Team:UC Davis/Adding secretion

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style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a
style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a
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href="https://2009.igem.org/Team:UC_Davis/homepage2"><img alt=""
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href="https://2009.igem.org/Team:UC_Davis"><img alt=""
src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC3.png"
src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC3.png"
style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a
style="border: 0px solid ; width: 83px; height: 36px;"></a> </big></big></big></big></span></b><a
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href="https://2009.igem.org/Team:UC_Davis/About_Us1"><b
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href="https://2009.igem.org/Team:UC_Davis/About_Us"><b
style="color: rgb(255, 255, 153);"><span
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style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><img
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style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a
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href="https://2009.igem.org/Team:UC_Davis/Project1"><img alt=""
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href="https://2009.igem.org/Team:UC_Davis/Project"><img alt=""
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"
src="https://static.igem.org/mediawiki/2009/b/b9/UCDAVIS_PIC8.png"
style="border: 0px solid ; width: 78px; height: 36px;"></a> </big></big></big></big></span></b><b
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style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a
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href="https://2009.igem.org/Team:UC_Davis/Notebook1"><img alt=""
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href="https://2009.igem.org/Team:UC_Davis/Notebook"><img alt=""
src="https://static.igem.org/mediawiki/2009/2/2f/UCDAVIS_PIC5.png"
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style="font-size: 13.5pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><big><big><big><a
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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"
style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a
style="border: 0px solid ; width: 78px; height: 37px;"></a>&nbsp;<a
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href="https://2009.igem.org/Team:UC_Davis/Contact1"><img alt=""
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href="https://2009.igem.org/Team:UC_Davis/Contacts_References"><img
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src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"
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alt="" src="https://static.igem.org/mediawiki/2009/1/1d/UCDAVIS_PIC7.png"
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alt="" src="https://static.igem.org/mediawiki/2009/c/ca/UCDAVIS_STOP.png"><br>
alt="" src="https://static.igem.org/mediawiki/2009/c/ca/UCDAVIS_STOP.png"><br>
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<br>
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</span></small></span><small><small><span
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</span></small></span></big></big></big>
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style="text-decoration: underline;"><br>
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</span></small></small></big></big></big>
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<p class="MsoNormal" style="text-align: left;"><big><span style="">&nbsp;&nbsp;&nbsp;
 +
&nbsp;&nbsp;&nbsp; &nbsp;&nbsp;&nbsp; <span
 +
style="font-family: Times New Roman,Times,serif;">The purpose of the
 +
secretion system is to
 +
introduce a method of secreting target proteins we wish to synthesize
 +
in the
 +
marvelous host: <i>E.coli</i>. Park </span><i
 +
style="font-family: Times New Roman,Times,serif;">et.
 +
al</i><span style="font-family: Times New Roman,Times,serif;"> showed
 +
that the truncated form of ice nucleation
 +
protein (INPNC) containing the complete coding region of phaZ1,
 +
including its
 +
signal sequence, could lead to stable secretion of the enzyme (</span><a
 +
style="font-family: Times New Roman,Times,serif;"
 +
href="https://2009.igem.org/Team:UC_Davis/Contacts_References">15</a><span
 +
style="font-family: Times New Roman,Times,serif;">). We have
 +
decided to see if we could co-opt this system to make a more
 +
generalized
 +
inducible system for protein secretion in which the INPNC and phaZ1
 +
signal
 +
sequence serve as carriers for a target protein.</span><o:p
 +
style="font-family: Times New Roman,Times,serif;"></o:p></span></big></p>
 +
<p class="MsoNormal" style="font-family: Times New Roman,Times,serif;"><big><span
 +
style=""><br>
 +
<big><b>Possible challenges</b>:</big><o:p></o:p></span></big></p>
<p class="MsoNormal"
<p class="MsoNormal"
-
