Team:ZJU-China/Notebook/September

From 2011.igem.org

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                 <div id="accordion">  
                 <div id="accordion">  
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  <h4>May&June</h4>
 +
<div class="pane" style="height:140px;"><a href="https://2011.igem.org/Team:ZJU-China/Notebook/Brainstorm">In the two months we discussed some new ideas and finally decided our project<br/>>> Click to see our brainstorm</a></div>
<h4 >July</h4>
<h4 >July</h4>
 
 
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</div>
</div>
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<h4>Protocol</h4>
 +
        <div class="pane"><a href="https://2011.igem.org/Team:ZJU-China/Protocol">>>Click to see our lab protocol about biofilm formation</a></div>
  </div>
  </div>
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</div>
</div>
            
            
-
   <div class="bigblock">
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   <div class="bigblock"><div class="inner" id="n_biobrick">
-
<div class="block" id="nsheet">  
+
<div class="block" id="nsheet">
-
<h3>Week1</h3><hr/>  
+
<h3>Week9</h3><hr/>
-
+
 
-
<table id="notesheet" width="650" border="1" cellspacing="0" cellpadding="1">  
+
<table id="notesheet" width="650" border="1" cellspacing="0" cellpadding="1">
-
   <tr><td width="76">Day</td><td width="349">Note</td></tr>  
+
   <tr><td width="76">Day</td><td width="349">Note</td></tr>
    
    
-
   <tr>  
+
   <tr>
-
     <td id="sheetleft">Jul.4th Monday</td>  
+
     <td id="sheetleft">Aug.29th
-
     <td><table id="intable" width="328" border="0" cellspacing="0" cellpadding="1">  
+
Monday
 +
 
 +
 
 +
 
 +
 
 +
</td>
 +
     <td><table id="intable" width="603" border="0" cellspacing="0" cellpadding="1">
    
    
-
     <td width="128">• Aerobic cultivation of DH5α</td>  
+
     <td width="191">• Meeting. Summarize all the experiment work so far.
 +
</td>
    
    
    
    
-
     <td width="196">• Preparation of apparatus for the formation of biofilm</td>  
+
     <td width="249" >• Cut: 22M(X+P), 1C3(E+P)
 +
</td>
 +
 
    
    
-
</table></td>  
+
</table></td>
-
   </tr>  
+
   </tr>
-
   <tr>  
+
   <tr>
-
     <td id="sheetleft">Jul 5th Tuesday
+
     <td id="sheetleft">Aug.30th
-
</td>  
+
Tuesday
-
     <td>&nbsp;</td>  
+
 
-
   </tr>  
+
 
-
   <tr>  
+
 
-
     <td id="sheetleft">Jul.6th
+
 
 +
 
 +
</td>
 +
     <td><table id="intable" width="615" border="0" cellspacing="0" cellpadding="1">
 +
 
 +
 
 +
 
 +
</table></td>
 +
   </tr>
 +
   <tr>
 +
     <td id="sheetleft">Aug.31st
Wednesday
Wednesday
-
</td>  
+
 
-
     <td><table width="217" border="0" cellspacing="0" cellpadding="1">  
+
 
-
   <tr>  
+
 
-
    <td width="215">•Receiving  primers ordered previously</td>  
+
 
-
   
+
 
-
   </tr>  
+
</td>
-
</table>  
+
     <td><table width="640" border="0" cellspacing="0" cellpadding="1">
-
</td>  
+
   <tr>
-
   </tr>  
+
  <td>• Cut vgb+YFP+tetR, fdhF+RFP+tetR.<br/>• Run the digestion results</td>
-
   <tr>  
+
  <td>• Culture the positive results</td>
-
     <td id="sheetleft">Jul.7th
+
   </tr>
 +
</table>
 +
</td>
 +
   </tr>
 +
   <tr>
 +
     <td id="sheetleft">Sept.1st
Thursday
Thursday
-
</td>  
+
 
-
     <td><table width="238" border="0" cellspacing="0" cellpadding="1">  
+
 
-
   <tr>  
+
 
-
    <td width="200">• Preparation of the aliquot of the primers</td>  
+
 
-
    <td width="200">• Something wrong with a shaking incubator</td>  
+
 
-
   </tr>  
+
</td>
-
</table>  
+
     <td><table width="618" border="0" cellspacing="0" cellpadding="1">
-
</td>  
+
   <tr>
-
   </tr>  
+
 
