Difference between revisions of "Team:USTC/Experiments"

 
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<title>子网页测试-队员</title>
 
<title>子网页测试-队员</title>
 
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<body>
 
<body>
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                   <li class="bg_all"><a href="https://2017.igem.org/Team:USTC/Description">Description</a></li>
 
                   <li class="bg_all"><a href="https://2017.igem.org/Team:USTC/Description">Description</a></li>
 
                   <li class="bg_all"><a href="https://2017.igem.org/Team:USTC/Design">Design</a></li>
 
                   <li class="bg_all"><a href="https://2017.igem.org/Team:USTC/Design">Design</a></li>
−
                   <li class="bg_all"><a href="https://2017.igem.org/Team:USTC/Results">Results</a></li>
+
                   <li class="bg_all"><a href="https://2017.igem.org/Team:USTC/Demonstrate">Results</a></li>
−
                   <li class="bg_all"><a href="https://2017.igem.org/Team:USTC/Demonstrate">Demonstrate</a></li>
+
                   <li><a href="https://2017.igem.org/Team:USTC/Demonstrate/1">&nbsp;&nbsp;>Conduction</a></li>
 +
                  <li><a href="https://2017.igem.org/Team:USTC/Demonstrate/2">&nbsp;&nbsp;>Photocatalyst</a></li>
 +
                  <li><a href="https://2017.igem.org/Team:USTC/Demonstrate/3">&nbsp;&nbsp;>Harvest</a></li>
 +
                 
 
                 </ul>
 
                 </ul>
 
               </div>
 
               </div>
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<a href="https://2017.igem.org/Team:USTC/Safety" class="waves-effect waves-teal">Safety</a>
 
<a href="https://2017.igem.org/Team:USTC/Safety" class="waves-effect waves-teal">Safety</a>
 
</li>
 
</li>
−
<li class="bold1 mintcream_choosed">
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<li class="bold honeydew_choosed"><a class="collapsible-header  waves-effect waves-teal">Model</a>
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<a href="https://2017.igem.org/Team:USTC/Model" class="waves-effect waves-teal">Model</a>
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              <div class="collapsible-body">
−
</li>
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                <ul>
 +
                  <li class="bg_all"><a href="https://2017.igem.org/Team:USTC/Model">Overview</a></li>
 +
                  <li class="bg_all"><a href="https://2017.igem.org/Team:USTC/Model/2">DLA Crystal</a></li>
 +
                  <li id="static_word" class="bg_all"><a>Electron transfer</a></li>
 +
                  <li><a href="https://2017.igem.org/Team:USTC/Model/4">&nbsp;&nbsp;>Semi-conductor</a></li>
 +
                  <li><a href="https://2017.igem.org/Team:USTC/Model/3">&nbsp;&nbsp;>Markov</a></li>
 +
                  <li><a href="https://2017.igem.org/Team:USTC/Model/1">&nbsp;&nbsp;>MeCiM</a></li>
 +
                  <li class="bg_all"><a href="https://2017.igem.org/Team:USTC/Model/5">UPEP</a></li>
 +
                </ul>
 +
              </div>
 +
            </li>
  
 
             <li class="bold honeydew_choosed"><a class="collapsible-header  waves-effect waves-teal">Parts</a>
 
             <li class="bold honeydew_choosed"><a class="collapsible-header  waves-effect waves-teal">Parts</a>
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<div >
 
<div >
 
                                                         <br>
 
                                                         <br>
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        <p id="first" class="scrollspy label label-pink">Protocols</p>
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        <p id="first" class="scrollspy label label-pink">Protocols</p><br><br>
 +
        <a href="https://static.igem.org/mediawiki/2017/f/f9/USTC-note-pro.pdf" class="indent_word">Click here to check out the protocols of our experiments!</a>
 +
                                                        <br><br><br>
 +
</div>
 +
<div >
 
                                                         <br>
 
                                                         <br>
−
                                                        <br>
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        <p id="second" class="scrollspy label label-pink">Materials</p><br><br>
−
                                                        <p>Plasmid Extraction</p>
+
        <a href="https://static.igem.org/mediawiki/2017/d/dd/USTC-note-mat.pdf" class="indent_word">Click here to check out the preparation for different materials we have used!</a>
−
                                                        <p>Materials
:</p>
+
                                                         <br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br>
−
                                                        <p>Buffer P1 (stored in 4 degree)
</p>
+
−
                                                        <p>Buffer P2
</p>
+
−
                                                        <p>Buffer p3
</p>
+
−
                                                        <p>VisualLyse</p>
+
−
                                                        <p>
Buffer DW1
</p>
+
−
                                                        <p>Wash Solution
dd </p>
+
−
                                                        <p>ddH<sub>2</sub>O</p>
+
−
                                                        <p>
Prepation
</p>
+
−
                                                        <p>1.Check out whether RNaseA has been added in Buffer P1.
</p>
+
−
                                                        <p>2.Check out whether ethyl alcohol has been added in Wash Solution
</p>
+
−
                                                        <p>3.Check out whether sediment exist in Buffer P2 and P2.