style="margin-bottom: 0.0001pt; line-height: normal; text-align: center;"><b><span
+
style="margin-left: 30pt; text-align: left; font-family: Times New Roman,Times,serif;"><big><span
-
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">Advantages
+
style="">1. Efficient recognition of the cleavage site may require<span
-
of having several models:</span></b><b><span
+
style="-moz-background-clip: -moz-initial; -moz-background-origin: -moz-initial; -moz-background-inline-policy: -moz-initial; background-attachment: scroll;"><span
-
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p></o:p></span></b></p>
+
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>
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<div style="text-align: left;"></div>
+
some of the non-signal sequence portion of the phaZ1 protein (we have
-
<p class="MsoNormal" style="line-height: normal; text-align: left;"><span
+
only cloned the
-
style="font-size: 12pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">&nbsp;&nbsp;&nbsp;
+
signal sequence up to the cleavage site).<br>
-
&nbsp;&nbsp; In
+
2. The system may only work for proteins in a narrow site range.<br>
-
our secretion system, we are using two genes with different sizes as
+
3. The expression levels of the system may be low.<o:p></o:p></span></big></p>
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our target
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<p class="MsoNormal"
-
secretion genes, GFP being short in length and Luciferase being
+
style="text-align: center; font-family: Times New Roman,Times,serif;"
-
comparably long
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align="center"><big><b><span style=""><br>
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would test our secretion system and its ability to secrete small and
+
<big>Advantages:</big></span></b><span style=""><o:p></o:p></span></big></p>
-
large
+
<ol style="text-align: left;" start="1" type="1">
-
proteins. <br>
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<li style="font-family: Times New Roman,Times,serif;"
-
&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; Testing multiple different combinations;
+
class="MsoNormal"><big><span style=""><u1:p></u1:p>&nbsp; In our
-
Signal
+
secretion system, we are using two genes with different sizes as our
-
Sequence alone, Signal Sequence plus ompA/INPNC and ompA/INPNC alone,
+
target secretion genes; GFP being short in length and Luciferase being
-
would
+
comparably long, these would test the ability of our secretion system
-
help us find the best combination for our secretion system based on
+
to secrete proteins of different sizes. <o:p></o:p></span></big></li>
-
their
+
<li style="font-family: Times New Roman,Times,serif;"
-
secretion ability and rank them from strongest to weakest in strength
+
class="MsoNormal"><big><span style="">&nbsp; Testing various
-
(in this
+
combinations of the Signal Sequence with either ompA or INPNC would
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specific system).</span></p>
+
help us
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<big><big><big><span style="font-weight: bold;"><small><span
+
find the best combination for our secretion system based on their
-
style="text-decoration: underline; background-color: rgb(255, 255, 0);"></span></small></span></big></big></big><br>
+
secretion ability and allow us to rank the combinations from strongest
 +
to weakest in strength.
 +
(in this specific system).<o:p></o:p></span></big></li>
 +
<li class="MsoNormal"><span style=""><big
 +
style="font-family: Times New Roman,Times,serif;">&nbsp; We have
 +
submitted the BioBricks to the parts registry; therefore they can be
 +
used in future studies (for different purposes).</big><o:p></o:p></span></li>
 +
</ol>
 +
<br>
 +
<div style="text-align: left;"><big><big
 +
style="font-family: Times New Roman,Times,serif;"><big><big><span
 +
style="font-weight: bold;"><small><span
 +
style="text-decoration: underline; background-color: rgb(255, 255, 0);"></span></small></span></big></big></big></big><br>
 +
</div>
<div style="text-align: left;">
<div style="text-align: left;">
<hr style="width: 100%; height: 2px;"><big><small><big
<hr style="width: 100%; height: 2px;"><big><small><big
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</small></span></big><span style="font-style: italic;"><big><span
</small></span></big><span style="font-style: italic;"><big><span
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style="text-decoration: underline;"><small></small></span></big></span></small></big>
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style="text-decoration: underline;"></span></big></span></small></big>
<div style="text-align: center;"><big><span style="font-weight: bold;"></span></big><big><span
<div style="text-align: center;"><big><span style="font-weight: bold;"></span></big><big><span
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style="font-weight: bold;"><a href="UCDAVIS_Model%201.html"><img alt=""
