Team:Warsaw/Calendar-Main/30 July 2008

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<h3>Checking if OmpA_omega_A_alpha gives ampicillin resistance </h3>
 
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<h4>Piotr</h4>
 
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<p>Inoculation of OmpA_omega_A_alpha from various IPTG concentrations: 0, 0.1, 0.25, 0.5, 0.75, 1 mM into same IPTG concentrations, but with various ampicillin concentrations (25, 50, 75, 100 mM) in ratio 1:50.</p>
 
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<table id="result" align="center">
 
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<thead>
 
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<b>Bacterial growth (OD) measurement in the evening:</b>
 
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</thead>
 
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<tr><th rowspan="2">ampicillin concentration (μg/mL):</th><th colspan="6">IPTG concentration (mmol/mL):</td></tr>
 
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<tr><th>0</th><th>0.1</th><th>0.25</th><th>0.5</th><th>0.75</th><th>1</th></tr>
 
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<tr><th>25</th><td class="live">1.054</td><td class="live">1.154</td><td class="live">1.051</td><td class="live">0.99</td><td class="live">1.096</td><td class="live">0.896</td></tr>
 
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<tr><th>50</th><td class="live">0.94</td><td class="live">0.891</td><td class="live">1.123</td><td class="live">0.847</td><td class="live">0.924</td><td class="live">0,81</td></tr>
 
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<tr><th>75</th><td class="live">0.5</td><td class="live">0.63</td><td class="live">0.743</td><td class="live">0.782</td><td class="live">0.631</td><td class="live">0.64</td></tr>
 
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<tr><th>100</th><td class="live">0.02</td><td class="live">0.396</td><td class="live">0.563</td><td class="live">0.678</td><td class="live">0.602</td><td class="live">0.611</td></tr>
 
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</table>
 
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<p>Conclusion: we obtained the best induction using 0.5 - 0.25 mM IPTG. Best ampicillin concentration is 50 - 75 μg/ml.</p>
 
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<h3>
 
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Checking if degradation of fusion with OmpA is a result of the activity of lon and iompt proteases. (present in <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#top10">top10</a>)<br>
 
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Piotr</h3>
 
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<p>Test was conducted in <i>E.coli</i> <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#rosetta">Rosetta</a> strain expressing omp_omega_A_alfa (with and without induction) and omp_A_alfa. </p>
 
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<p>Transformation of <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#rosetta">Rosettas</a> with omega_A_alpha and A_alpha.</p>
 
<h4>Michał K.:</h4>
<h4>Michał K.:</h4>

Revision as of 21:46, 11 October 2008

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  1. Separate transformant colonies (tranformation of ligation of pACYC177+OmpA_alpha and omega_A from previous day) inoculated to liquid LB with kanamycin.

Michał K.:

  1. Isolation of plasmids from cultures inocluated on previous day.
  2. Control digest of isolated plasmids (pACYC177+omega_deltaA) with HindIII and SacI (7 from 8 colonies confirmed).