-
   <tr>  
+
<td>• Miniprep: Y#4, R#2/3/4/5<br/>
-
     <td id="sheetleft">Jul.8th
+
• Cut Y#4, R#2/3/4/5
 +
</td>
 +
<td>•Run the gel.<br/>
 +
• Send R#3,Y#4 sample to sequencing
 +
</td>
 +
<td>• Hypoxia culture: RFP3, RFP4<br/>
 +
• Preserve Y#4, R#2/3/4/5
 +
</td>
 +
   </tr>
 +
</table>
 +
</td>
 +
   </tr>
 +
   <tr>
 +
     <td id="sheetleft">Sept.2nd
Friday
Friday
-
</td>  
+
 
-
     <td><table width="222" border="0" cellspacing="0" cellpadding="1">  
+
 
-
       <tr>  
+
 
-
        <td width="155">• Preparation of culture plates for the transformations</td>  
+
</td>
-
       </tr>  
+
     <td><table width="627" border="0" cellspacing="0" cellpadding="1">
-
     </table></td>  
+
       <tr>
-
   </tr>  
+
      <td >• Phusion PCR</td>
-
   <tr>  
+
      <td>• Hypoxia Culture: R, Y<br/>
-
     <td id="sheetleft">Jul.9th
+
• Run the PCR results. Bands are confirmed right.
 +
</td>
 +
<td>• Purification: vgb, YFP, tetR.</td>
 +
       </tr>
 +
     </table></td>
 +
   </tr>
 +
   <tr>
 +
     <td id="sheetleft">Sept.3rd
Saturday
Saturday
-
</td>  
+
 
-
     <td><table width="313" border="0" cellspacing="0" cellpadding="1">  
+
 
-
   <tr>  
+
 
-
     <td width="127">• Preparation of culture plates for the transformations</td>  
+
</td>
-
    <td width="182">• protocols of transformation</td>  
+
     <td><table width="620" border="0" cellspacing="0" cellpadding="1">
-
   </tr>  
+
   <tr>
-
</table>  
+
     <td>• Culture: YFP, RFP, A25<br/>
-
</td>  
+
• Culture in micro-oxygen: YFP, RFP, A25
-
   </tr>  
+
</td>
-
   <tr>  
+
  <td>• Culture in slope medium: RFP, YFP, A25<br/>
-
     <td id="sheetleft">Jul.10th
+
• Hypoxia culture: RFP, YFP, A25
 +
</td>
 +
<td>• Check the fluorescence.<br/>
 +
• Neither RFP in hypoxia or oxygen condition.
 +
Little YFP both in hypoxia and oxygen.
 +
</td>
 +
   </tr>
 +
</table>
 +
</td>
 +
   </tr>
 +
   <tr>
 +
     <td id="sheetleft">Sept.4th
Sunday
Sunday
-
</td>  
+
 
-
     <td><table width="246" border="0" cellspacing="0" cellpadding="1">  
+
 
-
       <tr>  
+
</td>
-
         <td>• Several colonies were picked up and cultivated in  5mL LB medium.
+
     <td><table width="620" border="0" cellspacing="0" cellpadding="1">
-
•Cryosectioning of biofilm
+
       <tr>
-
</td>  
+
         <td width="244">• Miniprep: RFP3, YFP4
-
       </tr>  
+
 
-
     </table></td>  
+
</td>
-
   </tr>  
+
<td width="164">• Cut: RFP3, YFP4, !C3.
-
+
</td>
-
</table>  
+
 
 +
       </tr>
 +
     </table></td>
 +
   </tr>
 +
 
 +
</table>
<p>&nbsp;</p>  
<p>&nbsp;</p>  
</div>  
</div>  
-
<script type="text/javascript">
 
-
</script>
+
<div class="block" id="nsheet">
 +
<h3>Week10</h3><hr/>
 +
<table id="notesheet" width="650" border="1" cellspacing="0" cellpadding="1">
 +
  <tr><td width="76">Day</td><td width="349">Note</td></tr>
 +
 
 +
  <tr>
 +
    <td id="sheetleft">Sept.5th
 +
Monday
 +
</td>
 +
    <td><table id="intable" width="603" border="0" cellspacing="0" cellpadding="1">
    
    
 +
    <td width="191">• Run the gel: Yu’s PCR results, 1C3, RFP3, YFP4<br/>
 +
• Purification: RFP3, YFP4
 +
 +
</td>
 +
 
 +
 
 +
    <td  >• Ligation: RFP3+1C3, YFP4+1C3<br/>
 +
• Hypoxia culture: RFP3, YFP4, A25.
 +
 +
</td>
 +
<td>• Transform the ligation results.<br/>
 +
• Confirmed RFP expression only in hypoxia condition.
 +
</td>
 +
 
