Team:XMU-China/Content parts

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<b><h4>Agarose Gel Electrophoresis Confirmation for all Plasmids</h4></b>
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<b><h4>Agarose Gel Electrophoresis Confirmation for all Plasmids</h4></b></br>
<b><p>pSB1C3-gfp-luxI</p></b>
<b><p>pSB1C3-gfp-luxI</p></b>
<p>This plasmid consists of two parts of target genes: gfp and luxI, and both of them are regulated by quorum sensing promoter. High copy number. (Fig.7-1)</p>
<p>This plasmid consists of two parts of target genes: gfp and luxI, and both of them are regulated by quorum sensing promoter. High copy number. (Fig.7-1)</p>
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<b><p>p3H-sfGFP-luxI</p></b>
<b><p>p3H-sfGFP-luxI</p></b>
<p>This plasmid consists of two parts of target genes: sfgfp and luxI, and both of them are regulated by quorum sensing promoter. sfGFP gene is included into this plasmid as a parallel experiment group of pSB1C3-gfp-luxI. Middle copy number. (Fig.7-4)</p>
<p>This plasmid consists of two parts of target genes: sfgfp and luxI, and both of them are regulated by quorum sensing promoter. sfGFP gene is included into this plasmid as a parallel experiment group of pSB1C3-gfp-luxI. Middle copy number. (Fig.7-4)</p>
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<table><tr><td><img src="https://2013.igem.org/File:Fig7_4.png" class="border" alt="" /> </td>
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<table><tr><td><img src="https://static.igem.org/mediawiki/2013/4/46/Fig7_4.png" class="border" alt="" /> </td>
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<tr><td>Fig.7-4  AGE of p3H-sfgfp construction</td></tr></table>
<tr><td>Fig.7-4  AGE of p3H-sfgfp construction</td></tr></table>
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<b><p>pSB4K5-ndh</p></b>
<b><p>pSB4K5-ndh</p></b>
<p>This plasmid has one target gene aiiA under the regulation of quorum sensing promoter. Low copy number. (Fig. 7-6)</p>
<p>This plasmid has one target gene aiiA under the regulation of quorum sensing promoter. Low copy number. (Fig. 7-6)</p>
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<table><tr><td><img src="https://static.igem.org/mediawiki/2013/8/80/Fig7_6.png" class="border" alt="" /> </td>
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<table><tr><td><img src="https://static.igem.org/mediawiki/2013/8/80/Fig7_6.png" width="900" class="border" alt="" /> </td>
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<tr><td>Fig.7-6 AGE confirmation of plasmid pSB4K5-ndh construction
<tr><td>Fig.7-6 AGE confirmation of plasmid pSB4K5-ndh construction
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<h4>SDS-PAGE Confirmation for Plasmids</h4>
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<p>Please refer to <a href="https://2013.igem.org/Team:XMU-China/Exploration"><b>Improvement</b></a> (For a Better Glee) in Strains comparisons.</p>
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<h3><a name="future"></a>Summary & Future work</h3>
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</div>
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</div>
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<div class="center-block-page clearfix">
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<h4>Future Work (Keep Practicing)</h4>
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<p>Oh, no, time is up for iGEM Competition, our glee has no more time to play, but pack their luggage and head for MIT. But let's see what they have made so far and what they still have to do if they really want to be a success. </p>
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<p><b>Construction Have done:</b></p>
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<table width="100%" border="1" cellspacing="0px" align="center">
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  <caption>
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  Table 6-1 Plasmids we have built
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  </caption>
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  <tr>
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    <th rowspan=2>No.</th>
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    <td rowspan=2>Plasmid</td>
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    <td rowspan=2>Replication</br>Origin</td>
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    <td rowspan=2>Copy</br>Number</td>
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    <td rowspan=2>Resistance</td>
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    <td colspan=2 >Size(bp)</td>
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  </tr>
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  <tr>
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    <td>Insert</td>
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    <td>Backbone</td>
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  </tr>
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  <tr>
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  <th >A1</th>
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    <td>pSB1C3-gfp-luxI</td>
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    <td>pSB1C3</td>
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    <td>high</br>(100~300)</td>
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    <td>Chloromycetin</br>(Cm)</td>
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    <td>4052</td>
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    <td>2070)</td>
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    </tr>
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      <tr>
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  <th >A2</th>
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    <td>pSB1C3-sfgfp-luxI</td>
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    <td>pSB1C3</td>
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    <td>high</br>(100~300)</td>
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    <td>Chloromycetin</br>(Cm)</td>
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    <td>4046</td>
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    <td>2070)</td>
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    </tr>
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          <tr>
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  <th >A3</th>