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style="font-weight: bold;"><a
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src="https://static.igem.org/mediawiki/2009/b/ba/UCDAVIS_PIC12.png"
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href="https://2009.igem.org/Team:UC_Davis/Adding_secretion/model_1"><img
 +
alt="" src="https://static.igem.org/mediawiki/2009/b/ba/UCDAVIS_PIC12.png"
style="border: 0px solid ; width: 95px; height: 63px;"></a><a
style="border: 0px solid ; width: 95px; height: 63px;"></a><a
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href="UCDAVIS_Model%202.html"><img alt=""
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href="https://2009.igem.org/Team:UC_Davis/Adding_secretion/model_2"><img
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src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC13.png"
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alt="" src="https://static.igem.org/mediawiki/2009/a/a4/UCDAVIS_PIC13.png"
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style="border: 0px solid ; width: 95px; height: 63px;"></a><a
+
-
href="UCDAVIS_Model%203.html"><img alt=""
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src="https://static.igem.org/mediawiki/2009/9/9a/UCDAVIS_PIC14.png"
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style="border: 0px solid ; width: 95px; height: 63px;"></a><a
+
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href="UCDAVIS_Model%204.html"><img alt=""
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src="https://static.igem.org/mediawiki/2009/b/ba/UCDAVIS_PIC15.png"
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style="border: 0px solid ; width: 95px; height: 63px;"></a><a
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href="UCDAVIS_Model%205.html"><img alt=""
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src="https://static.igem.org/mediawiki/2009/5/5d/UCDAVIS_PIC16.png"
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style="border: 0px solid ; width: 95px; height: 63px;"></a>&nbsp;&nbsp;&nbsp;
style="border: 0px solid ; width: 95px; height: 63px;"></a>&nbsp;&nbsp;&nbsp;
</span><span style="font-style: italic;"><big><span
</span><span style="font-style: italic;"><big><span
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<p class="MsoNormal"><b><span
<p class="MsoNormal"><b><span
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>
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>
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<big><big style="font-weight: bold;"><small></small>
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<big><big style="font-weight: bold;">
</big></big>
</big></big>
<hr style="width: 100%; height: 2px;">
<hr style="width: 100%; height: 2px;">
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</span></big></span></b></p>
</span></big></span></b></p>
<div style="text-align: center;"><b><span
<div style="text-align: center;"><b><span
-
style="font-size: 13.5pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><big><span
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style="font-size: 13.5pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><span
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style="text-decoration: underline;">Molecular data on some proteins</span></big><o:p></o:p></span></b></div>
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style="text-decoration: underline;">Molecular data on some proteins</span><o:p></o:p></span></b></div>
-
<div style="text-align: center;"></div>
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<br>
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<div style="text-align: center;"></div>
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<table
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<table class="MsoNormalTable"
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style="text-align: left; width: 422px; background-color: rgb(0, 0, 153); height: 268px; margin-left: auto; margin-right: auto;"
-
style="border: medium none ; border-collapse: collapse; text-align: left; margin-left: auto; margin-right: auto; width: 597px; height: 264px;"
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border="1" cellpadding="2" cellspacing="2">
-
border="1" cellpadding="0" cellspacing="0">
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<tbody>
<tbody>
-
<tr style="">
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<tr>
<td
<td
-
style="border: 1pt solid black; padding: 0in 5.4pt; width: 125.15pt; background-color: rgb(0, 0, 153); color: rgb(255, 255, 0);"
+
style="vertical-align: top; background-color: rgb(0, 0, 153); text-align: center;"><big><big
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valign="top" width="167">
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style="font-weight: bold;"><span style="color: rgb(255, 255, 0);">Name</span></big></big><br>
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<p class="MsoNormal"
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-
