="#general_model">General model for secretion</a>&nbsp;&nbsp;&nbsp;&nbsp;<br />
<a href="#secretion_model">Secretion Models</a>&nbsp;&nbsp;&nbsp;&nbsp;<br />
<a href="#different_genes">Why test different genes</a></big><br><br />
<hr style="width: 100%; height: 2px;"><big<br />
style="text-decoration: underline;"><big><span<br />
style="font-weight: bold;"></span></big></big></div><br />
<p class="MsoNormal" style="line-height: normal;"><span<br />
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span></p><br />
<b><u><span<br />
style="font-size: 20pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></u></b><big<br />
style="text-decoration: underline;"><big><span<br />
style="font-weight: bold;"></span></big></big><br />
<div style="text-align: center;"><big><big><big><span<br />
style="font-weight: bold;"><small><span<br />
style="text-decoration: underline; background-color: rgb(255, 255, 0);"><a<br />
name="general_model"></a>General<br />
model for secretion system:<br><br />
<img<br />
style="width: 242px; height: 58px; background-color: rgb(255, 255, 255);"<br />
alt=""<br />
src="https://static.igem.org/mediawiki/2009/4/4e/UCDAVIS_PROMOTER_1.png"><img<br />
style="width: 89px; height: 81px; background-color: rgb(255, 255, 255);"<br />
alt="" src="https://static.igem.org/mediawiki/2009/1/15/UCDAVIS_RBS1.png"><img<br />
style="width: 106px; height: 56px; background-color: rgb(255, 255, 255);"<br />
alt=""<br />
src="https://static.igem.org/mediawiki/2009/9/95/UCDAVIS_SECRETION.png"><img<br />
style="width: 94px; height: 69px; background-color: rgb(255, 255, 255);"<br />
alt="" src="https://static.igem.org/mediawiki/2009/9/9d/UCDAVIS_SS1.png"><img<br />
style="width: 106px; height: 59px; background-color: rgb(255, 255, 255);"<br />
alt="" src="https://static.igem.org/mediawiki/2009/0/0c/UCDAVIS_GENE1.png"><img<br />
style="width: 134px; height: 66px; background-color: rgb(255, 255, 255);"<br />
alt="" src="https://static.igem.org/mediawiki/2009/c/c0/UCDAVIS_6HIS.png"><img<br />
style="width: 94px; height: 83px; background-color: rgb(255, 255, 255);"<br />
alt="" src="https://static.igem.org/mediawiki/2009/c/ca/UCDAVIS_STOP.png"><br><br />
</span></small></span><small><small><span<br />
style="text-decoration: underline;"><br><br />
</span></small></small></big></big></big><br />
<p class="MsoNormal" style="text-align: left;"><big><span style="">&nbsp;&nbsp;&nbsp;<br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; <span<br />
style="font-family: Times New Roman,Times,serif;">The purpose of the<br />
secretion system is to<br />
introduce a method of secreting target proteins we wish to synthesize<br />
in the<br />
marvelous host: <i>E.coli</i>. Park </span><i<br />
style="font-family: Times New Roman,Times,serif;">et.<br />
al</i><span style="font-family: Times New Roman,Times,serif;"> showed<br />
that the truncated form of ice nucleation<br />
protein (INPNC) containing the complete coding region of phaZ1,<br />
including its<br />
signal sequence, could lead to stable secretion of the enzyme (</span><a<br />
style="font-family: Times New Roman,Times,serif;"<br />
href="https://2009.igem.org/Team:UC_Davis/Contacts_References">15</a><span<br />
style="font-family: Times New Roman,Times,serif;">). We have<br />
decided to see if we could co-opt this system to make a more<br />
generalized<br />
inducible system for protein secretion in which the INPNC and phaZ1<br />
signal<br />
sequence serve as carriers for a target protein.</span><o:p<br />
style="font-family: Times New Roman,Times,serif;"></o:p></span></big></p><br />
<p class="MsoNormal" style="font-family: Times New Roman,Times,serif;"><big><span<br />
style=""><br><br />
<big><b>Possible challenges</b>:</big><o:p></o:p></span></big></p><br />
<p class="MsoNormal"<br />
style="margin-left: 30pt; text-align: left; font-family: Times New Roman,Times,serif;"><big><span<br />
style="">1. Efficient recognition of the change site may<br />
<span<br />
style="-moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial; background-attachment: scroll;"><span<br />
style="background: yellow none repeat scroll 0% 50%; -moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial;">()</span></span><br />
some of the phaZ1 protein we have only cloned the<br />
signal sequence up to the cleavage site.<br><br />
2. The system may only work for proteins in a narrow site range.<br><br />
3. The expression levels of the system may be low.<o:p></o:p></span></big></p><br />
<p class="MsoNormal"<br />
style="text-align: center; font-family: Times New Roman,Times,serif;"<br />
align="center"><big><b><span style=""><br><br />
<big>Advantages:</big></span></b><span style=""><o:p></o:p></span></big></p><br />
<ol style="text-align: left;" start="1" type="1"><br />
<li style="font-family: Times New Roman,Times,serif;"<br />
class="MsoNormal"><big><span style=""><u1:p></u1:p>&nbsp; In our<br />
secretion system, we are using two genes with different sizes as our<br />
target secretion genes; GFP being short in length and Luciferase being<br />
comparably long, these would test the ability of our secretion system to secrete proteins of different sizes. <o:p></o:p></span></big></li><br />
<li style="font-family: Times New Roman,Times,serif;"<br />
class="MsoNormal"><big><span style="">&nbsp; Testing various<br />
combinations of the Signal Sequence with either ompA or INPNC would help us<br />
find the best combination for our secretion system based on their<br />
secretion ability and allow us to rank the combinations from strongest to weakest in strength.<br />
(in this specific system).<o:p></o:p></span></big></li><br />
<li class="MsoNormal"><span style=""><big<br />
style="font-family: Times New Roman,Times,serif;">&nbsp; We have<br />
submitted the BioBricks to the parts registry; therefore they can be<br />
used in future studies (for different purposes).</big><o:p></o:p></span></li><br />
</ol><br />
<br><br />
<div style="text-align: left;"><big><big<br />
style="font-family: Times New Roman,Times,serif;"><big><big><span<br />
style="font-weight: bold;"><small><span<br />
style="text-decoration: underline; background-color: rgb(255, 255, 0);"></span></small></span></big></big></big></big><br><br />
</div><br />
<div style="text-align: left;"><br />
<hr style="width: 100%; height: 2px;"><big><small><big<br />
style="font-weight: bold;"><span style="text-decoration: underline;"><small><big><a<br />
name="secretion_model"></a>Secretion<br />
Models:</big><br><br />
</small></span></big><span style="font-style: italic;">Click on a<br />
specific model for more information:</span><big<br />
style="font-weight: bold;"><span style="text-decoration: underline;"><small><br><br />
</small></span></big><span style="font-style: italic;"><big><span<br />
style="text-decoration: underline;"></span></big></span></small></big><br />
<div style="text-align: center;"><big><span style="font-weight: bold;"></span></big><big><span<br />
style="font-weight: bold;"><a<br />
href="https://2009.igem.org/Team:UC_Davis/Adding_secretion/model_1"><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/b/ba/UCDAVIS_PIC12.png"<br />
style="border: 0px solid ; width: 95px; height: 63px;"></a><a<br />
href="https://2009.igem.org/Team:UC_Davis/Adding_secretion/model_2"><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC13.png"<br />
style="border: 0px solid ; width: 95px; height: 63px;"></a>&nbsp;&nbsp;&nbsp;<br />
</span><span style="font-style: italic;"><big><span<br />
style="text-decoration: underline; font-weight: bold;"></span></big></span></big><br><br />
</div><br />
<div style="text-align: center;"><big><span style="font-style: italic;"><big><span<br />
style="text-decoration: underline; font-weight: bold;"></span></big></span></big></div><br />
<big><span style="font-style: italic;"></span></big><span<br />
style="text-decoration: underline;"><span style="font-weight: bold;"></span></span><br><br />
<big><span style="font-style: italic;"></span></big><br />
<p class="MsoNormal"><b><span<br />
style="font-size: 13.5pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span></b></p><br />
<big><big style="font-weight: bold;"><br />
</big></big><br />
<hr style="width: 100%; height: 2px;"><br />
<div style="text-align: left;"><b><u><span<br />
style="font-size: 20pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><small><a<br />
name="different_genes"></a>Why<br />
test different genes in our secretion system?</small><o:p></o:p></span></u></b></div><br />
<p class="MsoNormal"><b><span<br />
style="font-size: 13.5pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="">&nbsp;&nbsp;&nbsp;&nbsp; </span></span></b><span<br />
style="font-size: 13.5pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><small>There<br />
is wide range of genes present in our environment. Therefore, it is<br />
important<br />
to measure our secretion system’s ability to secrete genes of various<br />
sizes.</small></span><b><span<br />
style="font-size: 13.5pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><span<br />
style="text-decoration: underline;"><br><br />
</span></big></span></b></p><br />
<div style="text-align: center;"><b><span<br />
style="font-size: 13.5pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="text-decoration: underline;">Molecular data on some proteins</span><o:p></o:p></span></b></div><br />
<br><br />
<table<br />
style="text-align: left; width: 422px; background-color: rgb(0, 0, 153); height: 268px; margin-left: auto; margin-right: auto;"<br />
border="1" cellpadding="2" cellspacing="2"><br />
<tbody><br />
<tr><br />
<td<br />
style="vertical-align: top; background-color: rgb(0, 0, 153); text-align: center;"><big><big<br />
style="font-weight: bold;"><span style="color: rgb(255, 255, 0);">Name</span></big></big><br><br />
</td><br />
<td style="vertical-align: top; text-align: center;"><big<br />
style="color: rgb(255, 255, 0);"><big><span style="font-weight: bold;">Molecular<br />
Weight (kD)</span></big></big><br><br />
</td><br />
</tr><br />
<tr><br />
<td<br />
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>Myosin<br><br />
