Difference between revisions of "Team:Pasteur Paris/Protocol"

Revision as of 09:46, 17 October 2016

Protocols

During our lab work we used the following protocols. You can also download all the protocols in one file here:

Experiments	Aim:	Download
PCR	To reproduce (amplify) selected sections of DNA (inserts A1, A2, B1, B2, C1, C2, E1,E2).
Digestion	To linearize the different plasmids with appropriate enzymes. Perform restriction enzyme digestion in order to recover linear backbones of the plasmids. And chose the appropriate restriction sites based on the host plasmids.
Dephosphorylation	To reduce vector self-ligation and favor that of vector to insert instead.
Ligation	To link an insert to its host plasmid before the transformation. This step happens after the digestion of the insert and the plasmid with the same restriction sites enzymes.
Electrophoresis	To know the size of DNA fragments to check the efficiency of a digestion for instance.
Gel extraction kit	To get back the DNA purified thanks to the electrophoresis on agarose gel.
Transformation	To increase the amount of plasmid by transformation in competent cells. The amount of plasmid supplied is insufficient to perform all our future experiments.
Harvest and culture	In order to obtain a large amount of plasmid, we need to grow the bacteria overnight.
Stab culture	To store a clone of bacteria to be used later.
IPTG induction	To increase the production of our protein which depends on this molecule.
Protein Purification Fast Protein Liquid Chromatography	To check if the His-tag works and if our protein has really been produced
Cellulose-binding Protocol	Verify that the C2 protein binds to cellulose.
Silification & assay Protocol	Prepare a one pot mixture that will be compressed directly after incubation.
Patch compression Protocol	Make a patch by compression with cellulose and the silicated C2 protein.
One pot mixture-Patch protocol	Prepare a one pot mixture that will be compressed directly after incubation.
Antibody-binding Protocol

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        <td>In order to obtain a large amount of plasmid, we need to grow the bacteria overnight.</td>
        <td> <center><a href="https://static.igem.org/mediawiki/2016/4/4b/T--Pasteur_Paris--Bacterial_culture_protocol.pdf"><img src="https://static.igem.org/mediawiki/2016/c/c1/DownloadT--iGEM16_Pasteur_Paris--file.xxx.png" width="40%" alt=""></a></center></td>
+    </tr>
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+      <td><strong>Stab culture</strong></td>
+      <td>To store a clone of bacteria to be used later.</td>
+      <td> <center><a href="https://static.igem.org/mediawiki/2016/b/b1/Stab_culture_Pasteur_Paris_2016.pdf"><img src="https://static.igem.org/mediawiki/2016/c/c1/DownloadT--iGEM16_Pasteur_Paris--file.xxx.png" width="40%" alt=""></a></center></td>
      </tr>
      <tr>