style="margin-bottom: 0.0001pt; line-height: normal;"><b><span
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style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p>&nbsp;</o:p></span></b></p>
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</td>
</td>
 +
<td style="vertical-align: top; text-align: center;"><big
 +
style="color: rgb(255, 255, 0);"><big><span style="font-weight: bold;">Molecular
 +
Weight (kD)</span></big></big><br>
 +
</td>
 +
</tr>
 +
<tr>
<td
<td
-
style="border-style: solid solid solid none; border-color: black black black -moz-use-text-color; border-width: 1pt 1pt 1pt medium; padding: 0in 5.4pt; width: 117.9pt; background-color: rgb(0, 0, 153); color: rgb(255, 255, 0);"
+
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>Myosin<br>
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valign="top" width="157"><small><small> </small></small>
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</big></td>
-
<p class="MsoNormal"
+
-
style="margin-bottom: 0.0001pt; line-height: normal;"><small><small><b><span
+
-
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">Molecular
+
-
Weight<o:p></o:p></span></b></small></small></p>
+
-
<small><small> </small></small></td>
+
<td
<td
-
style="border-style: solid solid solid none; border-color: black black black -moz-use-text-color; border-width: 1pt 1pt 1pt medium; padding: 0in 5.4pt; width: 117.5pt; background-color: rgb(0, 0, 153); color: rgb(255, 255, 0);"
+
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>200.0<br>
-
valign="top" width="157"><small> </small>
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</big></td>
-
<p class="MsoNormal"
+
-
style="margin-bottom: 0.0001pt; line-height: normal;"><small><b><span
+
-
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">Number
+
-
of residues<o:p></o:p></span></b></small></p>
+
-
<small> </small></td>
+
-
<td
+
-
style="border-style: solid solid solid none; border-color: black black black -moz-use-text-color; border-width: 1pt 1pt 1pt medium; padding: 0in 5.4pt; width: 118.25pt; background-color: rgb(0, 0, 153); color: rgb(255, 255, 0);"
+
-
valign="top" width="158">
+
-
<p class="MsoNormal"
+
-
style="margin-bottom: 0.0001pt; line-height: normal;"><b><span
+
-
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">Number
+
-
of polypeptide<o:p></o:p></span></b></p>
+
-
</td>
+
</tr>
</tr>
-
<tr style="">
+
<tr>
<td
<td
-
style="border-style: none solid solid; border-color: -moz-use-text-color black black; border-width: medium 1pt 1pt; padding: 0in 5.4pt; width: 125.15pt; background-color: rgb(0, 0, 153); color: rgb(255, 255, 0);"
+
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>-Galactosidase<br>
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valign="top" width="167">
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</big></td>
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<p class="MsoNormal"
+
-
style="margin-bottom: 0.0001pt; line-height: normal;"><b><span
+
-
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">Cytochrome
+
-
<i>c </i>(human)<o:p></o:p></span></b></p>
+
-
</td>
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<td
<td
-
style="border-style: none solid solid none; border-color: -moz-use-text-color black black -moz-use-text-color; border-width: medium 1pt 1pt medium; padding: 0in 5.4pt; width: 117.9pt; background-color: rgb(0, 0, 153); color: rgb(255, 255, 0);"
+
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>116.3<br>
-
valign="top" width="157"><small> </small>
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</big></td>
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<p class="MsoNormal"
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</tr>
-
style="margin-bottom: 0.0001pt; line-height: normal;"><small><b><span
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<tr>
-
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">13,000<o:p></o:p></span></b></small></p>
+
-
<small> </small></td>
+
<td
<td
-
style="border-style: none solid solid none; border-color: -moz-use-text-color black black -moz-use-text-color; border-width: medium 1pt 1pt medium; padding: 0in 5.4pt; width: 117.5pt; background-color: rgb(0, 0, 153); color: rgb(255, 255, 0);"
+
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold; text-align: left;"><big>Phosphorylase
-
valign="top" width="157">
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b<br>
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<p class="MsoNormal"
+
</big></td>
-
style="margin-bottom: 0.0001pt; line-height: normal;"><b><span
+
-
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">104<o:p></o:p></span></b></p>
+
-
</td>
+
<td
<td
-
style="border-style: none solid solid none; border-color: -moz-use-text-color black black -moz-use-text-color; border-width: medium 1pt 1pt medium; padding: 0in 5.4pt; width: 118.25pt; background-color: rgb(0, 0, 153); color: rgb(255, 255, 0);"
+
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>97.4<br>
-
valign="top" width="158">