</big></td><br />
<td<br />
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>200.0<br><br />
</big></td><br />
</tr><br />
<tr><br />
<td<br />
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>-Galactosidase<br><br />
</big></td><br />
<td<br />
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>116.3<br><br />
</big></td><br />
</tr><br />
<tr><br />
<td<br />
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold; text-align: left;"><big>Phosphorylase<br />
b<br><br />
</big></td><br />
<td<br />
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>97.4<br><br />
</big></td><br />
</tr><br />
<tr><br />
<td<br />
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>Ovalbumin<br><br />
</big></td><br />
<td<br />
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>45.0<br><br />
</big></td><br />
</tr><br />
<tr><br />
<td<br />
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>Carbonic<br />
anhydrase<br><br />
</big></td><br />
<td<br />
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>31.0<br><br />
</big></td><br />
</tr><br />
<tr><br />
<td<br />
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>Aprotinin<br><br />
</big></td><br />
<td<br />
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>6.5<br><br />
</big></td><br />
</tr><br />
<tr><br />
<td<br />
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>Insulin,<br />
B chain, oxidized<br><br />
</big></td><br />
<td<br />
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>3.5<br><br />
</big></td><br />
</tr><br />
</tbody><br />
</table><br />
<br><br />
<p style="text-align: center;" class="MsoNormal"><b><span<br />
style="font-size: 14pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><br<br />
style=""><br />
<!--[if !supportLineBreakNewLine]--><br style=""><br />
<!--[endif]--></span></b><b><u><span<br />
style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span></u></b></p><br />
<big style="text-decoration: underline;"><big><span<br />
style="font-weight: bold;"></span></big></big></div><br />
</div><br />
<div style="text-align: center;"><br><br />
<big style="text-decoration: underline;"><big><span<br />
style="font-weight: bold;"></span></big></big></div><br />
</div><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_DavisTeam:UC Davis2009-10-21T18:07:57Z<p>Lydia: </p>
<hr />
<div><html><br />
<head><br />
<meta content="text/html;charset=ISO-8859-1" http-equiv="Content-Type"><br />
<title>Homepage_UCdavis</title><br />
</head><br />
<body style="background-color: rgb(255, 255, 255); color: rgb(0, 0, 0);"<br />
alink="#ee0000" link="#0000ee" vlink="#551a8b"><br />
<div style="text-align: center;"><span<br />
style="color: rgb(255, 255, 153);">&nbsp;<br />
</span><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><span<br />
style="color: rgb(255, 255, 153);"><img src="" alt=""<br />
style="width: 151px; height: 161px;"></span><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big>&nbsp;<br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/d/d1/UCDAVIS_PIC4.png"<br />
style="border: 0px solid ; width: 92px; height: 46px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 92px; height: 46px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 92px; height: 46px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Parts"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 92px; height: 46px;"></a>&nbsp;<a<br />
href="https://2009.igem.org/Team:UC_Davis/Contacts_References"><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"<br />
style="border: 0px solid ; width: 92px; height: 46px;"></a></big></big></big></big></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><small><br />
<img style="width: 191px; height: 147px;" alt="" src=""><br><br />
</small></big></big></big></big></span></b></div><br />
<hr style="width: 100%; height: 2px;"><b style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><big><span<br />
style="color: rgb(0, 0, 0);"></span></big><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="font-family: Symbol;"></span><span<br />
style="font-family: Arial Narrow; font-weight: bold;"></span></span></b><span<br />
style="color: rgb(0, 0, 0); font-family: Arial Narrow; font-weight: bold;"><span<br />
style="font-size: 13pt; line-height: 115%;"></span></span><br />
<p class="MsoNormal"<br />
style="margin-bottom: 0.0001pt; line-height: normal;"><b><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;">&nbsp;&nbsp;&nbsp;<br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp;<big> </big></span></b><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black; font-weight: bold;"><big>&nbsp;<br />
</big><big>It<br />
is estimated that about one in<br />
every 133 Americans suffer from an autoimmune disorder called Celiac<br />
Disease<br />
(14, 11). Celiac Disease is a condition in which the small intestines of<br />
an<br />
affected individual cannot properly digest gliadin, a protein present<br />
in gluten.<br />
When<br />
consumed, gliadin induces immune system responses, the<br />
side-effects of which are,<br />
but not limited to: abdominal pain, chronic diarrhea, and vomiting (14).<br />
Currently, the only accepted treatment for this disorder is adhering to<br />
a<br />
gluten-free diet (14, 12). This can be very difficult as great<br />
varieties of<br />
food contain gluten. This inspired us to imagine an alternative<br />
treatment based on a probiotic organism<br />
that can<br />
survive and take residence in the stomach, where it would secrete an<br />
enzyme that<br />
degrades gliadin.</big></span><big style="font-weight: bold;"><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; &nbsp;&nbsp; Then the design<br />
elements of this organism should satisfy at least two main<br />
criteria.<br />
First, it<br />
should be able to survive only in the stomach; second, it should secrete an<br />
enzyme<br />
that degrades gliadin<i>.</i> We envision that these parts would<br />
not<br />
only be useful for our project, but also to other future projects.</span><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span><br><br />
</big><span style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big<br />
style="font-weight: bold;">&nbsp;&nbsp;&nbsp;<br />
&nbsp; &nbsp;<br />
&nbsp;&nbsp; We decided to focus our summer project on building the<br />
first of the two key elements<br />
required for<br />
our system:<o:p></o:p><br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp;<br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; 1. <a<br />
href="https://2009.igem.org/Team:UC_Davis/secretion">Produce an<br />
inducible secretion<br />
system</a><br><br />
</big><br />
<big style="font-weight: bold;">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
2. <a href="https://2009.igem.org/Team:UC_Davis/ph_sensor1">Construct a<br />
biological pH sensor limiting this secretion system to<br />
the<br />
stomach</a></big></span><a<br />
href="https://2009.igem.org/Team:UC_Davis/ph_sensor1"><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="text-decoration: underline;"></span></span></a><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big<br />
style="font-weight: bold;"><br><br />
</big></span></p><br />
<p class="MsoNormal"<br />
style="margin-bottom: 0.0001pt; line-height: normal;"><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span><a<br />
href="https://2009.igem.org/Team:UC_Davis/ph_sensor1"><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b></a><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
&nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp;<br />
&nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp;<br />
&nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp;<br />
&nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp;<br />
&nbsp; &nbsp; &nbsp; &nbsp; &nbsp;<br />
&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
&nbsp;</span><a href="https://2009.igem.org/Team:UC_Davis/ph_sensor1"><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="text-decoration: underline;"></span></span></a><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project_Summary"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/ac/UCDAVIS_summary.png"<br />
style="border: 0px solid ; width: 63px; height: 50px;"></a><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
<span style="text-decoration: underline;"><br><br />
</span> </span><a href="https://2009.igem.org/Team:UC_Davis/ph_sensor1"><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b></a><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b></p><br />
<hr style="width: 100%; height: 2px;"><br />
<p class="MsoNormal"<br />
style="margin-bottom: 0.0001pt; line-height: normal; color: rgb(0, 0, 0); text-align: center; background-color: rgb(255, 255, 255);"><b><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b></p><br />
<ul style="background-color: rgb(255, 255, 255); color: rgb(0, 0, 0);"><br />
</ul><br />
<div style="text-align: center;"><big> </big></div><br />
<div style="text-align: center;"><br />
<div style="text-align: left;"><br />
<div style="text-align: center;"><big><span style="font-weight: bold;"><big>&nbsp;&nbsp;<br />
&nbsp;&nbsp;<small><br />
UC<br />
Davis iGEM team is sponsored by:</small></big></span></big><br><br />
<big><span style="font-weight: bold;"></span></big></div><br />
<div style="text-align: center;">&nbsp;&nbsp; &nbsp;&nbsp; <img<br />
src="https://static.igem.org/mediawiki/2009/8/88/UCDAVIS_Biomedical_engineering.jpg"<br />
alt="" style="width: 83px; height: 109px;"><img<br />
style="width: 105px; height: 109px;" alt=""<br />
src="https://static.igem.org/mediawiki/2009/4/43/UCDAVIS_college_of_engineering.png"><img<br />
style="width: 135px; height: 99px;" alt=""<br />
src="https://static.igem.org/mediawiki/2009/8/8e/UCDAVIS_Accutek1.jpg"><img<br />
style="width: 121px; height: 109px;" alt=""<br />
src="https://static.igem.org/mediawiki/2009/6/6b/UCDAVIS_mrgen.gif"><img<br />
style="width: 182px; height: 108px;" alt=""<br />
src="https://static.igem.org/mediawiki/2009/c/c6/UCDAVIS_genome_center.jpg"><br><br />
<font size="-1">We want to thank <span style="font-weight: bold;">Marc</span><br />