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    <td>p3H-GFP-luxI</td>
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    <td>p3H</td>
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    <td>Middle</br>(18~22)</td>
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    <td>Ampicillin</br>
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(Amp)</td>
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    <td>4052</td>
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    <td>/</td>
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    </tr>
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              <tr>
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  <th >A4</th>
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    <td>p3H-sfGFP-luxI</td>
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    <td>p3H</td>
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    <td>Middle</br>(18~22)</td>
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    <td>Ampicillin</br>
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(Amp)</td>
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    <td>4046</td>
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    <td>/</td>
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    </tr>
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                  <tr>
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  <th >B</th>
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    <td>pSB3T5-aiiA</td>
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    <td>p15A</td>
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    <td>Middle</br>(10~12)</td>
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    <td>Tetracycline</br>
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(Tet)
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</td>
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    <td>2135</td>
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    <td>2837</td>
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    </tr>
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                      <tr>
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  <th >C</th>
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    <td>pSB4K5-ndh</td>
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    <td>pSC101</td>
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    <td>Low</br>(5)</td>
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    <td>Kanamycin</br>
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(Kan)
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</td>
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    <td>2658</td>
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    <td>3004</td>
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    </tr>
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</table>
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<h4>Need to do:</h4>
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<h5 style="font-size:22px">Microfluidic</h5>
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<p><b>Have done:</b>Finished building two different arrays, find basic testing parameters.</p>
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<p style="line-height:0px"><b>To do:</b>Confirm more suitable microfluidic parameters for oscillation.</p>
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<h5 style="font-size:22px">Strain</h5>
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<p><b>Have done:</b>Confirmed DH5a's incompetent in express oscillation;</p>
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<!-- END CONTENT -->
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<p style="line-height:0px"><b>To do:</b>Make a comparison between BL21 (WT) and BL21 (DE3) in SDS-PAGE and so on.</p>
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</br>
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<h5 style="font-size:22px">Copy number</h5>
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<p><b>Have done:</b>Have done: Compared the arrangements of high-middle-low copy number and middle-middle-low copy number's effect on oscillation.</p>
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<p style="line-height:0px"><b>To do:</b>Try other arrangements..</p>
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</br>
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<h5 style="font-size:22px">Fast Degrading Tag</h5>
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<p><b>Have done:</b>Found LVA-tag is not fast enough for our oscillation circuit.</p>
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<p style="line-height:0px"><b>To do:</b>Try LAA or other fast degrading tag for comparison.</p>
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<li><a href="https://2013.igem.org/Team:XMU-China">Home</a></li>
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<li><a href="https://2013.igem.org/Team:XMU-China/Team1">Team</a></li>
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<li><a href="https://2013.igem.org/Team:XMU-China/Project">Project</a></li>
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<li><a href="https://2013.igem.org/Team:XMU-China/Notebook">Notebook</a></li>
<li><a href="https://2013.igem.org/Team:XMU-China/Notebook">Notebook</a></li>
<li><a href="https://2013.igem.org/Team:XMU-China/Software">Software</a></li>
<li><a href="https://2013.igem.org/Team:XMU-China/Software">Software</a></li>
<li><a href="https://2013.igem.org/Team:XMU-China/Human Practice">Human Practice</a></li>
<li><a href="https://2013.igem.org/Team:XMU-China/Human Practice">Human Practice</a></li>
<li><a href="https://2013.igem.org/Team:XMU-China/Safety">Safety</a></li>
<li><a href="https://2013.igem.org/Team:XMU-China/Safety">Safety</a></li>
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<li style="margin-left: 400px"><a href="https://2013.igem.org/Team:XMU-China/Team1#Attribution">Contact us</a></li>
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<div class="copyright">&copy; Copyright &copy; 2013.XMU-China iGEM All rights reserved.</div>
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<div class="copyright" style="margin-right: -150px">&copy; Copyright 2013.XMU-China iGEM All rights reserved.</div>
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Latest revision as of 19:06, 28 October 2013