 +
</table></td>
 +
  </tr>
 +
  <tr>
 +
    <td id="sheetleft">Sept.6th
 +
Tuesday
 +
 +
 +
 +
 +
 +
 +
</td>
 +
    <td><table id="intable" width="615" border="0" cellspacing="0" cellpadding="1">
 +
  <td>• Check the RFP, YFP plates.</td>
 +
  <td>• Colony PCR</td>
 +
 +
</table></td>
 +
  </tr>
 +
  <tr>
 +
    <td id="sheetleft">Sept.7th
 +
Wednesday
 +
 +
 +
 +
 +
 +
 +
</td>
 +
    <td><table width="640" border="0" cellspacing="0" cellpadding="1">
 +
  <tr>
 +
  <td>• Culture YFP1/2, RFP3/4</td>
 +
 
 +
  </tr>
 +
</table>
 +
</td>
 +
  </tr>
 +
  <tr>
 +
    <td id="sheetleft">Sept.8th
 +
Thurday
 +
 +
 +
</td>
 +
    <td><table width="618" border="0" cellspacing="0" cellpadding="1">
 +
  <tr>
 +
 
 +
<td>• Preserve the strain of RFP1, YFP1/2 in 1C3.<br/>
 +
• Miniprep the remains.
 +
 +
</td>
 +
<td>• Cut: YFP1/2, RFP1
 +
</td>
 +
<td>• Run the digestion results.<br/>
 +
• Culture RFP2
 +
 +
</td>
 +
  </tr>
 +
</table>
 +
</td>
 +
  </tr>
 +
  <tr>
 +
    <td id="sheetleft">Sept.9th
 +
Friday
 +
 +
 +
 +
 +
</td>
 +
    <td><table width="627" border="0" cellspacing="0" cellpadding="1">
 +
      <tr>
 +
      <td >• Miniprep: RFP2, YFP2, RFP1</td>
 +
      <td>• Cut them and run the gel.
 +
</td>
 +
 +
      </tr>
 +
    </table></td>
 +
  </tr>
 +
  <tr>
 +
    <td id="sheetleft">Sept.10th
 +
Saturday
 +
 +
 +
 +
 +
</td>
 +
    <td><table width="620" border="0" cellspacing="0" cellpadding="1">
 +
  <tr>
 +
    <td>• Cut YFP(1A3), RFP(1A3) with S+E
 +
 +
 +
</td>
 +
  <td>• Purification: RFP(1A3), YFP(1A3)
 +
</td>
 +
<td>• Ligation: RFP+1C3, YFP+1C3
 +
</td>
 +
<td>• Transform the ligation results.</td>
 +
  </tr>
 +
</table>
 +
</td>
 +
  </tr>
 +
  <tr>
 +
    <td id="sheetleft">Sept.11th
 +
Sunday
 +
 +
 +
 +
</td>
 +
    <td><table width="620" border="0" cellspacing="0" cellpadding="1">
 +
      <tr>
 +
        <td width="244">• Check the plates.
 +
 +
</td>
 +
<td width="164">• Culture lig RFY+YFP+IC3<br/>
 +
• Culture 1K3
 +
 +
</td>
 +
<td>• Colony PCR: RFP(1C3)， YFP(1C3)</td>
 +
      </tr>
 +
    </table></td>
 +
  </tr>
 +
 +
</table>
 +
<p>&nbsp;</p>
 +
</div>
 +
 +
<div class="block" id="nsheet">
 +
<h3>Week11</h3><hr/>
 +
 +
<table id="notesheet" width="650" border="1" cellspacing="0" cellpadding="1">
 +
  <tr><td width="76">Day</td><td width="349">Note</td></tr>
 +
 
 +
  <tr>
 +
    <td id="sheetleft">Sept.12th
 +
Monday
 +
 +
 +
</td>
 +
    <td><table id="intable" width="603" border="0" cellspacing="0" cellpadding="1">
 +
 
 +
    <td width="191">• Colony PCR: 1A3, 1K3, lig RFP+YFP+1C3.
 +
 +
</td>
 +
 
 +
 
 +
    <td  >• Run the PCR results.
 +
 +
</td>
 +
<td>• Miniprep: lig RFP+YFP+1C3
 +
</td>
 +
 