<span style="font-weight: bold;">Facciotti</span> for his helpful<br />
advice.<br><br />
Special thanks to <span style="font-weight: bold;">Suzy<br />
Fenton</span>,<span style="font-weight: bold;">Wayne Fenton, Jeanne<br />
Joe-Fenton </span>and<span style="font-weight: bold;"> Wayne Fenton Jr.</span><br />
<br><br />
Also we thank <span style="font-size: 10pt; line-height: 115%;"><b>Charles<br />
H. <span class="il">Halsted </span></b><span class="il">for reviewing<br />
our material regarding celiac disease.</span></span></font><br><br />
<br><br />
</div><br />
</div><br />
</div><br />
<ul style="background-color: rgb(255, 255, 255); color: rgb(0, 0, 0);"><br />
<span style="font-weight: bold;"></span><br />
</ul><br />
<p class="MsoNormal"<br />
style="margin-bottom: 0.0001pt; line-height: normal; color: rgb(0, 0, 0); text-align: left; background-color: rgb(255, 255, 255);"><b><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
</span></b></p><br />
<span style="color: rgb(0, 0, 0);"></span><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_DavisTeam:UC Davis2009-10-21T18:07:12Z<p>Lydia: </p>
<hr />
<div><html><br />
<head><br />
<meta content="text/html;charset=ISO-8859-1" http-equiv="Content-Type"><br />
<title>Homepage_UCdavis</title><br />
</head><br />
<body style="background-color: rgb(255, 255, 255); color: rgb(0, 0, 0);"<br />
alink="#ee0000" link="#0000ee" vlink="#551a8b"><br />
<div style="text-align: center;"><span<br />
style="color: rgb(255, 255, 153);">&nbsp;<br />
</span><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><span<br />
style="color: rgb(255, 255, 153);"><img src="" alt=""<br />
style="width: 151px; height: 161px;"></span><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big>&nbsp;<br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/d/d1/UCDAVIS_PIC4.png"<br />
style="border: 0px solid ; width: 92px; height: 46px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 92px; height: 46px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 92px; height: 46px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Parts"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 92px; height: 46px;"></a>&nbsp;<a<br />
href="https://2009.igem.org/Team:UC_Davis/Contacts_References"><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"<br />
style="border: 0px solid ; width: 92px; height: 46px;"></a></big></big></big></big></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><small><br />
<img style="width: 191px; height: 147px;" alt="" src=""><br><br />
</small></big></big></big></big></span></b></div><br />
<hr style="width: 100%; height: 2px;"><b style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><big><span<br />
style="color: rgb(0, 0, 0);"></span></big><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="font-family: Symbol;"></span><span<br />
style="font-family: Arial Narrow; font-weight: bold;"></span></span></b><span<br />
style="color: rgb(0, 0, 0); font-family: Arial Narrow; font-weight: bold;"><span<br />
style="font-size: 13pt; line-height: 115%;"></span></span><br />
<p class="MsoNormal"<br />
style="margin-bottom: 0.0001pt; line-height: normal;"><b><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black;">&nbsp;&nbsp;&nbsp;<br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp;<big> </big></span></b><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;; color: black; font-weight: bold;"><big>&nbsp;<br />
</big><big>It<br />
is estimated that about one in<br />
every 133 Americans suffer from an autoimmune disorder called Celiac<br />
Disease<br />
(14, 11). Celiac Disease is a condition in which the small intestines of<br />
an<br />
affected individual cannot properly digest gliadin, a protein present<br />
in gluten.<br />
When<br />
consumed, gliadin induces immune system responses, the<br />
side-effects of which are,<br />
but not limited to: abdominal pain, chronic diarrhea, and vomiting (14).<br />
Currently, the only accepted treatment for this disorder is adhering to<br />
a<br />
gluten-free diet (14, 12). This can be very difficult as great<br />
varieties of<br />
food contain gluten. This inspired us to imagine an alternative<br />
treatment based on a probiotic organism<br />
that can<br />
survive and take residence in the stomach, where it would secrete an<br />
enzyme that<br />
will degrade gliadin.</big></span><big style="font-weight: bold;"><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; &nbsp;&nbsp; Then the design<br />
elements of this organism should satisfy at least two main<br />
criteria.<br />
First, it<br />
should be able to survive only in the stomach; second, it should secrete an<br />
enzyme<br />
that degrades gliadin<i>.</i> We envision that these parts would<br />
not<br />
only be useful for our project, but also to other future projects.</span><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span><br><br />
</big><span style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big<br />
style="font-weight: bold;">&nbsp;&nbsp;&nbsp;<br />
&nbsp; &nbsp;<br />
&nbsp;&nbsp; We decided to focus our summer project on building the<br />
first of the two key elements<br />
required for<br />
our system:<o:p></o:p><br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp;<br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; 1. <a<br />
href="https://2009.igem.org/Team:UC_Davis/secretion">Produce an<br />
inducible secretion<br />
system</a><br><br />
</big><br />
<big style="font-weight: bold;">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
2. <a href="https://2009.igem.org/Team:UC_Davis/ph_sensor1">Construct a<br />
biological pH sensor limiting this secretion system to<br />
the<br />
stomach</a></big></span><a<br />
href="https://2009.igem.org/Team:UC_Davis/ph_sensor1"><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="text-decoration: underline;"></span></span></a><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big<br />
style="font-weight: bold;"><br><br />
</big></span></p><br />
<p class="MsoNormal"<br />
style="margin-bottom: 0.0001pt; line-height: normal;"><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span><a<br />
href="https://2009.igem.org/Team:UC_Davis/ph_sensor1"><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b></a><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
&nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp;<br />
&nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp;<br />
&nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp;<br />
&nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp;<br />
&nbsp; &nbsp; &nbsp; &nbsp; &nbsp;<br />
&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
&nbsp;</span><a href="https://2009.igem.org/Team:UC_Davis/ph_sensor1"><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span<br />
style="text-decoration: underline;"></span></span></a><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project_Summary"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/ac/UCDAVIS_summary.png"<br />
style="border: 0px solid ; width: 63px; height: 50px;"></a><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
<span style="text-decoration: underline;"><br><br />
</span> </span><a href="https://2009.igem.org/Team:UC_Davis/ph_sensor1"><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b></a><span<br />
style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b></p><br />
<hr style="width: 100%; height: 2px;"><br />
<p class="MsoNormal"<br />
style="margin-bottom: 0.0001pt; line-height: normal; color: rgb(0, 0, 0); text-align: center; background-color: rgb(255, 255, 255);"><b><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b></p><br />
<ul style="background-color: rgb(255, 255, 255); color: rgb(0, 0, 0);"><br />
</ul><br />
<div style="text-align: center;"><big> </big></div><br />
<div style="text-align: center;"><br />
<div style="text-align: left;"><br />
<div style="text-align: center;"><big><span style="font-weight: bold;"><big>&nbsp;&nbsp;<br />
&nbsp;&nbsp;<small><br />
UC<br />
Davis iGEM team is sponsored by:</small></big></span></big><br><br />
<big><span style="font-weight: bold;"></span></big></div><br />
<div style="text-align: center;">&nbsp;&nbsp; &nbsp;&nbsp; <img<br />
src="https://static.igem.org/mediawiki/2009/8/88/UCDAVIS_Biomedical_engineering.jpg"<br />
alt="" style="width: 83px; height: 109px;"><img<br />
style="width: 105px; height: 109px;" alt=""<br />
src="https://static.igem.org/mediawiki/2009/4/43/UCDAVIS_college_of_engineering.png"><img<br />
style="width: 135px; height: 99px;" alt=""<br />
src="https://static.igem.org/mediawiki/2009/8/8e/UCDAVIS_Accutek1.jpg"><img<br />
style="width: 121px; height: 109px;" alt=""<br />
src="https://static.igem.org/mediawiki/2009/6/6b/UCDAVIS_mrgen.gif"><img<br />
style="width: 182px; height: 108px;" alt=""<br />
src="https://static.igem.org/mediawiki/2009/c/c6/UCDAVIS_genome_center.jpg"><br><br />
<font size="-1">We want to thank <span style="font-weight: bold;">Marc</span><br />
<span style="font-weight: bold;">Facciotti</span> for his helpful<br />
advice.<br><br />
Special thanks to <span style="font-weight: bold;">Suzy<br />
Fenton</span>,<span style="font-weight: bold;">Wayne Fenton, Jeanne<br />
Joe-Fenton </span>and<span style="font-weight: bold;"> Wayne Fenton Jr.</span><br />
<br><br />
Also we thank <span style="font-size: 10pt; line-height: 115%;"><b>Charles<br />
H. <span class="il">Halsted </span></b><span class="il">for reviewing<br />
our material regarding celiac disease.</span></span></font><br><br />
<br><br />
</div><br />
</div><br />
</div><br />
<ul style="background-color: rgb(255, 255, 255); color: rgb(0, 0, 0);"><br />
<span style="font-weight: bold;"></span><br />
</ul><br />
<p class="MsoNormal"<br />
style="margin-bottom: 0.0001pt; line-height: normal; color: rgb(0, 0, 0); text-align: left; background-color: rgb(255, 255, 255);"><b><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<br />
</span></b></p><br />
<span style="color: rgb(0, 0, 0);"></span><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_Davis/assay_for_measuring_protein_localization_and_activityTeam:UC Davis/assay for measuring protein localization and activity2009-09-24T08:10:29Z<p>Lydia: </p>
