Team:UC Davis/Parts

From 2009.igem.org

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href="https://2009.igem.org/Team:UC_Davis/Parts"><img alt=""
href="https://2009.igem.org/Team:UC_Davis/Parts"><img alt=""
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"
src="https://static.igem.org/mediawiki/2009/a/a6/UCDAVIS_PIC6.png"
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style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a
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style="border: 0px solid ; width: 78px; height: 37px;"></a></big></big></big></big></span></b><b
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href="https://2009.igem.org/Team:UC_Davis/Contacts_References"><img
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alt="" src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"
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style="border: 0px solid ; width: 83px; height: 37px;"></a></big></big></big></big></span></b><b
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<hr style="width: 100%; height: 2px;"><big><big><big><span
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<hr style="width: 100%; height: 2px;">
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style="text-decoration: underline; font-weight: bold;"><small>Secretion
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<div style="text-align: left;"><big><big><b
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Model
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style="color: rgb(0, 0, 0); text-decoration: underline;">Parts:</b></big></big><b
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1:<br>
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style="color: rgb(0, 0, 0);">&nbsp;&nbsp; <br>
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<br>
<br>
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Parts related to
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</small></span></big></big></big><big><span style="font-style: italic;">Click
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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;&nbsp;&nbsp;&nbsp;
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on an
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Parts related to pH sensor:<br>
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individual part for more information.<br>
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</b>
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</span></big>
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<table
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<div style="text-align: center;"><big><big><big><span
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style="text-align: left; width: 100%; background-color: rgb(255, 255, 0);"
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style="font-weight: bold;"><small><span
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border="2" cellpadding="2" cellspacing="2">
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style="text-decoration: underline; background-color: rgb(255, 255, 0);"><a
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<tbody>
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href="#LacI"><img alt=""
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<tr>
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src="https://static.igem.org/mediawiki/2009/4/4e/UCDAVIS_PROMOTER_1.png"
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<td style="vertical-align: top;">Proteins:<br>
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style="border: 0px solid ; width: 242px; height: 58px; background-color: rgb(255, 255, 255);"></a><a
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</td>
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href="#RBS"><img alt=""
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<td style="vertical-align: top;">Promoters:<br>
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src="https://static.igem.org/mediawiki/2009/1/15/UCDAVIS_RBS1.png"
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</td>
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style="border: 0px solid ; width: 89px; height: 81px; background-color: rgb(255, 255, 255);"></a><a
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<td style="vertical-align: top;">Others:<br>
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href="#INPNC"><img alt=""
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</td>
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src="https://static.igem.org/mediawiki/2009/9/9c/UCDAVIS_ompA.png"
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<td style="vertical-align: top;">Proteins:<br>
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style="border: 0px solid ; width: 106px; height: 59px; background-color: rgb(255, 255, 255);"></a><a
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</td>
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href="#SS"><img alt=""
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<td style="vertical-align: top;">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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src="https://static.igem.org/mediawiki/2009/9/9d/UCDAVIS_SS1.png"
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Promoters:<br>
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style="border: 0px solid ; width: 94px; height: 69px; background-color: rgb(255, 255, 255);"></a><img
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</td>
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style="width: 106px; height: 59px; background-color: rgb(255, 255, 255);"
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</tr>
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alt="" src="https://static.igem.org/mediawiki/2009/0/0c/UCDAVIS_GENE1.png"><a
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<tr>
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href="#His"><img alt=""
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<td style="vertical-align: top;">
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src="https://static.igem.org/mediawiki/2009/c/c0/UCDAVIS_6HIS.png"
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<ul>
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style="border: 0px solid ; width: 134px; height: 66px; background-color: rgb(255, 255, 255);"></a><a
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<li><a href="#INPNC">INPNC</a></li>
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href="#Terminator"><img alt=""
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<li><a href="#OmpA">OmpA</a></li>
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src="https://static.igem.org/mediawiki/2009/c/ca/UCDAVIS_STOP.png"
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<li><a href="#Luciferase">Luciferase</a></li>
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style="border: 0px solid ; width: 94px; height: 83px; background-color: rgb(255, 255, 255);"></a><br>
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<li><a href="#GFP">GFP</a></li>
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</span></small></span></big></big></big><big><big><big><span
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</ul>
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style="font-weight: bold;"><small><span
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</td>
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style="text-decoration: underline; background-color: rgb(255, 255, 0);"><br>
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<td style="vertical-align: top;">
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</span>a.<a href="#LacI"><span style="text-decoration: underline;"><img
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<ul>
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alt=""
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<li><a href="#LacI">LacI</a></li>
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src="https://static.igem.org/mediawiki/2009/4/4e/UCDAVIS_PROMOTER_1.png"
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</ul>
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style="border: 0px solid ; width: 200px; height: 52px;"></span></a><span