 +
</table></td>
 +
  </tr>
 +
  <tr>
 +
    <td id="sheetleft">Sept.13th
 +
Tuesday
 +
 +
 +
 +
 +
 +
 +
 +
</td>
 +
    <td><table id="intable" width="615" border="0" cellspacing="0" cellpadding="1">
 +
  <td>• Transform 2M, 2I, 6I, 23L.</td>
 +
  <td>• Purification: 1K3</td>
 +
 +
</table></td>
 +
  </tr>
 +
  <tr>
 +
    <td id="sheetleft">Sept.14th
 +
Wednesday
 +
 +
 +
 +
 +
 +
 +
 +
</td>
 +
    <td><table width="640" border="0" cellspacing="0" cellpadding="1">
 +
  <tr>
 +
  <td>• Miniprep: 20H, 20J, 22B</td>
 +
  <td>• Cut: 1K3 with E+P, lig3 with S+E, 12I(CFP) with P+X,</td>
 +
  <td>• Gel excision and /or purification: 1K3, lig3, 12I(CFP)</td>
 +
  <td>• Run the gel.<br/>
 +
• Culture: 1I, 1K, 3L, 5E, 7C
 +
</td>
 +
<td>• Miniprep: RFP+YFP+1C3.<br/>
 +
• Run the gel of 1K3, lig3, CFP again.
 +
</td>
 +
<td>• Gel excision and purification<br/>
 +
• Ligation: Lig3+CFP+1K3
 +
</td>
 +
  </tr>
 +
</table>
 +
</td>
 +
  </tr>
 +
  <tr>
 +
    <td id="sheetleft">Sept.15th
 +
Thursday
 +
 +
 +
 +
</td>
 +
    <td><table width="618" border="0" cellspacing="0" cellpadding="1">
 +
  <tr>
 +
 
 +
<td>• Preserve: 1I, 1K, 3C, 5E, 7C<br/>
 +
• Transform the ligation results
 +
</td>
 +
<td>• Colony PCR: 2M, 2I，6I，23L<br/>
 +
• Cut: 20H, 20J, 22B with X+P
 +
 +
</td>
 +
<td>• Miniprep: 2M-1, 2I-1, 6I-1, 23L-1<br/>
 +
• Cut: 2M, 2I with E+S
 +
 +
 +
</td>
 +
  </tr>
 +
</table>
 +
</td>
 +
  </tr>
 +
  <tr>
 +
    <td id="sheetleft">Sept.16th
 +
Friday
 +
 +
 +
 +
 +
</td>
 +
    <td><table width="627" border="0" cellspacing="0" cellpadding="1">
 +
      <tr>
 +
      <td >• Colony PCR: RFP+YFP+1K3</td>
 +
      <td>• Cut 13K with E+S, 10I with P+X, 1C3 with E+P
 +
</td>
 +
<td>• Run the gel: 13K, 10I, 1C3, 2I, 2M, 20J, 22H, 22B</td>
 +
<td>• Ligation: 13K+10I+1C3<br/>
 +
• Culture: YFP(Amp), CFP(Amp)
 +
</td>
 +
      </tr>
 +
    </table></td>
 +
  </tr>
 +
  <tr>
 +
    <td id="sheetleft">Sept.17th
 +
Saturday
 +
 +
 +
 +
 +
 +
</td>
 +
    <td><table width="620" border="0" cellspacing="0" cellpadding="1">
 +
  <tr>
 +
    <td>• Order primers for 20H, 20J<br/>
 +
• Culture 20H-1, 20J-1, 22B-1
 +
</td>
 +
  <td>• Plate the ligation results<br/>
 +
• Culture RFP+YFP+CFP+1K3
 +
 +
</td>
 +
<td>• Colony PCR: 3A, 1C3<br/>
 +
• Preserve: 1K3, RFP+YFP+1C3.
 +
 +
</td>
 +
  </tr>
 +
</table>
 +
</td>
 +
  </tr>
 +
  <tr>
 +
    <td id="sheetleft">Sept.18th
 +
Sunday
 +
 +
 +
</td>
 +
    <td><table width="620" border="0" cellspacing="0" cellpadding="1">
 +
      <tr>
 +
        <td width="244">• Miniprep: 20J, 20H, RFP+YFP+CFP+1K3
 +
1A3
 +
</td>
 +
<td width="164">• Run the gel of PCR results
 +
 +
</td>
 +
<td>• Cut RFP+YFP+CFP+1K3
 +
with E+P<br/>
 +
• Colony PCR: 13K+10I+1C3
 +
</td>
 +
<td>• PCR: fdhF+rush+Vgb+tetR<br/>
 +
• Preserve: 20H, 20J, 22B
 +
</td>
 +
<td>• Run the gel: 20H, 20J, 22B<br/>
 +
• Cut the bands and purification.
 +
</td>
 +
<td>• Run the gel: 13K+10I+1C3, RFP+YFP+CFP+1K3<br/>
 +
• Culture: 13K+10I+1C3<br/>
 +
• Grad PCR: fdhF.
 +
</td>
 +
      </tr>
 +
    </table></td>
 +
  </tr>
 +
 +
</table>
 +
<p>&nbsp;</p>
 +
</div>
 +
 +
<div class="block" id="nsheet">
 +
<h3>Week12</h3><hr/>
 +
 +
<table id="notesheet" width="650" border="1" cellspacing="0" cellpadding="1">
 +
  <tr><td width="76">Day</td><td width="349">Note</td></tr>
 +
 