<hr />
<div><html><br />
<head><br />
<title></title><br />
</head><br />
<body><br />
<div style="text-align: center;"><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/homepage2"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC3.png"<br />
style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us1"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/d/d1/UCDAVIS_PIC4.png"<br />
style="border: 0px solid ; width: 82px; height: 36px;"></big></big></big></big></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project1"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 78px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook1"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 81px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Part1"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a<br />
href="https://2009.igem.org/Team:UC_Davis/Contact1"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"<br />
style="border: 0px solid ; width: 83px; height: 37px;"></a></big></big></big></big></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><small></small></big></big></big></big></span></b><br><br />
</div><br />
<hr style="width: 100%; height: 2px;"><br />
<div style="text-align: center;"><span<br />
style="font-weight: bold; text-decoration: underline;"><big>Assay for<br />
measuring protein localization and activity:</big><br><br />
</span><br />
<div style="text-align: left;"><br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; We will utilize both the Western<br />
blotting technique and the plate reader to identify and quantify our proteins, GFP and Luciferase.<br><br />
<hr style="width: 100%; height: 2px;"><br><br />
</div><br />
<span style="font-weight: bold; text-decoration: underline;"><br><br />
<br><br />
</span></div><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_Davis/assay_for_measuring_protein_localization_and_activityTeam:UC Davis/assay for measuring protein localization and activity2009-09-24T08:04:36Z<p>Lydia: </p>
<hr />
<div><html><br />
<head><br />
<title></title><br />
</head><br />
<body><br />
<div style="text-align: center;"><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/homepage2"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC3.png"<br />
style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us1"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/d/d1/UCDAVIS_PIC4.png"<br />
style="border: 0px solid ; width: 82px; height: 36px;"></big></big></big></big></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project1"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 78px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook1"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 81px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Part1"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a<br />
href="https://2009.igem.org/Team:UC_Davis/Contact1"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"<br />
style="border: 0px solid ; width: 83px; height: 37px;"></a></big></big></big></big></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><small></small></big></big></big></big></span></b><br><br />
</div><br />
<hr style="width: 100%; height: 2px;"><br />
<div style="text-align: center;"><span<br />
style="font-weight: bold; text-decoration: underline;"><big>Assay for<br />
measuring protein localization and activity:</big><br><br />
</span><br />
<div style="text-align: left;"><br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; We will utilize the Western<br />
blotting technique to identify and quantify our proteins. We will also<br />
use a plate reader to quantify fluorescence from GFP and Luciferase<br />
(our proteins).<br><br />
<hr style="width: 100%; height: 2px;"><br><br />
</div><br />
<span style="font-weight: bold; text-decoration: underline;"><br><br />
<br><br />
</span></div><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_Davis/assay_for_measuring_protein_localization_and_activityTeam:UC Davis/assay for measuring protein localization and activity2009-09-24T07:55:35Z<p>Lydia: </p>
<hr />
<div><html><br />
<head><br />
<title></title><br />
</head><br />
<body><br />
<div style="text-align: center;"><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/homepage2"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC3.png"<br />
style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a<br />
href="https://2009.igem.org/Team:UC_Davis/About_Us1"><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><img<br />
alt="" src="https://static.igem.org/mediawiki/2009/d/d1/UCDAVIS_PIC4.png"<br />
style="border: 0px solid ; width: 82px; height: 36px;"></big></big></big></big></span></b></a><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><br />
</big></big></big></big></span></b><b style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Project1"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"<br />
style="border: 0px solid ; width: 78px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Notebook1"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"<br />
style="border: 0px solid ; width: 81px; height: 36px;"></a> </big></big></big></big></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></b><b<br />
style="color: rgb(255, 255, 153);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a<br />
href="https://2009.igem.org/Team:UC_Davis/Part1"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"<br />
style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a<br />
href="https://2009.igem.org/Team:UC_Davis/Contact1"><img alt=""<br />
src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"<br />
style="border: 0px solid ; width: 83px; height: 37px;"></a></big></big></big></big></span></b><b<br />
style="color: rgb(0, 0, 0);"><span<br />
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><small></small></big></big></big></big></span></b><br><br />
</div><br />
<hr style="width: 100%; height: 2px;"><br />
<div style="text-align: center;"><span<br />
style="font-weight: bold; text-decoration: underline;"><big>Assay for<br />
measuring protein localization and activity:</big><br><br />
</span><br />
<div style="text-align: left;"><br><br />
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; We will utilize the Western<br />
blotting technique to identify and quantify our proteins, GFP and Luciferase, and also use a plate reader to measure the amount of fluorescence. <br />
<br><br />
<hr style="width: 100%; height: 2px;"><br><br />
</div><br />
<span style="font-weight: bold; text-decoration: underline;"><br><br />
<br><br />
</span></div><br />
</body><br />
</html></div>Lydiahttp://2009.igem.org/Team:UC_Davis/pH_Sensor/promotersTeam:UC Davis/pH Sensor/promoters2009-09-23T20:20:58Z<p>Lydia: /* Why is There a Need to Test Different Promoters Responsive to the ChvI Gene in Our System? */</p>
<hr />
<div>=Why is There a Need to Test Different Promoters Responsive to the ChvI Gene in Our System?=<br />
<br />
In response to environmental stimuli, due to the amino acids similarity and common mechanisms of different genes, there is a chance of cross-talk among our pH sensor device’s promoter and the genes already present in the ''E. coli'' cells (6). To minimize the effects of cross-talk in ''E. coli'', we need to find a gene that does not have a response to any other environmental changes other than an acidic environment.<br />
<br />
<br />
[[https://2009.igem.org/Team:UC_Davis/About_Us About Us]] [[https://2009.igem.org/Team:UC_Davis/Celiac Celiac Disease]] [[https://2009.igem.org/Team:UC_Davis/Treatments Current Treatment]] [[https://2009.igem.org/Team:UC_Davis/Approach Our Approach]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor pH Sensor]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/parts pH Sensor Parts]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/promoters pH Promoters]] [[https://2009.igem.org/Team:UC_Davis/Secretion/parts Secretion Parts]]</div>Lydiahttp://2009.igem.org/Team:UC_Davis/Secretion/partsTeam:UC Davis/Secretion/parts2009-09-23T20:20:31Z<p>Lydia: </p>
<hr />
<div>=More Parts=<br />
<br />
==INPNC== <br />
<br />
Ice-nucleation protein (INP) from Pseudomonas Syringae was suggested to be used for display of foreign proteins on the surface of E. ''coli''(7).Furthermore, researches have shown that an INP derivative constituting the N-and C-terminal domains can and has been used to display foreign proteins on the surface of E. ''coli''(9). In our project we are intending to harness and make use of this feature by fusing a specific protein to it. <br />
<br />
We have modified this protein to Biobrick standard, Tom Knights Standard. <br />
<br />
==OmpA==<br />
<br />
OmpA is one of the proteins on the outer membrane of E. ''coli'' (13). OmpA has been found to be useful as utilizable fusion part that can fuse our protein to and display on the surface of E. ''coli''. This part has already been documented on the parts registry; however, it has not been tested via fusion with a target protein linked with a cleavable signal sequence.<br />
<br />
We have modified this protein to Biobrick standard, Tom Knights Standard. <br />
<br />
''Note: “It has remained essentially unknown how proteins of Escherichia coli outer membrane are sorted and incorporated into this membrane” (10)''<br />
<br />
For more information [[http://partsregistry.org/wiki/index.php?title=Part:BBa_J36836 click here]]<br />
<br />
<br />
==RBS== <br />
<br />
Ribosome Binding site number 32 (BBa_J61132) from the registry is being used in our secretion system.<br />
For more information [http://partsregistry.org/wiki/index.php/Part:BBa_J61132 Click Here]]<br />
<br />
==Terminator==<br />
<br />
We are using BBa_B0015, a double terminator, as our terminator in both our secretion and pH system.<br />
<br />
For more information [[http://partsregistry.org/wiki/index.php?title=Part:BBa_B0015 Click Here]]<br />
<br />
==GFP==<br />
<br />
We are using Green Fluorescent Protein as a reporter that also serves as a small protein in testing our secretion system.<br />
