Team:Calgary/Notebook/Protocols/AmpicillinSurvivalAssay1

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<div id="Banner"><h1>Ampicillin Survival Assay 2</h1></div>
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<h1>Ampicillin Survival Assay 1</h1>
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<p class="noIndent">We used this protocol to verify if, after protein purification, beta-lactamase was functional when linked to one of our TALEs.</p>
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<h1>Ampicillin Survival Assay 2</h1>
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<h2>Reagents and Materials</h2>
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<li>LB Broth</li>
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<li>Ampicillin (100mg/mL)</li>
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<li>Chloramphenicol (30mg/mL)</li>
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<li>Falcon tubes</li>
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<li>Bacteria resistant to chloramphenicol but not ampicillin</li>
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<li>Bacteria resistant to both ampicillin and chloramphenicol</li>
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<li>Purified beta-lactamase protein</li>
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<h2>Protocol</h2>
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<li>To each Falcon tube, add 5mL of LB Broth, 5µL of ampicillin and 3µL of chloramphenicol</li>
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<li>Add your purified proteins in multiple tubes with different amounts of proteins; for construct we added the following: 20µg, 15µg, 10µg, 5µg, 1µg, 0.1µg and 0.01µg</li>
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<li>Incubate the tubes for 1h at 37°C</li>
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<li>For the positive control, inoculate a tube that doesn’t have protein on it with a colony of  your bacteria resistant to both antibiotics</li>
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<li>Add colonies of your bacteria resistant to chloramphenicol but not ampicillin to the other tubes</li>
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<li>Grow the cells overnight</li>
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Revision as of 23:50, 27 September 2013

Ampicillin Survival Assay 2

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