 +
  <tr>
 +
    <td id="sheetleft">Sept.19th
 +
Monday
 +
 +
 +
 +
</td>
 +
    <td><table id="intable" width="603" border="0" cellspacing="0" cellpadding="1">
 +
 
 +
    <td width="191">• Miniprep: 13K+10I+1C3, 1C3<br/>
 +
• Run the gel of digestion results.
 +
 +
</td>
 +
 
 +
 
 +
    <td  >• Cut: 13K+10I+1C3, 1A3, fdhF
 +
 +
</td>
 +
<td>• Run the gel: 20H, 20J, rush+vgb, RYC+1K3, fdhF, 13K+10I, 1A3
 +
</td>
 +
  <td>• Cut and purification: fdhF, 13K+10I, 1A3<br/>
 +
• Culture: CFP, YFP
 +
</td>
 +
<td>• Purification: Rush PCR<br/>
 +
• Cut: 22B with X, rush PCR results with S
 +
</td>
 +
 +
</table></td>
 +
  </tr>
 +
  <tr>
 +
    <td id="sheetleft">Sept.20th
 +
Tuesday
 +
 +
 +
 +
 +
 +
 +
 +
 +
</td>
 +
    <td><table id="intable" width="615" border="0" cellspacing="0" cellpadding="1">
 +
  <td>• Run the 20H, 20J from PCR results.<br/>
 +
• Transform: fdhF+13K+10I+1A3
 +
</td>
 +
  <td>• Purification: 20B, Rush<br/>
 +
• Ligation: 22B+Rush
 +
</td>
 +
<td>• Purification: ligation results<br/>
 +
• Cut: the ligation results with X+S
 +
</td>
 +
<td>• PCR 20H<br/>
 +
• Self-ligation: rush+22M
 +
</td>
 +
</table></td>
 +
  </tr>
 +
  <tr>
 +
    <td id="sheetleft">Sept.21st
 +
Wednesday
 +
 +
 +
 +
 +
 +
 +
 +
 +
</td>
 +
    <td><table width="640" border="0" cellspacing="0" cellpadding="1">
 +
  <tr>
 +
  <td>• Colony PCR: fdhF+13K+10I+1A3 on plates</td>
 +
  <td>• Run the gel of PCR results</td>
 +
  <td>• Culture the positive results.</td>
 +
 +
  </tr>
 +
</table>
 +
</td>
 +
  </tr>
 +
  <tr>
 +
    <td id="sheetleft">Sept.22nd
 +
Thursday
 +
 +
 +
 +
</td>
 +
    <td><table width="618" border="0" cellspacing="0" cellpadding="1">
 +
  <tr>
 +
 
