Team:Toulouse/Notebook/Protocols

From 2014.igem.org

(Difference between revisions)
 
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- Put the tubes 20 minutes in the ice </i>
- Put the tubes 20 minutes in the ice </i>
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- Put the tubes 2 minutes at 42°C in the water bath
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- Put the tubes 2 minutes at 42°C in the water bath to create the thermic shock
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- Put the tubes back in ice immediately to create the thermic shock
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- Put the tubes back in ice immediately  
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- Add 1mL of LB medium
- Add 1mL of LB medium
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<p class="texte">
<p class="texte">
<B> Day 0 </B>
<B> Day 0 </B>
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<br/>- Streak out the <i>Bacillus</i> strain and plate this on an LB agar plate overnight at 37°C</p>
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<br/>- Streak out the <i>B. subtilis</i> strain and plate this on an LB agar plate overnight at 37°C</p>
<p class="texte"><B> Day 1 </B>
<p class="texte"><B> Day 1 </B>
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<br/>- Pick up a nice big colony of <I>B. Subtilis </I> strain and drop it in 2ml of completed 1x MC
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<br/>- Pick up a nice big colony of <I>B. subtilis </I> strain and drop it in 2ml of completed 1x MC
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- Grow at 37°C for 5 hours
- Grow at 37°C for 5 hours
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<p class="title1" id="select7">Test of the pSB<sub>BS</sub>4S plasmid integration in <i>Bacillus subtilis</i> genome on the threonine site</p>
<p class="title1" id="select7">Test of the pSB<sub>BS</sub>4S plasmid integration in <i>Bacillus subtilis</i> genome on the threonine site</p>
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<p class="texte">
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<br>- Plate the transformed <i>Bacillus</i> strain on a selective medium (LB + spectinomycin) overnight  
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<br>- Plate the transformed <i>B. subtilis</i> strain on a selective medium (LB + spectinomycin) overnight  
<br>- The obtained clones are then plated on different media:  Medium Competence (Thr+), Medium Competence (Thr-) and LB + Spectinomycin.  
<br>- The obtained clones are then plated on different media:  Medium Competence (Thr+), Medium Competence (Thr-) and LB + Spectinomycin.  
<br>When the plasmid is integrated, the clone can grow on minimum medium with threonine and on LB + Spectinomycin but can not grow on the minimum medium without thronine.
<br>When the plasmid is integrated, the clone can grow on minimum medium with threonine and on LB + Spectinomycin but can not grow on the minimum medium without thronine.

Latest revision as of 03:37, 18 October 2014