+
</big></td>
-
<p class="MsoNormal"
+
-
style="margin-bottom: 0.0001pt; line-height: normal;"><b><span
+
-
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">1<o:p></o:p></span></b></p>
+
-
</td>
+
</tr>
</tr>
-
<tr style="height: 51.25pt;">
+
<tr>
<td
<td
-
style="border-style: none solid solid; border-color: -moz-use-text-color black black; border-width: medium 1pt 1pt; padding: 0in 5.4pt; width: 125.15pt; height: 51.25pt; background-color: rgb(0, 0, 153); color: rgb(255, 255, 0);"
+
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>Ovalbumin<br>
-
valign="top" width="167">
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</big></td>
-
<p class="MsoNormal"
+
-
style="margin-bottom: 0.0001pt; line-height: normal;"><b><span
+
-
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">Ribonuclease
+
-
A(bovine pancreas)<o:p></o:p></span></b></p>
+
-
</td>
+
<td
<td
-
style="border-style: none solid solid none; border-color: -moz-use-text-color black black -moz-use-text-color; border-width: medium 1pt 1pt medium; padding: 0in 5.4pt; width: 117.9pt; height: 51.25pt; background-color: rgb(0, 0, 153); color: rgb(255, 255, 0);"
+
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>45.0<br>
-
valign="top" width="157">
+
</big></td>
-
<p class="MsoNormal"
+
</tr>
-
style="margin-bottom: 0.0001pt; line-height: normal;"><b><span
+
<tr>
-
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">13,700<o:p></o:p></span></b></p>
+
-
</td>
+
<td
<td
-
style="border-style: none solid solid none; border-color: -moz-use-text-color black black -moz-use-text-color; border-width: medium 1pt 1pt medium; padding: 0in 5.4pt; width: 117.5pt; height: 51.25pt; background-color: rgb(0, 0, 153); color: rgb(255, 255, 0);"
+
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>Carbonic
-
valign="top" width="157">
+
anhydrase<br>
-
<p class="MsoNormal"
+
</big></td>
-
style="margin-bottom: 0.0001pt; line-height: normal;"><b><span
+
-
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">124<o:p></o:p></span></b></p>
+
-
</td>
+
<td
<td
-
style="border-style: none solid solid none; border-color: -moz-use-text-color black black -moz-use-text-color; border-width: medium 1pt 1pt medium; padding: 0in 5.4pt; width: 118.25pt; height: 51.25pt; background-color: rgb(0, 0, 153); color: rgb(255, 255, 0);"
+
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>31.0<br>
-
valign="top" width="158">
+
</big></td>
-
<p class="MsoNormal"
+
-
style="margin-bottom: 0.0001pt; line-height: normal;"><b><span
+
-
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">1<o:p></o:p></span></b></p>
+
-
</td>
+
</tr>
</tr>
-
<tr style="">
+
<tr>
<td
<td
-
style="border-style: none solid solid; border-color: -moz-use-text-color black black; border-width: medium 1pt 1pt; padding: 0in 5.4pt; width: 125.15pt; background-color: rgb(0, 0, 153); color: rgb(255, 255, 0);"
+
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>Aprotinin<br>
-
valign="top" width="167">
+
</big></td>
-
<p class="MsoNormal"
+
-
style="margin-bottom: 0.0001pt; line-height: normal;"><b><span
+
-
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">Serum
+
-
albumin (human)<o:p></o:p></span></b></p>
+
-
</td>
+
<td
<td
-
style="border-style: none solid solid none; border-color: -moz-use-text-color black black -moz-use-text-color; border-width: medium 1pt 1pt medium; padding: 0in 5.4pt; width: 117.9pt; background-color: rgb(0, 0, 153); color: rgb(255, 255, 0);"
+
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>6.5<br>
-
valign="top" width="157">
+
</big></td>
-
<p class="MsoNormal"
+
</tr>
-
style="margin-bottom: 0.0001pt; line-height: normal;"><b><span
+
<tr>
-
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">68,500<o:p></o:p></span></b></p>
+
-
</td>
+
<td
<td
-
style="border-style: none solid solid none; border-color: -moz-use-text-color black black -moz-use-text-color; border-width: medium 1pt 1pt medium; padding: 0in 5.4pt; width: 117.5pt; background-color: rgb(0, 0, 153); color: rgb(255, 255, 0);"
+
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>Insulin,
-
valign="top" width="157">
+
B chain, oxidized<br>
-
<p class="MsoNormal"
+
</big></td>
-
style="margin-bottom: 0.0001pt; line-height: normal;"><b><span
+
-
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">609<o:p></o:p></span></b></p>
+
-
</td>
+
<td
<td
-
style="border-style: none solid solid none; border-color: -moz-use-text-color black black -moz-use-text-color; border-width: medium 1pt 1pt medium; padding: 0in 5.4pt; width: 118.25pt; background-color: rgb(0, 0, 153); color: rgb(255, 255, 0);"
+
style="vertical-align: top; color: rgb(255, 255, 0); font-weight: bold;"><big>3.5<br>
-
valign="top" width="158">
+
</big></td>
-
<p class="MsoNormal"
+
-
style="margin-bottom: 0.0001pt; line-height: normal;"><b><span
+
-
style="font-size: 14pt; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">1<o:p></o:p></span></b></p>
+
-
</td>
+
</tr>
</tr>
</tbody>
</tbody>
</table>
</table>
-
<p style="text-align: center;" class="MsoNormal"><b><span
 