<br />
==Luciferase==<br />
<br />
Luciferase is a firefly protein that also fluoresces, so it serves as a reporter as well as a testable large protein.<br />
<br />
More can be found in:<br />
<br />
==LacI==<br />
<br />
One inducible Promoter which was found in the part registry.<br />
<br />
More can be found [[http://partsregistry.org/wiki/index.php?title=Part:BBa_R0010 Here]]<br />
<br />
==SS== <br />
<br />
This signal sequence, when placed between INPNC, contains a cleavable site that allows the target fusion protein to ‘secrete’ from INPNC. We will do the same with OmpA. <br />
<br />
We have modified this protein to Biobrick standard, Tom Knights Standard. <br />
<br />
==6-His Tag==<br />
<br />
The 6-Histidine Tag serves as a tag for Western Blotting if our fluorescent reporters are not expressed as highly as we would like.<br />
<br />
''Note: We are using this tag, just in case if the GFP or Luciferase does not work under a plate reader.''<br />
<br />
<br />
[[https://2009.igem.org/Team:UC_Davis/About_Us About Us]] [[https://2009.igem.org/Team:UC_Davis/Celiac Celiac Disease]] [[https://2009.igem.org/Team:UC_Davis/Treatments Current Treatment]] [[https://2009.igem.org/Team:UC_Davis/Approach Our Approach]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor pH Sensor]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/parts pH Sensor Parts]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/promoters pH Promoters]] [[https://2009.igem.org/Team:UC_Davis/Secretion/parts Secretion Parts]]</div>Lydiahttp://2009.igem.org/Team:UC_Davis/pH_Sensor/promotersTeam:UC Davis/pH Sensor/promoters2009-09-23T20:20:09Z<p>Lydia: /* Why is There a Need to Test Different Promoters Responsive to the ChvI Gene in Our System? */</p>
<hr />
<div>=Why is There a Need to Test Different Promoters Responsive to the ChvI Gene in Our System?=<br />
<br />
In response to environmental stimuli, due to the amino acids similarity and common mechanisms of different genes, there is a chance of cross-talk among our pH sensor device’s promoter and the genes already present in the E. ''coli'' cells (6). To minimize the effects of cross-talk in ''E. coli'', we need to find a gene that does not have a response to any other environmental changes other than an acidic environment.<br />
<br />
<br />
[[https://2009.igem.org/Team:UC_Davis/About_Us About Us]] [[https://2009.igem.org/Team:UC_Davis/Celiac Celiac Disease]] [[https://2009.igem.org/Team:UC_Davis/Treatments Current Treatment]] [[https://2009.igem.org/Team:UC_Davis/Approach Our Approach]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor pH Sensor]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/parts pH Sensor Parts]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/promoters pH Promoters]] [[https://2009.igem.org/Team:UC_Davis/Secretion/parts Secretion Parts]]</div>Lydiahttp://2009.igem.org/Team:UC_Davis/pH_Sensor/partsTeam:UC Davis/pH Sensor/parts2009-09-23T20:19:39Z<p>Lydia: </p>
<hr />
<div>==ChvG==<br />
Acidic condition plays an important role in Agrobacterium-plant interactions (1).<br />
<br />
Numerous studies have investigated the possible connection between the physiological response to acidic conditions and the activity of ChvG and ChvI. Studies have shown that ChvG /ChvI two component systems specifically get induced in an acidic environment (1).<br />
<br />
As a result, there is growing consensus that the ChvG/ChvI, two-component regulatory system, may be the primary global pH sensor in Agrobacterium ''tumefaciens'' and as such, serves in coordinating the expression of acid-induced genes (2).<br />
<br />
==ChvI==<br />
<br />
ChvI is the response regulator of the ChvG/ChvI two component regulatory system found in Agrobacterium (4).<br />
<br />
<br />
===ChvG ---> ChvI===<br />
<br />
ChvG and ChvI is the sensor kinase, and cognate response regulator, respectively, of two component regulatory system in Agrobacterium thought to respond to extracellular pH changes (2).<br />
<br />
<br />
==PhoB==<br />
<br />
PhoB is a response regulator in E.coli that gets activated by a histidine kinase and phosphate sensor called PhoR (6).<br />
<br />
==PhoR==<br />
<br />
PhoR is a histidine kinase and a phosphate sensor in E. ''coli'' that activates a response regulator called PhoB (6).<br />
<br />
===PhoR-->ChvI===<br />
<br />
It has been hypothesized that due to the amino acid similarity of ChvI to PhoB, ChvI is able to act as a phosphate regulatory in E. ''coli'', as a PhoB substitute (3).<br />
<br />
Since ChvI showed no abnormal activity inside Agrobacterium ''tumefaciens'' under phosphate limitation, it cannot serve as the primary phosphate regulator in Agrobacterium (3).<br />
<br />
''Note: ChvI was not required for phosphate regulation in Agrobacterium Tumefaciens (3), but it might be involved in phosphate regulation in Agrobacterium ''tumefaciens'' in some way (3). Therefore, the possibility of crosstalk still exists.''<br />
<br />
<br />
Due to the structural similarity of ChvI and PhoB, the possibility of crosstalk exists (3).<br />
<br />
''Note: Since we are going to use Escherichia ''coli'' as our model chassis for this project; we had to make sure that our proteins will keep their characteristics of our interest after being placed inside the Escherichia ''coli''. For example, VirG gene (promoter 1) after being introduced into E.coli cells, started to behave as if it were a member of phosphate regulation (note: PhoB function was necessary for virG induction by phosphate starvation)(6). Studies have suggested that the reason for this behavior was due to the fact that virG promoter 1 has two hexamer blocks which have the same positions as the pho genes (6). ''<br />
<br />
Studies have shown that ChvI probably activated Alkaline Phosphatase (AP) activity by activating transcription of PhoA (PhoA also gets activated by PhoB) (3). It was not proven that ChvI activates other PhoB-dependent promoters or recognizes the same DNA sequence as PhoB (3).But, there has been speculation that since PhoR mediates PhoB phosphorylation and dephosphorylation, it may do the same to ChvI(3).<br />
<br />
Therefore, the best option would be to find a promoter from Agrobacterium that gets activated by ChvG/ChvI, a two-component regulatory system only; and does not interact with any E. ''coli'' system.<br />
<br />
===PhoR-->PhoB===<br />
<br />
In E. ''coli'', the PhoR/PhoB two component system senses phosphate levels to activate genes that mediate phosphate starvation (3).<br />
<br />
<br />
[[https://2009.igem.org/Team:UC_Davis/About_Us About Us]] [[https://2009.igem.org/Team:UC_Davis/Celiac Celiac Disease]] [[https://2009.igem.org/Team:UC_Davis/Treatments Current Treatment]] [[https://2009.igem.org/Team:UC_Davis/Approach Our Approach]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor pH Sensor]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/parts pH Sensor Parts]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/promoters pH Promoters]] [[https://2009.igem.org/Team:UC_Davis/Secretion/parts Secretion Parts]]</div>Lydiahttp://2009.igem.org/Team:UC_Davis/pH_SensorTeam:UC Davis/pH Sensor2009-09-23T20:19:24Z<p>Lydia: </p>
<hr />
<div>=“Wiring the pH Sensor Into Our System”=<br />
<br />
==The pH Sensor==<br />
<br />
ChvG is a suspected pH sensor and histidine kinase and ChvI is its cognate response regulator (2). Together they are believed to sense extracellular pH between 5.5 to 7 and relaying this signal to regulate the expression of key virulence related genes. (2)<br />
<br />
In our design, we used this two-component system from ''Agrobacterium tumefaciens'', to regulate the expression of select target genes.<br />
<br />
==Possibility of Cross Talk==<br />
<br />
Bacteria have numerous related kinase/response regulated based, two component systems (5). For instance, the ''Agrobacterium tumefaciens'' genome is known to contain at least 25 two-component pathways (2).Therefore, there are possibilities of crosstalk between some genes present in ''E. coli'' that have the same mechanism as ChvI/ChvG system. In fact, there is evidence supporting that ChvI can complement a PhoB deletion in ''E. coli''. (3) Therefore it is highly likely that there is a possibility of cross talk between our new sensory systems elements, present in ''E. coli''.<br />
Many protein-protein interactions are not fully understood (5), which makes this project more challenging but very interesting.<br />
<br />
=Finding an Appropriate Biological pH Sensor=<br />
<br />
==Why is There a Need for a pH Sensor?==<br />
<br />
We would like our probiotic organism to confine itself to the stomach. Since low pH is a hallmark of the stomach, it seems reasonable to assume that linking the production of a suicide gene to a dramatic increase in pH (to values found in the intestine) should allow us to sequester our organism to the stomach. There are no pH sensors available on the registry of biological parts. So, we will engineer a device to sense extracellular pH changes and design it so that it can be used to regulate the expression of any target gene in confinement by pH levels, and then add it to the registry!<br />
Finding an appropriate biological pH sensor:<br />
<br />
We have decided to design our pH sensor by adopting one of the pH sensors of ''Agrobacterium tumefaciens''. The ChvG/ChvI two component system of Agrobacterium is suspected to sense extracellular pH changes (2) and relay this environmental signal to numerous virulence related promoters (2). <br />
<br />
For more information regarding ChvG, go to ChvG<br />
<br />
Once our pH sensor is built, we will need to test its behavior in response to changes to external pH.<br />
<br />
For more information go to assay for pH sensor<br />
<br />
<br />
[[https://2009.igem.org/Team:UC_Davis/About_Us About Us]] [[https://2009.igem.org/Team:UC_Davis/Celiac Celiac Disease]] [[https://2009.igem.org/Team:UC_Davis/Treatments Current Treatment]] [[https://2009.igem.org/Team:UC_Davis/Approach Our Approach]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor pH Sensor]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/parts pH Sensor Parts]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/promoters pH Promoters]] [[https://2009.igem.org/Team:UC_Davis/Secretion/parts Secretion Parts]]</div>Lydiahttp://2009.igem.org/Team:UC_Davis/pH_SensorTeam:UC Davis/pH Sensor2009-09-23T20:19:10Z<p>Lydia: /* “Wiring the pH Sensor Into Our System” */</p>