LinkUp - Multipurpose HTML Template

Parts

Table 1. The Bio-bricks used in the study

Part

Backbone

Type

Location

Size (bp)

Description

part

backbone

BBa_K546000

pSB1C3

Signaling

2013-P1-12D

1964

2070

Lux pL controlled LuxR with lux pR autoinducing LuxI (lva tag)- AHL.

BBa_I763020

pSB1C3

Intermediate

RBS-GFP-TT

2013-P3-11H

914

2070

RBS-GFP (+LVA Tag)-Terminators

BBa_F2621

pSB1A2

Signaling

2013-P2-21F

1158

2079

3OC6HSL Receiver Device

BBa_K546001

pSB1C3

Device

2013-P1-12F

2135

2070

AIIA

AHL Reporter and Quencher

BBa_J04450

pSB4K5

Reporter

2013-P5-5G

1069

3419

RFP Coding Device

BBa_J04450

pSB3T5

Reporter

2013-P5-7C

1069

3241

RFP Coding Device

BBa_J23022

BBa_J23006

Composite

2013-P5-3H

234

2356

[key3d][TT]


Favorite XMU-China 2013 iGEM Team Parts

W/N Name Type Description Designer Length
W BBa_K1036000 Composite lux pL controlled luxR with lux pR controlled ndh (LVA-tag) coding for NADH dehydrogenase II Shengquan Zeng
Zhaopeng Cheng		 2687
W BBa_K1036001 Coding the ndh gene coding for respiratory NADH dehydrogenase II Shengquan Zeng
Zhaopeng Cheng		 1366
W BBa_K1036003 Composite lux pL controlled luxR with lux pR controlled gfp (LVA-tag) Xin Guo
Xi Xi		 4052
W BBa_K1036006 Composite lux pL controlled luxR with lux pR controlled sfgfp (with LVA-tag) Zhaopeng Cheng
Xiyu Wu		 4046


XMU-China 2013 iGEM Team Parts Sandbox

W/N Name Type Description Designer Length
W BBa_K1036002 Composite gfp (with LVA-tag) under plux R control Xin Guo
Xi Xi		 2866
W BBa_K1036004 Reporter sfgfp with LVA-tag Zhaopeng Cheng
Xiyu Wu		 753
W BBa_K1036005 Composite sfgfp (with LVA-tag) under plux R control Zhaopeng Cheng
Xiyu Wu		 2880


Agarose Gel Electrophoresis Confirmation for all Plasmids


pSB1C3-gfp-luxI

This plasmid consists of two parts of target genes: gfp and luxI, and both of them are regulated by quorum sensing promoter. High copy number. (Fig.7-1)

Fig.7-1 AGE confirmation of plasmid pSB1C3-gfp-luxI construction

pSB1C3-sfgfp-luxI

This plasmid consists of two parts of target genes: sfgfp and luxI, and both of them are regulated by quorum sensing promoter. sfGFP gene is included into this plasmid as a parallel experiment group of pSB1C3-gfp-luxI. High copy number. (Fig.7-2)

Fig.7-2 AGE confirmation of plasmid pSB1C3-sfgfp-luxI construction

p3H-GFP-luxI

This plasmid consists of two parts of target genes: gfp and luxI, and both of them are regulated by quorum sensing promoter. Middle copy number. (Fig.7-3)

Fig.7-3 AGE of p3H-gfp construction

p3H-sfGFP-luxI

This plasmid consists of two parts of target genes: sfgfp and luxI, and both of them are regulated by quorum sensing promoter. sfGFP gene is included into this plasmid as a parallel experiment group of pSB1C3-gfp-luxI. Middle copy number. (Fig.7-4)

Fig.7-4 AGE of p3H-sfgfp construction

pSB3T5-aiiA

This plasmid has one target gene aiiA under the regulation of quorum sensing promoter. Middle copy number. (Fig. 7-5)

Fig.7-5 AGE confirmation of plasmid pSB3T5-aiiA construction

pSB4K5-ndh

This plasmid has one target gene aiiA under the regulation of quorum sensing promoter. Low copy number. (Fig. 7-6)

Fig.7-6 AGE confirmation of plasmid pSB4K5-ndh construction

SDS-PAGE Confirmation for Plasmids

Please refer to Improvement (For a Better Glee) in Strains comparisons.

Summary & Future work

Future Work (Keep Practicing)

Oh, no, time is up for iGEM Competition, our glee has no more time to play, but pack their luggage and head for MIT. But let's see what they have made so far and what they still have to do if they really want to be a success.

Construction Have done:

Table 6-1 Plasmids we have built
No. Plasmid Replication
Origin		 Copy
Number		 Resistance Size(bp)
Insert Backbone
A1 pSB1C3-gfp-luxI pSB1C3 high
(100~300)		 Chloromycetin
(Cm)		 4052 2070)
A2 pSB1C3-sfgfp-luxI pSB1C3 high
(100~300)		 Chloromycetin
(Cm)		 4046 2070)
A3 p3H-GFP-luxI p3H Middle
(18~22)		 Ampicillin
(Amp)		 4052 /
A4 p3H-sfGFP-luxI p3H Middle
(18~22)		 Ampicillin
(Amp)		 4046 /
B pSB3T5-aiiA p15A Middle
(10~12)		 Tetracycline
(Tet) 		2135 2837
C pSB4K5-ndh pSC101 Low
(5)		 Kanamycin
(Kan) 		2658 3004

Need to do:

Microfluidic

Have done:Finished building two different arrays, find basic testing parameters.

To do:Confirm more suitable microfluidic parameters for oscillation.


Strain

Have done:Confirmed DH5a's incompetent in express oscillation;

To do:Make a comparison between BL21 (WT) and BL21 (DE3) in SDS-PAGE and so on.


Copy number

Have done:Have done: Compared the arrangements of high-middle-low copy number and middle-middle-low copy number's effect on oscillation.

To do:Try other arrangements..


Fast Degrading Tag

Have done:Found LVA-tag is not fast enough for our oscillation circuit.

To do:Try LAA or other fast degrading tag for comparison.

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