-
style="font-size: 13.5pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;">(Page
 
-
128, third edition “Lehninger
 
-
Principles of Biochemistry” by David L. Nelson and Michael M. Cox)</span></b><b><span
 
-
style="font-size: 14pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><o:p>
 
<br>
<br>
-
</o:p></span></b></p>
 
-
<div style="text-align: center;"></div>
 
<p style="text-align: center;" class="MsoNormal"><b><span
<p style="text-align: center;" class="MsoNormal"><b><span
style="font-size: 14pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><br
style="font-size: 14pt; line-height: 115%; font-family: &quot;Times New Roman&quot;,&quot;serif&quot;;"><br

Latest revision as of 02:35, 22 October 2009

Adding Secretion

 

Adding Secretion:
General model for secretion     Secretion Models     Why test different genes

General model for secretion system:


            The purpose of the secretion system is to introduce a method of secreting target proteins we wish to synthesize in the marvelous host: E.coli. Park et. al showed that the truncated form of ice nucleation protein (INPNC) containing the complete coding region of phaZ1, including its signal sequence, could lead to stable secretion of the enzyme (15). We have decided to see if we could co-opt this system to make a more generalized inducible system for protein secretion in which the INPNC and phaZ1 signal sequence serve as carriers for a target protein.


Possible challenges:

1. Efficient recognition of the cleavage site may require some of the non-signal sequence portion of the phaZ1 protein (we have only cloned the signal sequence up to the cleavage site).
2. The system may only work for proteins in a narrow site range.
3. The expression levels of the system may be low.


Advantages:

  1.   In our secretion system, we are using two genes with different sizes as our target secretion genes; GFP being short in length and Luciferase being comparably long, these would test the ability of our secretion system to secrete proteins of different sizes.
  2.   Testing various combinations of the Signal Sequence with either ompA or INPNC would help us find the best combination for our secretion system based on their secretion ability and allow us to rank the combinations from strongest to weakest in strength. (in this specific system).
  3.   We have submitted the BioBricks to the parts registry; therefore they can be used in future studies (for different purposes).



Secretion Models:
Click on a specific model for more information:
   


Why test different genes in our secretion system?

     There is wide range of genes present in our environment. Therefore, it is important to measure our secretion system’s ability to secrete genes of various sizes.

Molecular data on some proteins

Name
Molecular Weight (kD)
Myosin
200.0
-Galactosidase
116.3
Phosphorylase b
97.4
Ovalbumin
45.0
Carbonic anhydrase
31.0
Aprotinin
6.5
Insulin, B chain, oxidized
3.5