<hr />
<div>=“Wiring the pH Sensor Into Our System”=<br />
<br />
==The pH Sensor==<br />
<br />
ChvG is a suspected pH sensor and histidine kinase and ChvI is its cognate response regulator (2). Together they are believed to sense extracellular pH between 5.5 to 7 and relaying this signal to regulate the expression of key virulence related genes. (2)<br />
<br />
In our design, we used this two-component system from ''Agrobacterium tumefaciens'', to regulate the expression of select target genes.<br />
<br />
==Possibility of Cross Talk==<br />
<br />
Bacteria have numerous related kinase/response regulated based, two component systems (5). For instance, the ''Agrobacterium tumefaciens'' genome is known to contain at least 25 two-component pathways (2).Therefore, there are possibilities of crosstalk between some genes present in ''E. coli'' that have the same mechanism as ChvI/ChvG system. In fact, there is evidence supporting that ChvI can complement a PhoB deletion in ''E. coli''. (3) Therefore it is highly likely that there is a possibility of cross talk between our new sensory systems elements, present in ''E. coli''.<br />
Many protein-protein interactions are not fully understood (5), which makes this project more challenging but very interesting.<br />
<br />
=Finding an Appropriate Biological pH Sensor=<br />
<br />
==Why is There a Need for a pH Sensor?==<br />
<br />
We would like our probiotic organism to confine itself to the stomach. Since low pH is a hallmark of the stomach, it seems reasonable to assume that linking the production of a suicide gene to a dramatic increase in pH (to values found in the intestine) should allow us to sequester our organism to the stomach. There are no pH sensors available on the registry of biological parts. So, we will engineer a device to sense extracellular pH changes and design it so that it can be used to regulate the expression of any target gene in confinement by pH levels, and then add it to the registry!<br />
Finding an appropriate biological pH sensor:<br />
<br />
We have decided to design our pH sensor by adopting one of the pH sensors of ''Agrobacterium tumefaciens''. The ChvG/ChvI two component system of Agrobacterium is suspected to sense extracellular pH changes (2) and relay this environmental signal to numerous virulence related promoters (2). <br />
<br />
For more information regarding ChvG, go to ChvG<br />
<br />
Once our pH sensor is built, we will need to test its behavior in response to changes to external pH.<br />
<br />
For more information go to assay for pH sensor</div>Lydiahttp://2009.igem.org/Team:UC_Davis/pH_SensorTeam:UC Davis/pH Sensor2009-09-23T20:18:51Z<p>Lydia: /* “Wiring the pH Sensor Into Our System” */</p>
<hr />
<div>=“Wiring the pH Sensor Into Our System”=<br />
<br />
==The pH Sensor==<br />
<br />
ChvG is a suspected pH sensor and histidine kinase and ChvI is its cognate response regulator (2). Together they are believed to sense extracellular pH between 5.5 to 7 and relaying this signal to regulate the expression of key virulence related genes. (2)<br />
<br />
In our design, we used this two-component system from ''Agrobacterium tumefaciens'', to regulate the expression of select target genes.<br />
<br />
==Possibility of Cross Talk==<br />
<br />
Bacteria have numerous related kinase/response regulated based, two component systems (5). For instance, the ''Agrobacterium tumefaciens'' genome is known to contain at least 25 two-component pathways (2).Therefore, there are possibilities of crosstalk between some genes present in ''E. coli'' that have the same mechanism as ChvI/ChvG system. In fact, there is evidence supporting that ChvI can complement a PhoB deletion in ''E. coli''. (3) Therefore it is highly likely that there is a possibility of cross talk between our new sensory systems elements, present in ''E. coli''.<br />
Many protein-protein interactions are not fully understood (5), which makes this project more challenging but very interesting.<br />
<br />
<br />
[[https://2009.igem.org/Team:UC_Davis/About_Us About Us]] [[https://2009.igem.org/Team:UC_Davis/Celiac Celiac Disease]] [[https://2009.igem.org/Team:UC_Davis/Treatments Current Treatment]] [[https://2009.igem.org/Team:UC_Davis/Approach Our Approach]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor pH Sensor]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/parts pH Sensor Parts]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/promoters pH Promoters]] [[https://2009.igem.org/Team:UC_Davis/Secretion/parts Secretion Parts]]<br />
<br />
=Finding an Appropriate Biological pH Sensor=<br />
<br />
==Why is There a Need for a pH Sensor?==<br />
<br />
We would like our probiotic organism to confine itself to the stomach. Since low pH is a hallmark of the stomach, it seems reasonable to assume that linking the production of a suicide gene to a dramatic increase in pH (to values found in the intestine) should allow us to sequester our organism to the stomach. There are no pH sensors available on the registry of biological parts. So, we will engineer a device to sense extracellular pH changes and design it so that it can be used to regulate the expression of any target gene in confinement by pH levels, and then add it to the registry!<br />
Finding an appropriate biological pH sensor:<br />
<br />
We have decided to design our pH sensor by adopting one of the pH sensors of ''Agrobacterium tumefaciens''. The ChvG/ChvI two component system of Agrobacterium is suspected to sense extracellular pH changes (2) and relay this environmental signal to numerous virulence related promoters (2). <br />
<br />
For more information regarding ChvG, go to ChvG<br />
<br />
Once our pH sensor is built, we will need to test its behavior in response to changes to external pH.<br />
<br />
For more information go to assay for pH sensor</div>Lydiahttp://2009.igem.org/Team:UC_Davis/ApproachTeam:UC Davis/Approach2009-09-23T20:18:39Z<p>Lydia: /* Our Approach */</p>
<hr />
<div>=Our Approach=<br />
We are going to break down gluten inside the stomach, because gliadin does not prompt an immune system response until absorption in the small intestines. Another worry is our delivery system taking residence somewhere outside the stomach, so we designed a pH inducible apoptosis system. This is achieved through sensing the change of pH from the stomach to the small intestine, a difference of about four to five.<br />
<br />
Unlike other suggested treatments requiring the consumption of capsules or tablets, our method of using ''Escherichia coli'' is more cost- and time-efficient.<br />
<br />
<br />
==Induced Secretion==<br />
<br />
*Gene sequence for secretion system<br />
*Adding Secretion<br />
*Assay for measuring protein localization and activity<br />
<br />
==Sensing pH and Inducing Cell Death==<br />
<br />
*Finding an appropriate biological pH sensor<br />
*Wiring the pH sensor into our system<br />
*Assay for measuring the behavior of the pH sensor<br />
<br />
<br />
[[https://2009.igem.org/Team:UC_Davis/About_Us About Us]] [[https://2009.igem.org/Team:UC_Davis/Celiac Celiac Disease]] [[https://2009.igem.org/Team:UC_Davis/Treatments Current Treatment]] [[https://2009.igem.org/Team:UC_Davis/Approach Our Approach]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor pH Sensor]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/parts pH Sensor Parts]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/promoters pH Promoters]] [[https://2009.igem.org/Team:UC_Davis/Secretion/parts Secretion Parts]]</div>Lydiahttp://2009.igem.org/Team:UC_Davis/TreatmentsTeam:UC Davis/Treatments2009-09-23T20:11:50Z<p>Lydia: </p>
<hr />
<div>=Current Treatments=<br />
<br />
Currently, there is no cure for this genetic condition. Prevention of the symptoms associated with celiac disease is accomplished through adopting a gluten-free diet (14, 12, 11). However, gluten is present in most everyday diets, making it difficult to have a gluten-free diet.<br />
<br />
"Oral supplementation with prolyl oligopeptidases has therefore been proposed as a potential therapeutic approach."(8) However, enzymes studied earlier were not able to degrade gluten inside stomach before it reaches small intestine because they were "irreversibly inactivated by pepsin and acidic pH, both present in the stomach."(8)<br />
<br />
Over the past years, researchers discovered an enzyme from the bacterium ''Aspergillus niger''. This recently identified prolyl endoprotease was observed to "work optimally at 4-5 pH and [remain] stable at 2 pH".(8) It is possible that this enzyme may provide an alternative treatment for celiac disease. (8) Studies have shown that prolyl endoprotease is able to "degrade gluten ''in vitro'' and under conditions similar to the ones present in the gastrointestinal tract." (8) However, due to licensing restrictions we have opted not to work with this protein.<br />
<br />
In 2007, a study suggested an alternative approach by combining a glutamine-specific endoprotease (EP-B2 from barley) and a prolyl endopeptidase (SC PEP from ''Sphingomonas capsulata''); with gastric activity and complementary substrate specificity there is a possibility of increasing the safe threshold of ingested gluten (12). An advantage of this “combination product is that both enzymes are active and stable in stomach and can therefore be administered as lyophilized powders or simple capsules or tablets” (12).<br />
<br />
''Note: This study was first “evaluated via in vitro digestion of whole-wheat bread and then confirmed by in vivo studies in rats (12), unlike other earlier studies which were performed on synthetic gluten oligopeptides, recombinant gliadin proteins, or uncooked gluten” (12).''<br />
<br />