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</td>
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style="text-decoration: underline; background-color: rgb(255, 255, 0);"><a
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<td style="vertical-align: top;">
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href="#RBS"><img alt=""
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<ul>
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src="https://static.igem.org/mediawiki/2009/1/15/UCDAVIS_RBS1.png"
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<li><a href="#SS">SS:signal sequence</a></li>
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style="border: 0px solid ; width: 69px; height: 63px; background-color: rgb(255, 255, 255);"></a><a
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<li><a href="#His">6-His-Tag</a></li>
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href="#INPNC"><img alt=""
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<li><a href="#RBS">RBS</a></li>
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src="https://static.igem.org/mediawiki/2009/9/9c/UCDAVIS_ompA.png"
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<li><a href="#Terminator">Terminator</a></li>
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style="border: 0px solid ; width: 106px; height: 51px; background-color: rgb(255, 255, 255);"></a><a
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</ul>
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href="#SS"><img alt=""
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</td>
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src="https://static.igem.org/mediawiki/2009/9/9d/UCDAVIS_SS1.png"
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<td style="vertical-align: top;">
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style="border: 0px solid ; width: 73px; height: 65px; background-color: rgb(255, 255, 255);"></a><a
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<ul>
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href="#GFP"><img alt=""
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<li>ChvI</li>
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src="https://static.igem.org/mediawiki/2009/7/7d/UCDAVIS_GFP.png"
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<li>ChvG</li>
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style="border: 0px solid ; width: 83px; height: 55px; background-color: rgb(255, 255, 255);"></a><a
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</ul>
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href="#His"><img alt=""
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</td>
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src="https://static.igem.org/mediawiki/2009/c/c0/UCDAVIS_6HIS.png"
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<td style="vertical-align: top;">
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style="border: 0px solid ; width: 110px; height: 58px; background-color: rgb(255, 255, 255);"></a><a
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<ul>
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href="#Terminator"><img alt=""
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<li><a href="#ChvI_promoter">ChvI promoter</a></li>
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src="https://static.igem.org/mediawiki/2009/c/ca/UCDAVIS_STOP.png"
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<li><a href="#katA">KatA promoter</a></li>
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style="border: 0px solid ; width: 78px; height: 71px; background-color: rgb(255, 255, 255);"></a></span></small></span></big></big></big><br>
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<li><a href="#aopB">AopB promoter</a></li>
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<div style="text-align: center;"><big><big><big><span
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<li><a href="#PhoA">PhoA
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style="font-weight: bold;"><small>b.<span
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promoter</a>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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style="text-decoration: underline;"><a href="#LacI"><img alt=""
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&nbsp;&nbsp; </li>
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src="https://static.igem.org/mediawiki/2009/4/4e/UCDAVIS_PROMOTER_1.png"
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<li>I<a href="#impA">mpA promoter</a></li>
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style="border: 0px solid ; width: 195px; height: 52px;"></a><a
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</ul>
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href="#RBS"><img alt=""
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</td>
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src="https://static.igem.org/mediawiki/2009/1/15/UCDAVIS_RBS1.png"
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</tr>
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style="border: 0px solid ; width: 68px; height: 63px;"></a><a
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</tbody>
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href="#INPNC"><img alt=""
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</table>
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src="https://static.igem.org/mediawiki/2009/9/9c/UCDAVIS_ompA.png"
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<b style="color: rgb(0, 0, 0);">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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style="border: 0px solid ; width: 105px; height: 51px;"></a><a
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</b><br>
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href="#SS"><img alt=""
 +
src="https://static.igem.org/mediawiki/2009/9/9d/UCDAVIS_SS1.png"
 +
style="border: 0px solid ; width: 73px; height: 65px;"></a><a
 +
href="#Luciferase"><img alt=""
 +
src="https://static.igem.org/mediawiki/2009/9/98/UCDAVIS_Luciferase.png"
 +
style="border: 0px solid ; width: 87px; height: 49px;"></a><a
 +
href="#His"><img alt=""
 +
src="https://static.igem.org/mediawiki/2009/c/c0/UCDAVIS_6HIS.png"
 +
style="border: 0px solid ; width: 110px; height: 58px;"></a><a
 +
href="#Terminator"><img alt=""
 +
src="https://static.igem.org/mediawiki/2009/c/ca/UCDAVIS_STOP.png"
 +
style="border: 0px solid ; width: 78px; height: 71px;"></a></span></small></span></big></big></big><big><big><big><span
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style="font-weight: bold;"><small><span
 +
style="text-decoration: underline; background-color: rgb(255, 255, 0);"></span></small></span></big></big></big></div>
 +
</div>
 +
<div style="text-align: center;"><big><span style="font-style: italic;"></span></big><br>
 +
</div>
 +
<big><big><big><span
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style="text-decoration: underline; font-weight: bold;"></span></big></big></big>
<hr style="width: 100%; height: 2px;">
<hr style="width: 100%; height: 2px;">
<p class="MsoNormal" style="line-height: normal;"><b><span
<p class="MsoNormal" style="line-height: normal;"><b><span
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style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">INPNC:</span></b><span
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style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><a
 +
name="INPNC"></a>INPNC:</span></b><span
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">Ice-nucleation
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">Ice-nucleation
protein (INP) from Pseudomonas
protein (INP) from Pseudomonas
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">
<hr align="center" size="2" width="100%"></span></div>
<hr align="center" size="2" width="100%"></span></div>
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<p class="MsoNormal" style="line-height: normal;"><a
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<b><i><span
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name="ChvI_promoter"></a><b><span