+
−
                                                        <p>Procedure
</p>
+
−
                                                        <p>4.Absorb 1.5 to 5 mL bacteria solution in to EP tubes, centrifuge them at 8,000xg 2 mins and then discard the culture medium.</p>
+
−
                                                        <p>5.Add 250 ul Buffer P1 into sediment, and use spearhead to make bacteria suspended.
</p>
+
−
                                                        <p>6.Add 250 ul Buffer P2, and overturn the EP tubes 5 to 10 times immediately and tenderly. Stand tubes about 2 to 4 mins.
</p>
+
−
                                                        <p>7.Add 350 ul Buffer P3 and overturn the EP tubes 5 to 10 times again immediately and tenderly.
</p>
+
−
                                                        <p>8.Centrifuge tubes at 12,000xg about 5 to 10 mins.
</p>
+
−
                                                        <p>9.Pour the supernatant liquid into absorption columns ,centrifuge them at 8,000xg about 30 sec and then discard the liquid in the collecting pipe.
</p>
+
−
                                                        <p>10.optional:Add 500 ul Buffer DW1 and centrifuge them at 9,000xg about 30 sec. Then discard the liquid in the collecting pipe.
</p>
+
−
                                                        <p>11.Add 500 ul Wash Solution, centrifuge them at 9,000xg about 30 sec and discard the liquid in the collecting pipe.</p>
+
−
                                                        <p>
12.Do the step 11 again.
</p>
+
−
                                                        <p>13.Centrifuge the empty columns at 9,000xg about 1 min.</p>
+
−
                                                        <p>
14.Put the columns in clean 1.5mL EP tubes, add 50 to 100 ul Elution Buffer on the absorption film, stand 10 min and centrifuge about 1 min at 9,000xg.
</p>
+
−
                                                        <p>15.Keep the DNA solution for further work.
The Protocol is based on SanPrep Column Type DNA Plasmid Extraction Kit.</p>
+
−
 
+
−
                                                        <br>
+
−
                                                        <br>
+
−
                                                        <p>PCR System Preparation and Conditions Setting</p>
+
−
                                                        <p>PCR system(set 50ul system as an example): 
The amount of substance of premier is written on the EP tubes. Calculating the volume of 1XTE required for 10uM primer solution is required before the system preparation.</p>
+
−
                                                        <p>
1.Template DNA xul as required.
</p>
+
−
                                                        <p>2.Forward Primer(10 uM) about 1 to 2 ul. The final concentration will be 0.2 to 0.4 uM.
</p>
+
−
                                                        <p>3.Reverse Primer(10 uM) about 1 to 2 ul. The final concentration will be 0.2 to 0.4 uM.
</p>
+
−
                                                        <p>4.5xTransStart FastPfu Fly Buffer 10 ul.
</p>
+
−
                                                        <p>5.2.5 mM dNTPs 5 uM. The final concentration will be 0.25 mM.
</p>
+
−
                                                        <p>6.TransStart FastPfu Fly DNA Polymerase 1 ul.
</p>
+
−
                                                        <p>7.Double distilled water added to 50 ul.
Reaction conditions
</p>
+
−
                                                        <p>8.Calculate the Tm Value. If the complementary strip is smaller than 20 bp, Tm=4X(A+T)+2X(C+G), and the annealing temperature is Tm-5 degree centigrade. If not, use software to figure out the exact Tm value.(Also Tm value exist in report)
PCR cycle:</p>
+
−
                                                        <p>
9.1 cycle of 95 degree centigrade about 2 min for pre-degeneration;
</p>
+
−
                                                         <p>10.30 to 35 cycles of 95 degree centigrade about 10 sec, Tm-5 degree centigrade about 20 sec and 72 degree centigrade 1kb per min;
</p>
+
−
                                                        <p>11.1 cycle of 75 degree centigrade about 5 mins and stay the device in 4 degree centigrade.
</p>
+
−
                                                        <p>The protocol is based on TransStart FastPfu Fly DNA Polymerase.</p>
+
−
 