[[https://2009.igem.org/Team:UC_Davis/About_Us About Us]] [[https://2009.igem.org/Team:UC_Davis/Celiac Celiac Disease]] [[https://2009.igem.org/Team:UC_Davis/Treatments Current Treatment]] [[https://2009.igem.org/Team:UC_Davis/Approach Our Approach]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor pH Sensor]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/parts pH Sensor Parts]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/promoters pH Promoters]] [[https://2009.igem.org/Team:UC_Davis/Secretion/parts Secretion Parts]]</div>Lydiahttp://2009.igem.org/Team:UC_Davis/CeliacTeam:UC Davis/Celiac2009-09-23T20:02:28Z<p>Lydia: /* What happens in celiac disease small intestines? */</p>
<hr />
<div>=What is Celiac Disease?=<br />
<br />
==Description==<br />
Celiac disease is a form of autoimmune disorder that occurs inside the small intestines of about one out of every 133 Americans. Affected individuals cannot properly digest gliadin (a component of gluten), leading to immune responses that may include pain and/or vomiting.<br />
<br />
==What happens in the normal small intestines?==<br />
The small intestines are lined with numerous protrusions called "microvilli", which constitute the brush border membrane. In normal small intestines, small peptides and molecules like water diffuse through the brush border membrane and into the bloodstream. Larger molecules such as gliadin are also allowed to diffuse through, but only after they have been converted through a transcellular route.<br />
<br />
==What happens in celiac disease small intestines?== <br />
The brush border membrane in the small intestines of people suffering from celiac disease allows large molecules such as gliadin to go directly into the blood stream without routing them through the transcellular path. Once gliadin passes through, antigen-presenting cells recognize gliadin as a foreign object and attack. This immune response causes pain and other adverse side-effects.<br />
<br />
Over time, these side-effects may also damage intestinal villi, which are important for absorbing nutrients. Lack of nutrients can lead to other illnesses such as autoimmune thyroid disease, autoimmune liver disease, and rheumatoid arthritis (''diseases in which body immune system attacks healthy cells/tissues'') (14).<br />
<br />
[[https://2009.igem.org/Team:UC_Davis/About_Us About Us]] [[https://2009.igem.org/Team:UC_Davis/Celiac Celiac Disease]] [[https://2009.igem.org/Team:UC_Davis/Treatments Current Treatment]] [[https://2009.igem.org/Team:UC_Davis/Approach Our Approach]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor pH Sensor]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/parts pH Sensor Parts]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/promoters pH Promoters]] [[https://2009.igem.org/Team:UC_Davis/Secretion/parts Secretion Parts]]</div>Lydiahttp://2009.igem.org/Team:UC_Davis/CeliacTeam:UC Davis/Celiac2009-09-23T19:59:21Z<p>Lydia: /* What happens in the normal small intestines? */</p>
<hr />
<div>=What is Celiac Disease?=<br />
<br />
==Description==<br />
Celiac disease is a form of autoimmune disorder that occurs inside the small intestines of about one out of every 133 Americans. Affected individuals cannot properly digest gliadin (a component of gluten), leading to immune responses that may include pain and/or vomiting.<br />
<br />
==What happens in the normal small intestines?==<br />
The small intestines are lined with numerous protrusions called "microvilli", which constitute the brush border membrane. In normal small intestines, small peptides and molecules like water diffuse through the brush border membrane and into the bloodstream. Larger molecules such as gliadin are also allowed to diffuse through, but only after they have been converted through a transcellular route.<br />
<br />
==What happens in celiac disease small intestines?== <br />
The brush border membrane in the small intestines of people suffering from celiac disease allows large molecules such as gliadin to go directly into the blood stream without routing them through the transcellular path. Once gliadin passes through, antigen-presenting cells recognize gliadin as a foreign object and will attack. This immune response causes pain and other adverse side-effects.<br />
<br />
These side-effects over time may also damage intestinal villi, important for absorbing nutrients. Lack of nutrients can lead to other illnesses such as autoimmune thyroid disease, autoimmune liver disease, and rheumatoid arthritis (''diseases in which body immune system attacks healthy cells/tissues'') (14).<br />
<br />
[[https://2009.igem.org/Team:UC_Davis/About_Us About Us]] [[https://2009.igem.org/Team:UC_Davis/Celiac Celiac Disease]] [[https://2009.igem.org/Team:UC_Davis/Treatments Current Treatment]] [[https://2009.igem.org/Team:UC_Davis/Approach Our Approach]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor pH Sensor]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/parts pH Sensor Parts]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/promoters pH Promoters]] [[https://2009.igem.org/Team:UC_Davis/Secretion/parts Secretion Parts]]</div>Lydiahttp://2009.igem.org/Team:UC_Davis/CeliacTeam:UC Davis/Celiac2009-09-23T19:42:21Z<p>Lydia: /* Description */</p>
<hr />
<div>=What is Celiac Disease?=<br />
<br />
==Description==<br />
Celiac disease is a form of autoimmune disorder that occurs inside the small intestines of about one out of every 133 Americans. Affected individuals cannot properly digest gliadin (a component of gluten), leading to immune responses that may include pain and/or vomiting.<br />
<br />
==What happens in the normal small intestines?==<br />
In normal small intestines, the brush border membrane lets small peptides and molecules like water through and into the bloodstream. The brush border membrane also lets larger molecules into the blood stream, similar to gliadin; however, these larger molecules are converted through a transcellular route, which does not prompt an immune system response.<br />
<br />
==What happens in celiac disease small intestines?== <br />
The brush border membrane in the small intestines of people suffering from celiac disease allows large molecules such as gliadin to go directly into the blood stream without routing them through the transcellular path. Once gliadin passes through, antigen-presenting cells recognize gliadin as a foreign object and will attack. This immune response causes pain and other adverse side-effects.<br />
<br />
These side-effects over time may also damage intestinal villi, important for absorbing nutrients. Lack of nutrients can lead to other illnesses such as autoimmune thyroid disease, autoimmune liver disease, and rheumatoid arthritis (''diseases in which body immune system attacks healthy cells/tissues'') (14).<br />
<br />
[[https://2009.igem.org/Team:UC_Davis/About_Us About Us]] [[https://2009.igem.org/Team:UC_Davis/Celiac Celiac Disease]] [[https://2009.igem.org/Team:UC_Davis/Treatments Current Treatment]] [[https://2009.igem.org/Team:UC_Davis/Approach Our Approach]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor pH Sensor]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/parts pH Sensor Parts]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/promoters pH Promoters]] [[https://2009.igem.org/Team:UC_Davis/Secretion/parts Secretion Parts]]</div>Lydiahttp://2009.igem.org/Team:UC_DavisTeam:UC Davis2009-09-23T19:40:43Z<p>Lydia: </p>
<hr />
<div>=Overview=<br />
An estimated one in every 133 Americans suffer from an autoimmune disorder called celiac disease (14, 11). Celiac disease is a condition in which the small intestines are unable to digest gliadin, a protein present in gluten. When gliadin is consumed by an individual with celiac disease, the immune system responds by causing a range of side-effects, which may include abdominal pain, chronic diarrhea, and/or vomiting (14). A friend of one of our teammates currently suffers from this illness, which prompted us to look into celiac disease as the focus of our project. Currently, the only accepted treatment for this disorder is adhering to a gluten-free diet (14, 12). This inspired us to design a probiotic organism that could survive and take residence in the stomach, where it would secrete an enzyme that can degrade gliadin. This would prevent gliadin from progressing through the small intestine and inducing immune system response.<br />
<br />
This synthetically engineered organism should be able to satisfy two main criteria. First, it should be able to survive ''only'' in the stomach and secondly, it should secrete an enzyme that can degrade gliadin. <br />
<br />
Studies have shown that the enzyme, Prolyl-endoprotease, is able to degrade gliadin at the pH level of the stomach (8). Prolyl-endoprotease is one of the most promising enzymes which are being used in recent studies regarding treatment for celiac disease.<br />
<br />
Due to licensing restrictions, we have opted not to work with this protein.<br />
<br />
We have decided to focus our summer project on building two key elements required for our desired organism:<br />
<br />
# An inducible secretion system<br />
# A biological pH sensor limiting this secretion system to the stomach<br />
<br />
The pH system would sense the pH change when the probiotic organism leaves the stomach and prompt apoptosis to occur.<br />
We are confident that our two systems can be built in time to present to the iGEM community.<br />
<br />
[[https://2009.igem.org/Team:UC_Davis/About_Us About Us]] [[https://2009.igem.org/Team:UC_Davis/Celiac Celiac Disease]] [[https://2009.igem.org/Team:UC_Davis/Treatments Current Treatment]] [[https://2009.igem.org/Team:UC_Davis/Approach Our Approach]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor pH Sensor]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/parts pH Sensor Parts]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/promoters pH Promoters]] [[https://2009.igem.org/Team:UC_Davis/Secretion/parts Secretion Parts]]</div>Lydiahttp://2009.igem.org/User:LydiaUser:Lydia2009-09-23T18:48:57Z<p>Lydia: Replacing page with 'My name is Lydia. I am a second year Biomedical Engineering major at UC Davis.'</p>