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style="font-size: 12pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></i></b><big><big><big><span
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style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">ChvI
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style="text-decoration: underline; font-weight: bold;"></span></big></big></big>
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promoter</span></b><span
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style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">: </span><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">Gene
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fusion studies confirmed that ChvI gene
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was induced by acidic conditions (1). Also, it has been known to
+
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implicate in
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virulence (1). This gene is one of the candidates to be use in our
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biological
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pH sensor as a promoter.<o:p></o:p></span></p>
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<div class="MsoNormal"
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style="margin-bottom: 0.0001pt; text-align: center; line-height: normal;"
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align="center"><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">
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<hr align="center" size="2" width="100%"></span></div>
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<p class="MsoNormal" style="line-height: normal;"><a name="katA"></a><b><span
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style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">KatA
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promoter</span></b><span
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style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"> :</span><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">This
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Chromosomal gene is located on the linear
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chromosome (2) and it seems to be induced under an acidic environment
+
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as well
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as being involved in the <i>Agrobacterium tumorigenesis</i>
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(2).Research has
+
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suggested that ChvG is needed for "responsiveness of&nbsp; gene
+
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expression
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to low pH "(2). This gene has become a candidate to complete our pH
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sensor
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device from this evidence.<o:p></o:p></span></p>
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<div class="MsoNormal"
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style="margin-bottom: 0.0001pt; text-align: center; line-height: normal;"
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align="center"><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">
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<hr align="center" size="2" width="100%"></span></div>
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<p class="MsoNormal" style="line-height: normal;"><a name="aopB"></a><b><span
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style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">AopB
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promoter</span></b><span
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style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">: </span><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">This
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Chromosomal gene located on the circular
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chromosome (2) encodes an outer member protein exposed on the bacterial
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cell
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surface (2). Also, ChvG was shown to be absolutely required for this
+
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gene
+
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expression (2)It seems to get induced under an acidic environment as
+
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well as
+
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being involved in the <i>Agrobacterium</i> <i>tumorigenesis </i>(2).
+
-
Therefore,
+
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we have chosen this gene to be one of our candidates to complete our pH
+
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sensor
+
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device.<o:p></o:p></span></p>
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<div class="MsoNormal"
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style="margin-bottom: 0.0001pt; text-align: center; line-height: normal;"
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align="center"><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">
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<hr align="center" size="2" width="100%"></span></div>
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<p class="MsoNormal" style="line-height: normal;"><a name="PhoA"></a><b><span
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style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">PhoA
+
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promoter</span></b><span
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style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">: </span><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">There
+
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has been a suggestion that ChvI can
+
-
activate AP activity by activating transcription of this gene, PhoA
+
-
(3).
+
-
Therefore, this gene has become one of our candidates to complete our
+
-
pH sensor
+
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device.<o:p></o:p></span></p>
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<div class="MsoNormal"
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style="margin-bottom: 0.0001pt; text-align: center; line-height: normal;"
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align="center"><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">
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<hr align="center" size="2" width="100%"></span></div>
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<p class="MsoNormal" style="line-height: normal;"><a name="impA"></a><b><span
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style="font-size: 13pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">ImpA
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promoter:</span></b><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">Gene
+
-
fusion studies confirmed that impA genes
+
-
was induced by acidic conditions (1), therefore, this is one of our
+
-
candidates
+
-
to complete our pH sensor device.<o:p></o:p></span></p>
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-
<p class="MsoNormal" style="line-height: normal;"><span
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style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">&nbsp;<o:p></o:p></span></p>
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<p class="MsoNormal"><span
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style="font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p>&nbsp;</o:p></span></p>
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<hr style="width: 100%; height: 2px;"><small style="font-weight: bold;"><span
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style="font-size: 18pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"></span></small></div>
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Revision as of 04:19, 26 September 2009

