PennState/Project/Diauxie/PromoterTest

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'''Promoter Test Constructs'''
'''Promoter Test Constructs'''
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To test each of the four xyl promoters, each is placed in a construct followed by a ribosome binding site, the reporter green florescent protein, and a terminator. The strength of each is measured by gfp emission.
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The xyl bidirectional promoter region promotes in the direction of the genes encoding for xylAB and xylFGH metabolization and transport proteins. The CPR-cAMP, and xylR binding sites regulate both directions. This means that there are four possibilities for making promoters: xylF facing including the CRP-cAMP binding site, xylF facing without the CRP-cAMP binding site, xylA facing including the CRP-cAMP binding site, and xylA facing without the CRP-cAMP binding site.  
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<html><img src="http://openwetware.org/images/8/88/Pctest.jpg" id="logo" name="logo" height="150" width="300" alt="" /></html>This construct uses the xylF and xylA facing promoters that includes the CRP binding region.
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To test the four xyl promoters, each is placed in a construct followed by a ribosome binding site, the reporter green florescent protein, and a terminator. The strength of each is measured by gfp emission.
-
<html><img src="http://openwetware.org/images/b/b1/Fpnctest.jpg" id="logo" name="logo" height="150" width="300" alt="" /></html>This construct uses the xylA facing promoter that does not include the CRP binding region.
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<html><img src="http://openwetware.org/images/8/88/Pctest.jpg" id="logo" name="logo" height="150" width="300" alt="" /></html>This construct uses the xylF and xylA facing promoters that include the CRP-cAMP binding region.
-
After each of the four promoter constructs are tested individually, the two
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<html><img src="http://openwetware.org/images/d/d1/Pnctest.jpg" id="logo" name="logo" height="150" width="300" alt="" /></html>This construct uses the xylF and xylA facing promoters that do not include the CRP-cAMP binding region.
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 +
 
 +
After each of the four promoter constructs are tested individually, the two which include the CRP-cAMP are changed so that a gene expressing the mutant CRP* is included. This would act as an effective increase in the CRP-cAMP levels in the cell, increasing the strength of the promoter through positive feedback.
 +
 
 +
<html><img src="http://openwetware.org/images/9/99/Pctestcrp.jpg" id="logo" name="logo" height="150" width="420" alt="" /></html>
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 +
This construct uses the xylF and xylA facing promoters that include the CRP-cAMP binding region followed by the CRP* gene.
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<html>
 +
<!-- START OF LUC'S CODE -->
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<!-- START CSS: the following css needs to be on the page to style the podcast divs properly. Diagonal span background based on www.mezzoblue.com/zengarden/alldesigns/ -->
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<style type="text/css">
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.column {
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width:  200px;
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position: relative;
 +
display: block;
 +
float: left;
 +
clear: none;
 +
margin-right: 10px;
 +
}
 +
.podcast{
 +
position: relative;
 +
float: none;
 +
border: 1px dotted #c7cbce;
 +
height: 110px;
 +
background-color: #f1f1f1;
 +
clear: both;
 +
margin-bottom: 42px;
 +
font: 12px/1.4em "Lucida Grande", "Lucida Sans Unicode", Verdana, Tahoma, Arial, sans-serif;
 +
padding: 0;
 +
width: 175px;
 +
}
 +
.podcast h3{
 +
color: #3d3d3d;
 +
margin: .2em 0 .5em;
 +
padding: 0 0 0 4px;
 +
font-family: Verdana, sans-serif;
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font-size: 100%;
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}
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.podcast a:link, .podcast a:visited{
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text-decoration: none;
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color: #3d3d3d;
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}
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.podcast a:hover {
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color: #aaaaff;
 +
}
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a:hover.screenshot {
 +
}
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.podcast .screenshot {
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float: left;
 +
position: relative;
 +
clear: none;
 +
display: block;
 +
margin: 0px 0px 0px 16px;
 +
}
 +
.podcast .screenshot img {
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background: #ffffff;
 +
border: 1px solid #c7cbce;
 +
padding: 4px;
 +
}
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.podcast .screenshot span {
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position: absolute;
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bottom:0px;
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right: 1px;
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padding: 4px 20px 4px 26px;
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background: url(https://static.igem.org/mediawiki/2007/0/09/Podcast-download-bg.gif) no-repeat;
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font-size: 0.8em;
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line-height: 1;
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}
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.podcast .screenshot span a{
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padding: 0 0 0 0.5em;
 +
}
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.podcast .description {
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display: block;
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position: relative;
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float: left;
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width: 220px;
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margin-left: 3px;
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padding-left: 1.2em;
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}
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.podcast .description a{
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border-bottom: 1px dotted #aaaaff;
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}
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.podcast .description a:hover{
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border-bottom: 1px dotted #aaaaff;
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}
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.screenshot img:hover {
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border: 1px solid #aaaaff;
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}
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</style></html>
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<html>
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<!-- PODCAST HTML: the following html structures the information for each podcast. Each podcast entry is contained in its own podcast div, which are arranged into two columns by two column divs.  The last column must be cleared.-->
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<div class="column">
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<div class="podcast">
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<!-- this is where you need to post the link to where it will go (the second one)-->
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<h3><a rel="enclosure" href="https://2007.igem.org/PennState/Project/Diauxie" ="window.popup_player_258600 = window.open('https://2007.igem.org/PennState/Project/Diauxie','post_258600','toolbar=no,scrollbars=no,directories=no,resizable=yes,width=360,height=305,top=20,left=20,location=no,menubar=no,status=yes,'); return false;">
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Introduction
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</a></h3>
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<div class="screenshot">
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<!-- little white box thing --> <a rel="enclosure" href="https://2007.igem.org/PennState/Project/Diauxie"
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<img src="http://openwetware.org/images/9/97/Psuigem2007promoterimage.jpg" width="133" height="100" alt="" />
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</a>
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<span class="download">
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<a href="http://openwetware.org/images/9/97/Psuigem2007promoterimage.jpg">Image Link</a>
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</span>
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</div>
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</div>
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 +
</div><div class="column">
 +
 
