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Team:Warsaw/Calendar-Main/21 August 2008
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<h3>Tests for ampicillin resistance given by protein added to medium</h3><h4>Piotr</h4> | <h3>Tests for ampicillin resistance given by protein added to medium</h3><h4>Piotr</h4> | ||
| - | Results of growth from | + | Results of growth from previous day |
<br><br> | <br><br> | ||
<table id="result"> | <table id="result"> | ||
<tr><th>OmpA variant ("hunter")</th><th>"Prey" variant</th><th>Growth with prey</th><th>Growth without prey</th></tr> | <tr><th>OmpA variant ("hunter")</th><th>"Prey" variant</th><th>Growth with prey</th><th>Growth without prey</th></tr> | ||
<tr><th>OmpA-alpha</th><td>His+Z+omega</td><td>+</td><td>+</td></tr> | <tr><th>OmpA-alpha</th><td>His+Z+omega</td><td>+</td><td>+</td></tr> | ||
| - | <tr><th>OmpA-alpha</th><td>His+Z+ | + | <tr><th>OmpA-alpha</th><td>His+Z+alpha</td><td>+</td><td>+</td></tr> |
<tr><th>OmpA-A-alpha</th><td>His+Z+omega</td><td>+</td><td>-</td></tr> | <tr><th>OmpA-A-alpha</th><td>His+Z+omega</td><td>+</td><td>-</td></tr> | ||
| - | |||
<tr><th>Ompa-A-omega</th><td>His+Z+alpha</td><td>+</td><td>-</td></tr> | <tr><th>Ompa-A-omega</th><td>His+Z+alpha</td><td>+</td><td>-</td></tr> | ||
<tr><th>OmpA-omega</th><td>His+Z+alpha</td><td>+</th><td>-</td></tr> | <tr><th>OmpA-omega</th><td>His+Z+alpha</td><td>+</th><td>-</td></tr> | ||
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</table> | </table> | ||
| - | + | <br><br> | |
| + | <ol><li> <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation">Isolation of plasmids</a> from above cultures. </li> | ||
| + | <li> Control | ||
| + | <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#digest">digest</a> of isolated plasmids with BamHI and SacI.</li> <li> Gel electrophoresis - all constructs apart from <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-alpha>pACYC177+OmpA_alpha</a> were successfully recovered.</li> | ||
| + | </ol></p> | ||
<br> | <br> | ||
| - | + | <p><b>Conclusions:</b> | |
| + | <ol> | ||
| + | <li>Almost none of tested constructs give resistance to ampicillin without prey protein added to culture medium.</li> | ||
| + | <li>Interaction between hunter and prey is not necessary to make cells ampicillin resistant (OmpA_alpha and OmpA_omega grow with prey).</li> | ||
| + | <li><a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-alpha>pACYC177+OmpA_alpha</a> culture may be contaminated with antibiotic-resistant bacteria, alpha doesn't need interaction of A and Z proteins to bind to omega - maybe we must give bacteria less prey.</li></ol></p> | ||
| - | |||
| - | |||
<h3>Cloning of protein A DNA to GeneArt plasmid</h3><h4>Antoni</h4> | <h3>Cloning of protein A DNA to GeneArt plasmid</h3><h4>Antoni</h4> | ||
<p><ol> | <p><ol> | ||
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<a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#AP+NotI">AP+NotI</a> and <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#AL+SacI">AL+SacI</a>. </li> | <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#AP+NotI">AP+NotI</a> and <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#AL+SacI">AL+SacI</a>. </li> | ||
<li>Gel electrophoresis.</li> | <li>Gel electrophoresis.</li> | ||
| - | <li> Confirmed transformant colonies inoculated to liquid LB with ampicillin | + | <li> Confirmed transformant colonies inoculated to liquid LB with ampicillin.</li> |
| - | + | ||
| - | + | ||
</ol></p> | </ol></p> | ||
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<h3>Cloning of alpha to <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-A-omega>pACYC177+OmpA_A_omega</a> and <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-Z-omega>pACYC177+OmpA_Z_omega</a></h3><h4>Michał K.</h4> | <h3>Cloning of alpha to <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-A-omega>pACYC177+OmpA_A_omega</a> and <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-Z-omega>pACYC177+OmpA_Z_omega</a></h3><h4>Michał K.</h4> | ||
<p><ol> | <p><ol> | ||
| - | <li> <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation">Isolation of plasmids</a> from cultures | + | <li> <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation">Isolation of plasmids</a> from cultures inoculated on previous day. </li> |
<li> Control | <li> Control | ||
| - | <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#digest">digest</a> of isolated plasmids with FastBamHI and FastAcc65I (Fast Digest buffer).</li> <li> Gel electrophoresis - no proper clones | + | <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#digest">digest</a> of isolated plasmids with FastBamHI and FastAcc65I (Fast Digest buffer).</li> |
| + | <li> Gel electrophoresis - no proper clones found (<a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/21_August_2008#fig1">Fig. 1.</a>).</li> | ||
</ol></p> | </ol></p> | ||
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| + | <a name="fig1"><img src="https://static.igem.org/mediawiki/2008/f/f3/Traw_04_09_2008.jpg" width=300/></a> <var><b>Fig. 1.</b> Control Acc65I/BamHI digests of isolated plasmids<br> | ||
| + | 1. Marker<br> | ||
| + | 2-3. digested plasmids pACYC177+OmpA_A_omega+alpha <br> | ||
| + | 4-5. digested plasmids pACYC177+OmpA_Z_omega+alpha <br></var> | ||
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Latest revision as of 19:09, 26 October 2008
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Tests for ampicillin resistance given by protein added to mediumPiotrResults of growth from previous day
Conclusions:
Cloning of protein A DNA to GeneArt plasmidAntoni
Cloning of alpha to pACYC177+OmpA_A_omega and pACYC177+OmpA_Z_omegaMichał K.
 Fig. 1. Control Acc65I/BamHI digests of isolated plasmids1. Marker 2-3. digested plasmids pACYC177+OmpA_A_omega+alpha 4-5. digested plasmids pACYC177+OmpA_Z_omega+alpha
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