<hr />
<div>My name is Lydia. I am a second year Biomedical Engineering major at UC Davis.</div>Lydiahttp://2009.igem.org/User:LydiaUser:Lydia2009-09-23T18:40:08Z<p>Lydia: </p>
<hr />
<div>My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia. My name is Lydia.</div>Lydiahttp://2009.igem.org/User:LydiaUser:Lydia2009-09-23T18:37:59Z<p>Lydia: </p>
<hr />
<div>School: UC Davis '12<br />
Sex: Female<br />
Birthday: May 15, 1990<br />
Religious Views: Christian</div>Lydiahttp://2009.igem.org/User:LydiaUser:Lydia2009-09-23T18:36:09Z<p>Lydia: New page: JAMJI!!!!!!</p>
<hr />
<div>JAMJI!!!!!!</div>Lydiahttp://2009.igem.org/Team:UC_Davis/CeliacTeam:UC Davis/Celiac2009-09-23T01:19:00Z<p>Lydia: /* What happens in the normal small intestines? */</p>
<hr />
<div>=What is Celiac Disease?=<br />
<br />
==Description==<br />
Celiac disease is a form of autoimmune disorder that occurs inside the small intestines of about one out of every 133 Americans. Affected individuals cannot properly digest gliadin, a component of gluten, leading to immune responses that may include pain and/or vomiting.<br />
<br />
==What happens in the normal small intestines?==<br />
In normal small intestines, the brush border membrane lets small peptides and molecules like water through and into the bloodstream. The brush border membrane also lets larger molecules into the blood stream, similar to gliadin; however, these larger molecules are converted through a transcellular route, which does not prompt an immune system response.<br />
<br />
==What happens in celiac disease small intestines?== <br />
The brush border membrane in the small intestines of people suffering from celiac disease allows large molecules such as gliadin to go directly into the blood stream without routing them through the transcellular path. Once gliadin passes through, antigen-presenting cells recognize gliadin as a foreign object and will attack. This immune response causes pain and other adverse side-effects.<br />
<br />
These side-effects over time may also damage intestinal villi, important for absorbing nutrients. Lack of nutrients can lead to other illnesses such as autoimmune thyroid disease, autoimmune liver disease, and rheumatoid arthritis (''diseases in which body immune system attacks healthy cells/tissues'') (14).</div>Lydiahttp://2009.igem.org/Team:UC_Davis/CeliacTeam:UC Davis/Celiac2009-09-23T01:16:56Z<p>Lydia: /* Description */</p>
<hr />
<div>=What is Celiac Disease?=<br />
<br />
==Description==<br />
Celiac disease is a form of autoimmune disorder that occurs inside the small intestines of about one out of every 133 Americans. Affected individuals cannot properly digest gliadin, a component of gluten, leading to immune responses that may include pain and/or vomiting.<br />
<br />
==What happens in the normal small intestines?==<br />
In normal small intestines, the brush border membrane lets small peptides and molecules like water through and into the bloodstream. The brush border membrane also lets larger molecules into the blood stream, similar to gliadin; however, these larger molecules are converted through a transcellular route, which does not prompt an immune system response.<br />
<br />
==What happens in celiac disease small intestines?== <br />
The brush border membrane in the small intestines of people suffering from celiac disease allows large molecules such as gliadin to go directly into the blood stream without routing them through the transcellular path. Once gliadin passes through, antigen-presenting cells recognize gliadin as a foreign object and will attack. This immune response causes pain and other adverse side-effects.<br />
<br />
These side-effects over time may also damage intestinal villi, important for absorbing nutrients. Lack of nutrients can lead to other illnesses such as autoimmune thyroid disease, autoimmune liver disease, and rheumatoid arthritis (''diseases in which body immune system attacks healthy cells/tissues'') (14).</div>Lydiahttp://2009.igem.org/Team:UC_Davis/About_UsTeam:UC Davis/About Us2009-09-23T00:36:59Z<p>Lydia: /* About Us */</p>
<hr />
<div>=About Us=<br />
<br />
This is the first time that UC Davis has decided to enter the iGEM competition. This team is comprised of six undergraduate students from three different academic fields of interest: College of Engineering, College of Agricultural and Environmental Sciences, and College of Biological Science. This project has been supervised by Dr. Marc Facciotti and Professor Johnathon Eisen. UC Davis is new to the iGEM competition, but the members are very enthusiastic and ready to work. Furthermore, two of the members on the UC Davis team have competed in iGEM before.</div>Lydiahttp://2009.igem.org/Team:UC_DavisTeam:UC Davis2009-09-23T00:32:54Z<p>Lydia: /* Overview */</p>
<hr />
<div>=Overview=<br />
An estimated one in every 133 Americans suffer from an autoimmune disorder called celiac disease (14, 11). Celiac disease is a condition in which the small intestines are unable to digest gliadin, a protein present in gluten. When gliadin is consumed by an individual with celiac disease, the immune system responds by causing a range of side-effects, which may include abdominal pain, chronic diarrhea, and/or vomiting (14). A friend of one of our teammates currently suffers from this illness, which prompted us to look into celiac disease as the focus of our project. Currently, the only accepted treatment for this disorder is adhering to a gluten-free diet (14, 12). This inspired us to design a probiotic organism that could survive and take residence in the stomach, where it would secrete an enzyme that can degrade gliadin. This would prevent gliadin from progressing through the small intestine and inducing immune system response.<br />
<br />
This synthetically engineered organism should be able to satisfy two main criteria. First, it should be able to survive ''only'' in the stomach and secondly, it should secrete an enzyme that can degrade gliadin. <br />
<br />
Studies have shown that the enzyme, Prolyl-endoprotease, is able to degrade gliadin at the pH level of the stomach (8). Prolyl-endoprotease is one of the most promising enzymes which are being used in recent studies regarding treatment for celiac disease.<br />
<br />
Due to licensing restrictions, we have opted not to work with this protein.<br />
<br />
We have decided to focus our summer project on building two key elements required for our desired organism. Therefore, our project has been split into two tasks:<br />
<br />
# Produce an inducible secretion system<br />
# Construct a biological pH sensor limiting this secretion system to the stomach<br />
<br />
This pH system would sense the pH change when the probiotic organism leaves the stomach.<br />
We are confident that our two systems can be built in time to present to the iGEM community.<br />
<br />
[[https://2009.igem.org/Team:UC_Davis/About_Us About Us]] [[https://2009.igem.org/Team:UC_Davis/Celiac Celiac Disease]] [[https://2009.igem.org/Team:UC_Davis/Treatments Current Treatment]] [[https://2009.igem.org/Team:UC_Davis/Approach Our Approach]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor pH Sensor]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/parts pH Sensor Parts]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/promoters pH Promoters]] [[https://2009.igem.org/Team:UC_Davis/Secretion/parts Secretion Parts]]</div>Lydiahttp://2009.igem.org/Team:UC_DavisTeam:UC Davis2009-09-23T00:19:57Z<p>Lydia: /* Overview */</p>
<hr />
<div>=Overview=<br />
An estimated one in every 133 Americans suffer from an autoimmune disorder called celiac disease (14, 11). Celiac disease is a condition in which the small intestines are unable to digest gliadin, a protein present in gluten. When gliadin is consumed by an individual with celiac disease, the immune system responds by causing a range of side-effects, which may include abdominal pain, chronic diarrhea and vomiting (14). A friend of one of our teammates currently suffers from this illness, which prompted us to look into celiac disease as the focus of our project. Currently, the only accepted treatment for this disorder is adhering to a gluten-free diet (14, 12). This inspired us to design a probiotic organism that could survive and take residence in the stomach, where it would secrete an enzyme that can degrade gliadin. This would prevent gliadin from progressing through the small intestine and inducing immune system response.<br />
<br />
This synthetically engineered organism should be able to satisfy at least two main criteria. First, it should be able to survive ''only'' in the stomach and, secondly, secrete an enzyme that can degrade gliadin. <br />
<br />
Studies have shown that the enzyme, Prolyl-endoprotease is able to degrade gliadin at the pH level of the stomach (8). Prolyl-endoprotease is one of the most promising enzymes which being used in recent studies regarding celiac disease treatment.<br />
<br />
Due to licensing restrictions, we have opted not to work with this protein.<br />
<br />
We have decided to focus our summer project on building two key elements required for our desired organism. Therefore, our project has been split into two tasks:<br />
<br />
# Produce an inducible secretion system<br />
# Construct a biological pH sensor limiting this secretion system to the stomach<br />
<br />
This pH system will be sensitive to the change in pH when the probiotic leaves the stomach.<br />
We are confident that our two systems can be built in time to present to the iGEM community.<br />
<br />
[[https://2009.igem.org/Team:UC_Davis/About_Us About Us]] [[https://2009.igem.org/Team:UC_Davis/Celiac Celiac Disease]] [[https://2009.igem.org/Team:UC_Davis/Treatments Current Treatment]] [[https://2009.igem.org/Team:UC_Davis/Approach Our Approach]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor pH Sensor]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/parts pH Sensor Parts]] [[https://2009.igem.org/Team:UC_Davis/pH_Sensor/promoters pH Promoters]] [[https://2009.igem.org/Team:UC_Davis/Secretion/parts Secretion Parts]]</div>Lydia