 +
<div class="podcast">
 +
<!-- this is where you need to post the link to where it will go (the second one)-->
 +
<h3><a rel="enclosure" href="https://2007.igem.org/PennState/Project/Diauxie/PromoterTest" ="window.popup_player_258600 = window.open('https://2007.igem.org/PennState/Project/Diauxie/PromoterTest','post_258600','toolbar=no,scrollbars=no,directories=no,resizable=yes,width=360,height=305,top=20,left=20,location=no,menubar=no,status=yes,'); return false;">
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Promoter Test Constructs
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</a></h3>
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<div class="screenshot">
 +
<!-- little white box thing --> <a rel="enclosure" href="https://2007.igem.org/PennState/Project/Diauxie/PromoterTest"
 +
<img src="http://openwetware.org/images/b/be/Psuigem2007RBSimage.jpg" width="133" height="100" alt="" />
 +
</a>
 +
<span class="download">
 +
<a href="http://openwetware.org/images/b/be/Psuigem2007RBSimage.jpg">Image Link</a>
 +
</span>
 +
</div>
 +
</div>
 +
</div>
 +
<div class="column"><div class="podcast">
 +
<!-- this is where you need to post the link to where it will go (the second one)-->
 +
<h3><a rel="enclosure" href="https://2007.igem.org/PennState/Project/Diauxie/DiauxieElimination" ="window.popup_player_258600 = window.open('http://openwetware.org/images/c/c9/XylRdimer1.png','post_258600','toolbar=no,scrollbars=no,directories=no,resizable=yes,width=360,height=305,top=20,left=20,location=no,menubar=no,status=yes,'); return false;">
 +
Eliminating Diauxie
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</a></h3>
 +
<div class="screenshot">
 +
<!-- little white box thing --> <a rel="enclosure" href="https://2007.igem.org/PennState/Project/Diauxie/DiauxieElimination"
 +
<img src="http://openwetware.org/images/2/21/Psuigem2007geneimage.jpg" width="133" height="100" alt="" />
 +
</a>
 +
<span class="download">
 +
<a href="http://openwetware.org/images/2/21/Psuigem2007geneimage.jpg">Image Link</a>
 +
</span>
 +
</div>
 +
</div>
 +
</div>
 +
<div class="column"><div class="podcast">
 +
<!-- this is where you need to post the link to where it will go (the second one)-->
 +
<h3><a rel="enclosure" href="https://2007.igem.org/PennState/Project/Diauxie/FinalConstructs" ="window.popup_player_258600 = window.open('https://2007.igem.org/PennState/Project/Diauxie/FinalConstructs','post_258600','toolbar=no,scrollbars=no,directories=no,resizable=yes,width=360,height=305,top=20,left=20,location=no,menubar=no,status=yes,'); return false;">
 +
Final Constructs
 +
</a></h3>
 +
<div class="screenshot">
 +
<!-- little white box thing --> <a rel="enclosure" href="https://2007.igem.org/PennState/Project/Diauxie/FinalConstructs"
 +
<img src="http://openwetware.org/images/9/9b/Psuigem2007TTimage.jpg" width="133" height="100" alt="" />
 +
</a>
 +
<span class="download">
 +
<a href="http://openwetware.org/images/9/9b/Psuigem2007TTimage.jpg">Image Link</a>
 +
</span>
 +
</div>
 +
</div>
 +
 
 +
<div style="clear:both"></div>
 +
<!-- END OF LUC'S CODE -->
 +
</html>

Latest revision as of 03:05, 27 October 2007

Promoter Test Constructs

The xyl bidirectional promoter region promotes in the direction of the genes encoding for xylAB and xylFGH metabolization and transport proteins. The CPR-cAMP, and xylR binding sites regulate both directions. This means that there are four possibilities for making promoters: xylF facing including the CRP-cAMP binding site, xylF facing without the CRP-cAMP binding site, xylA facing including the CRP-cAMP binding site, and xylA facing without the CRP-cAMP binding site.


To test the four xyl promoters, each is placed in a construct followed by a ribosome binding site, the reporter green florescent protein, and a terminator. The strength of each is measured by gfp emission.


This construct uses the xylF and xylA facing promoters that include the CRP-cAMP binding region.


This construct uses the xylF and xylA facing promoters that do not include the CRP-cAMP binding region.


After each of the four promoter constructs are tested individually, the two which include the CRP-cAMP are changed so that a gene expressing the mutant CRP* is included. This would act as an effective increase in the CRP-cAMP levels in the cell, increasing the strength of the promoter through positive feedback.

This construct uses the xylF and xylA facing promoters that include the CRP-cAMP binding region followed by the CRP* gene.


Introduction

Image Link

Promoter Test Constructs

Image Link

Eliminating Diauxie

Image Link

Final Constructs

Image Link

Retrieved from "http://2007.igem.org/wiki/index.php/PennState/Project/Diauxie/PromoterTest"