 +
<td>• Miniprep: fdhF+13K+10I+1A3<br/>
 +
• Preserve the above.
 +
 +
</td>
 +
<td>• Cut and run the results:
 +
fdhF+13K+10I+1A3, 13K+10I+1C3
 +
 +
 +
</td>
 +
<td>• Cut: RFP with E+S, YFP with X+P, 1K3 with E+P, 1C3 with E+P
 +
</td>
 +
<td>• Purification the digestion results.</td>
 +
  </tr>
 +
</table>
 +
</td>
 +
  </tr>
 +
  <tr>
 +
    <td id="sheetleft">Sept.23rd
 +
Friday
 +
 +
 +
 +
 +
 +
</td>
 +
    <td><table width="627" border="0" cellspacing="0" cellpadding="1">
 +
      <tr>
 +
      <td >• Ligate: RFP+1A3/ YFP+1C3/1K3, RFP+1A3/1C3</td>
 +
      <td>• Transform the ligation results.
 +
</td>
 +
 +
      </tr>
 +
    </table></td>
 +
  </tr>
 +
  <tr>
 +
    <td id="sheetleft">Sept. 24th
 +
Saturday
 +
 +
 +
 +
 +
 +
 +
</td>
 +
    <td><table width="620" border="0" cellspacing="0" cellpadding="1">
 +
  <tr>
 +
    <td>• Order primers for 20H, 20J<br/>
 +
• Culture 20H-1, 20J-1, 22B-1
 +
</td>
 +
  <td>• Plate the ligation results<br/>
 +
• Culture RFP+YFP+CFP+1K3
 +
 +
</td>
 +
<td>• Colony PCR: 3A, 1C3<br/>
 +
• Preserve: 1K3, RFP+YFP+1C3.
 +
 +
</td>
 +
  </tr>
 +
</table>
 +
</td>
 +
  </tr>
 +
  <tr>
 +
    <td id="sheetleft">Sept.25th
 +
Sunday
 +
 +
</td>
 +
    <td><table width="620" border="0" cellspacing="0" cellpadding="1">
 +
      <tr>
 +
        <td width="244">• Miniprep: 20J, 20H, RFP+YFP+CFP+1K3
 +
1A3
 +
</td>
 +
<td width="164">• Run the gel of PCR results
 +
 +
</td>
 +
<td>• Cut RFP+YFP+CFP+1K3
 +
with E+P<br/>
 +
• Colony PCR: 13K+10I+1C3
 +
</td>
 +
<td>• PCR: fdhF+rush+Vgb+tetR<br/>
 +
• Preserve: 20H, 20J, 22B
 +
</td>
 +
<td>• Run the gel: 20H, 20J, 22B<br/>
 +
• Cut the bands and purification.
 +
</td>
 +
<td>• Run the gel: 13K+10I+1C3, RFP+YFP+CFP+1K3<br/>
 +
• Culture: 13K+10I+1C3<br/>
 +
• Grad PCR: fdhF.
 +
</td>
 +
      </tr>
 +
    </table></td>
 +
  </tr>
 +
 +
</table>
 +
<p>&nbsp;</p>
 +
</div>
 +
</div>
 +
 +
 +
 +
 +
<div class="inner" id="biofilm_jul">
 +
<div class="block" id="nsheet">
 +
<h3>3rd Sep</h3><hr/>
 +
 +
 +
<p>Directly use DH5α fdhF+RFP colony to inoculate silicone tube set with 5ml LB. rest for
 +
12h for biofilm attachment.</p>
 +
</div>
 +
<div class="block" id="nsheet">
 +
<h3>4th Sep</h3><hr/>
 +
<p>Add 50mlLB+Amp and start circulation.</p>
 +
</div>
 +
<div class="block" id="nsheet">
 +
<h3>6th Sep</h3><hr/>
 +
<p>Biofilm with direct inoculation is optimum. Expressed red florescence but only one small
 +
fraction of the biofilm remained after cutting off the part of the tube.</p>
 +
</div>
 +
<div class="block" id="nsheet">
 +
<h3>12th Sep</h3><hr/>
 +
<p>4 silicone tube sets with biofilm attachement time of 3h, 5h, 7.5h and 10h; 4 bubbling sets.</p>
 +
</div>
 +
<div class="block" id="nsheet">
 +
<h3>15th Sep</h3><hr/>
 +
<p>The silicone tube set with 3h of attachment achieves optimum biofilm formation. Bubbling sets
 +
glass slides are froze in -80℃ for later observation with laser confocal. Biofilm formed on glass
 +
slide is about 10μm thick.</p>
 +
</div>
 +
<div class="block" id="nsheet">
 +
<h3>16th Sep</h3><hr/>
 +
<p>New sets with silicone tube, a mixture of RFP, CFP and YFP expressing e.coli. Attachment 3h.</p>
 +
</div>
 +
<div class="block" id="nsheet">
 +
<h3>17th Sep</h3><hr/>
 +
<p>Glass slide with bubbling set with the same mixture of e.coli.<br/>
 +
Place two filter film on solid culture medium and placed the mixture of e.coli on the filter to form
 +
gas-solid interface biofilm.</p>
 +
</div>
 +
<div class="block" id="nsheet">
 +
<h3>20th Sep</h3><hr/>
 +
<p>Observation of glass slide and gas-solid biofilm under laser confocal. Glass slide are observed
 +
directly and gas-solid biofilm is first pressed by a covering slide so that it would stick to it and
 +
peeled off from the filter for observation.<br/>
 +
Biofilm on glass slide is about 8.6μm thick. Biofilm formed on gas-solid interface is too
 +
concentrated in bacteria and culture medium that the florescence of too strong to see any clear
 +
structure.</p>
 +
 +
 +
<p>&nbsp;</p>
 +
</div>
 +
</div>
 +
  <script type="text/javascript">
 +
document.getElementById('p_intro').onclick= function(){
 +
document.getElementById('biofilm_jul').style.display='none'
 +
document.getElementById('n_biobrick').style.display='block'}
 +
document.getElementById('p_model').onclick= function(){
 +
document.getElementById('n_biobrick').style.display='none'
 +
document.getElementById('biofilm_jul').style.display='block'}
 +
</script>
 +
</div>
</div>

Latest revision as of 07:00, 3 October 2011

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Labnote

 Biobrick group | Biofilm group 

This group...........