Model 1

=
Secretion Model 1:

Click on an individual part for more information.


a.
b.

INPNC: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.

We have modified this protein to Biobrick standard, Tom Knights Standard.

OmpA: 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.

We have modified this protein to Biobrick standard, Tom Knights Standard.

Note: “It has remained essentially unknown how proteins of E. coli outer membrane are sorted and incorporated into this membrane” (10)

For more information go to: http://partsregistry.org/wiki/index.php?title=Part:BBa_J36836

RBS:  Ribosome Binding site number 32 (BBa_J61132) from the registry is being used in our secretion system.

For more information go to: http://partsregistry.org/wiki/index.php/Part:BBa_J61132

Terminator: We are using BBa_B0015, a double terminator, as our terminator in both our secretion and pH system.

For more information go to: http://partsregistry.org/wiki/index.php?title=Part:BBa_B0015

GFP: We are using Green Fluorescent Protein as a reporter that also serves as a small protein in testing our secretion system.

Luciferase: Luciferase is a firefly protein that also fluoresces, so it serves as a reporter as well as a testable large protein.

LacI: One inducible Promoter which was found in the part registry.

More can be found in: http://partsregistry.org/wiki/index.php?title=Part:BBa_R0010

SS: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.

We have modified this protein to Biobrick standard, Tom Knights Standard.

6-His Tag:The 6-Histidine Tag serves as a tag for Western Blotting if our fluorescent reporters are not expressed as highly as we would like.

Note: We are using this tag, just in case if the GFP or Luciferase does not work under a plate reader.

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