+
−
                                                        <br>
+
−
                                                        <br>
+
−
                                                        <p>PCR of Bacteria System Preparation and Conditions Setting</p>
+
−
                                                        <p>PCR system(set 50ul system as an example): 
</p>
+
−
                                                        <p>The amount of substance of premier is written on the EP tubes. Calculating the volume of 1XTE required for 10uM primer solution is required before the system preparation.
</p>
+
−
                                                        <p>1.Template(bacteria fluid) about 1 ul.
</p>
+
−
                                                        <p>2.Forward Primer(10 uM) about 1 to 2 ul. The final concentration will be 0.2 to 0.4 uM.
</p>
+
−
                                                        <p>3.Reverse Primer(10 uM) about 1 to 2 ul. The final concentration will be 0.2 to 0.4 uM.</p>
+
−
                                                        <p>
4.5xTransStart FastPfu Fly Buffer 10 ul.
</p>
+
−
                                                        <p>5.2.5 mM dNTPs 5 uM. The final concentration will be 0.25 mM.
</p>
+
−
                                                        <p>6.TransStart FastPfu Fly DNA Polymerase 1 ul.
</p>
+
−
                                                        <p>7.Double distilled water added to 50 ul.
Reaction conditions
.</p>
+
−
                                                        <p>8.Calculate the Tm Value. If the complementary strip is smaller than 20 bp, Tm=4X(A+T)+2X(C+G), and the annealing temperature is Tm-5 degree centigrade. If not, use software to figure out the exact Tm value.(Also Tm value exist in report)
PCR cycle:</p>
+
−
                                                        <p>
9.1 cycle of 95 degree centigrade about 2 min for pre-degeneration;
</p>
+
−
                                                        <p>10.30 to 35 cycles of 95 degree centigrade about 10 sec, Tm-5 degree centigrade about 20 sec and 72 degree centigrade 1kb per min;</p>
+
−
                                                        <p>
11.1 cycle of 75 degree centigrade about 5 mins and stay the device in 4 degree centigrade.
The protocol is based on TransStart FastPfu Fly DNA Polymerase.</p>
+
−
                                                        <p>PCR System Preparation and Conditions Setting
PCR system(set 50ul system as an example): </p>
+
−
                                                        <p>
The amount of substance of premier is written on the EP tubes. Calculating the volume of 1XTE required for 10uM primer solution is required before the system preparation.
</p>
+
−
                                                        <p>1.Template DNA xul as required.
</p>
+
−
                                                        <p>2.Forward Primer(10 uM) about 1 to 2 ul. The final concentration will be 0.2 to 0.4 uM.</p>
+
−
                                                        <p>
3.Reverse Primer(10 uM) about 1 to 2 ul. The final concentration will be 0.2 to 0.4 uM.
</p>
+
−
                                                        <p>4.5xTransStart FastPfu Fly Buffer 10 ul.</p>
+
−
                                                        <p>
5.2.5 mM dNTPs 5 uM. The final concentration will be 0.25 mM.
</p>
+
−
                                                        <p>6.TransStart FastPfu Fly DNA Polymerase 1 ul.
</p>
+
−
                                                        <p>7.Double distilled water added to 50 ul.
Reaction conditions
</p>
+
−
                                                        <p>8.Calculate the Tm Value. If the complementary strip is smaller than 20 bp, Tm=4X(A+T)+2X(C+G), and the annealing temperature is Tm-5 degree centigrade. If not, use software to figure out the exact Tm value.(Also Tm value exist in report)
PCR cycle:</p>
+
−
                                                        <p>
9.1 cycle of 95 degree centigrade about 2 min for pre-degeneration;
</p>
+
−
                                                        <p>10.30 to 35 cycles of 95 degree centigrade about 10 sec, Tm-5 degree centigrade about 20 sec and 72 degree centigrade 1kb per min;
</p>
+
−
                                                        <p>11.1 cycle of 75 degree centigrade about 5 mins and stay the device in 4 degree centigrade.
</p>
+
−
                                                        <p>The protocol is based on TransStart FastPfu Fly DNA Polymerase.</p>
+
−
 
+
 
</div>
 
</div>
−
 
−
                                        <div>
 
−
                                                  <br>
 
−
  <span id="second" class="scrollspy label label-pink">Materials</span>
 
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                                                  <br>
 
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<img src="https://static.igem.org/mediawiki/2017/4/49/Ustc_USTCIF.jpg" width="8%" height="8%">
 
<img src="https://static.igem.org/mediawiki/2017/4/4f/Ustc_ustc_teach.png" width="45%" height="78%">
 
<img src="https://static.igem.org/mediawiki/2017/4/4f/Ustc_ustc_teach.png" width="45%" height="78%">
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<img src="https://static.igem.org/mediawiki/2017/thumb/6/65/Ustc_ustc_bio.jpg/1200px-Ustc_ustc_bio.jpg.png" width="45%" height="77%" style="margin-bottom: -7px">
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<img src="https://static.igem.org/mediawiki/2017/thumb/6/65/Ustc_ustc_bio.jpg/1200px-Ustc_ustc_bio.jpg.png" width="45%" height="77%" style="margin-bottom: -12px">
 
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Latest revision as of 15:21, 29 October 2017

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