Week9

DayNote
Aug.29th Monday
• Meeting. Summarize all the experiment work so far. • Cut: 22M(X+P), 1C3(E+P)
Aug.30th Tuesday
Aug.31st Wednesday
• Cut vgb+YFP+tetR, fdhF+RFP+tetR.
• Run the digestion results		 • Culture the positive results
Sept.1st Thursday
• Miniprep: Y#4, R#2/3/4/5
• Cut Y#4, R#2/3/4/5 		•Run the gel.
• Send R#3,Y#4 sample to sequencing 		• Hypoxia culture: RFP3, RFP4
• Preserve Y#4, R#2/3/4/5
Sept.2nd Friday
• Phusion PCR • Hypoxia Culture: R, Y
• Run the PCR results. Bands are confirmed right. 		• Purification: vgb, YFP, tetR.
Sept.3rd Saturday
• Culture: YFP, RFP, A25
• Culture in micro-oxygen: YFP, RFP, A25 		• Culture in slope medium: RFP, YFP, A25
• Hypoxia culture: RFP, YFP, A25 		• Check the fluorescence.
• Neither RFP in hypoxia or oxygen condition. Little YFP both in hypoxia and oxygen.
Sept.4th Sunday
• Miniprep: RFP3, YFP4 • Cut: RFP3, YFP4, !C3.

 

Week10

DayNote
Sept.5th Monday
• Run the gel: Yu’s PCR results, 1C3, RFP3, YFP4
• Purification: RFP3, YFP4 		• Ligation: RFP3+1C3, YFP4+1C3
• Hypoxia culture: RFP3, YFP4, A25. 		• Transform the ligation results.
• Confirmed RFP expression only in hypoxia condition.
Sept.6th Tuesday
• Check the RFP, YFP plates. • Colony PCR
Sept.7th Wednesday
• Culture YFP1/2, RFP3/4
Sept.8th Thurday
• Preserve the strain of RFP1, YFP1/2 in 1C3.
• Miniprep the remains. 		• Cut: YFP1/2, RFP1 • Run the digestion results.
• Culture RFP2
Sept.9th Friday
• Miniprep: RFP2, YFP2, RFP1 • Cut them and run the gel.
Sept.10th Saturday
• Cut YFP(1A3), RFP(1A3) with S+E • Purification: RFP(1A3), YFP(1A3) • Ligation: RFP+1C3, YFP+1C3 • Transform the ligation results.
Sept.11th Sunday
• Check the plates. • Culture lig RFY+YFP+IC3
• Culture 1K3 		• Colony PCR: RFP(1C3)， YFP(1C3)

 

Week11

DayNote
Sept.12th Monday
• Colony PCR: 1A3, 1K3, lig RFP+YFP+1C3. • Run the PCR results. • Miniprep: lig RFP+YFP+1C3
Sept.13th Tuesday
• Transform 2M, 2I, 6I, 23L. • Purification: 1K3
Sept.14th Wednesday
• Miniprep: 20H, 20J, 22B • Cut: 1K3 with E+P, lig3 with S+E, 12I(CFP) with P+X, • Gel excision and /or purification: 1K3, lig3, 12I(CFP) • Run the gel.
• Culture: 1I, 1K, 3L, 5E, 7C 		• Miniprep: RFP+YFP+1C3.
• Run the gel of 1K3, lig3, CFP again. 		• Gel excision and purification
• Ligation: Lig3+CFP+1K3
Sept.15th Thursday
• Preserve: 1I, 1K, 3C, 5E, 7C
• Transform the ligation results 		• Colony PCR: 2M, 2I，6I，23L
• Cut: 20H, 20J, 22B with X+P 		• Miniprep: 2M-1, 2I-1, 6I-1, 23L-1
• Cut: 2M, 2I with E+S
Sept.16th Friday
• Colony PCR: RFP+YFP+1K3 • Cut 13K with E+S, 10I with P+X, 1C3 with E+P • Run the gel: 13K, 10I, 1C3, 2I, 2M, 20J, 22H, 22B • Ligation: 13K+10I+1C3
• Culture: YFP(Amp), CFP(Amp)
Sept.17th Saturday
• Order primers for 20H, 20J
• Culture 20H-1, 20J-1, 22B-1 		• Plate the ligation results
• Culture RFP+YFP+CFP+1K3 		• Colony PCR: 3A, 1C3
• Preserve: 1K3, RFP+YFP+1C3.
Sept.18th Sunday
• Miniprep: 20J, 20H, RFP+YFP+CFP+1K3 1A3 • Run the gel of PCR results • Cut RFP+YFP+CFP+1K3 with E+P
• Colony PCR: 13K+10I+1C3 		• PCR: fdhF+rush+Vgb+tetR
• Preserve: 20H, 20J, 22B 		• Run the gel: 20H, 20J, 22B
• Cut the bands and purification. 		• Run the gel: 13K+10I+1C3, RFP+YFP+CFP+1K3
• Culture: 13K+10I+1C3
• Grad PCR: fdhF.

 

Week12

DayNote
Sept.19th Monday
• Miniprep: 13K+10I+1C3, 1C3
• Run the gel of digestion results. 		• Cut: 13K+10I+1C3, 1A3, fdhF • Run the gel: 20H, 20J, rush+vgb, RYC+1K3, fdhF, 13K+10I, 1A3 • Cut and purification: fdhF, 13K+10I, 1A3
• Culture: CFP, YFP 		• Purification: Rush PCR
• Cut: 22B with X, rush PCR results with S
Sept.20th Tuesday
• Run the 20H, 20J from PCR results.
• Transform: fdhF+13K+10I+1A3 		• Purification: 20B, Rush
• Ligation: 22B+Rush 		• Purification: ligation results
• Cut: the ligation results with X+S 		• PCR 20H
• Self-ligation: rush+22M
Sept.21st Wednesday
• Colony PCR: fdhF+13K+10I+1A3 on plates • Run the gel of PCR results • Culture the positive results.
Sept.22nd Thursday
• Miniprep: fdhF+13K+10I+1A3
• Preserve the above. 		• Cut and run the results: fdhF+13K+10I+1A3, 13K+10I+1C3 • Cut: RFP with E+S, YFP with X+P, 1K3 with E+P, 1C3 with E+P • Purification the digestion results.
Sept.23rd Friday
• Ligate: RFP+1A3/ YFP+1C3/1K3, RFP+1A3/1C3 • Transform the ligation results.
Sept. 24th Saturday
• Order primers for 20H, 20J
• Culture 20H-1, 20J-1, 22B-1 		• Plate the ligation results
• Culture RFP+YFP+CFP+1K3 		• Colony PCR: 3A, 1C3
• Preserve: 1K3, RFP+YFP+1C3.
Sept.25th Sunday
• Miniprep: 20J, 20H, RFP+YFP+CFP+1K3 1A3 • Run the gel of PCR results • Cut RFP+YFP+CFP+1K3 with E+P
• Colony PCR: 13K+10I+1C3 		• PCR: fdhF+rush+Vgb+tetR
• Preserve: 20H, 20J, 22B 		• Run the gel: 20H, 20J, 22B
• Cut the bands and purification. 		• Run the gel: 13K+10I+1C3, RFP+YFP+CFP+1K3
• Culture: 13K+10I+1C3
• Grad PCR: fdhF.

 

3rd Sep

Directly use DH5α fdhF+RFP colony to inoculate silicone tube set with 5ml LB. rest for 12h for biofilm attachment.

4th Sep

Add 50mlLB+Amp and start circulation.

6th Sep

Biofilm with direct inoculation is optimum. Expressed red florescence but only one small fraction of the biofilm remained after cutting off the part of the tube.

12th Sep

4 silicone tube sets with biofilm attachement time of 3h, 5h, 7.5h and 10h; 4 bubbling sets.

15th Sep

The silicone tube set with 3h of attachment achieves optimum biofilm formation. Bubbling sets glass slides are froze in -80℃ for later observation with laser confocal. Biofilm formed on glass slide is about 10μm thick.

16th Sep

New sets with silicone tube, a mixture of RFP, CFP and YFP expressing e.coli. Attachment 3h.

17th Sep

Glass slide with bubbling set with the same mixture of e.coli.
Place two filter film on solid culture medium and placed the mixture of e.coli on the filter to form gas-solid interface biofilm.

20th Sep

Observation of glass slide and gas-solid biofilm under laser confocal. Glass slide are observed directly and gas-solid biofilm is first pressed by a covering slide so that it would stick to it and peeled off from the filter for observation.
Biofilm on glass slide is about 8.6μm thick. Biofilm formed on gas-solid interface is too concentrated in bacteria and culture medium that the florescence of too strong to see any clear structure.

 

Retrieved from "http://2011.igem.org/Team:ZJU-China/Notebook/September"