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| − | <h1 class="sectionedit1"><a name="sporulation" id="sporulation">Sporulation</a></h1>
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| − | <div class="level1">
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| − | <p> | + | <div class="column full_size" > |
| − | <strong>Checklist:</strong><br/> | + | <h5 style="text-align: center"> Lab Journals </h5> |
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| − | </p>
| + | <div class=" para_center_20"> |
| − | <ul>
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| − | <li class="level1"><div class=" li" > Use the right Medium.</div> | + | |
| − | </li>
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| − | <li class="level1"><div class="li"> Vortex cuvettes if you make a dilution</div>
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| − | </li>
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| − | <li class="level1"><div class="li"> Set Nanodrop to cuvettes</div>
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| − | </li>
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| − | </ul>
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| − | <p>
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| − | 1. Overnight culture
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| − | </p>
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| − | <ul>
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| − | <li class="level1"><div class="li"> inoculate your culture in 5ml LB-Medium</div>
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| − | </li>
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| − | <li class="level1"><div class="li"> let them grow over night at 37°C, 200rpm</div>
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| − | </li>
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| − | </ul>
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| − | <p>
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| − | <br/>
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| − | 2. Exponential growth
| + | </div> |
| − | </p> | + | |
| − | <ul>
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| − | <li class="level1"><div class="li"> measure the OD600 of your overnight culture</div>
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| − | </li>
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| − | <li class="level1"><div class="li"> inoculate 2ml + Xml to measure OD (personal preference) LB-Medium to OD600=0,1 from overnight culture </div>
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| − | </li>
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| − | <li class="level1"><div class="li"> let the cells grow to an OD600=0,8 (37°C, 200rpm)</div>
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| − | </li>
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| − | </ul>
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| − | | + | |
| − | <p>
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| − | 3. Sporulation
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| − | </p>
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| − | <ul>
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| − | <li class="level1"><div class="li"> centrifuge the 2 ml of cells at 13.000 rpm for 1 minute</div>
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| − | </li>
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| − | <li class="level1"><div class="li"> wash the pellet with PBS</div>
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| − | </li>
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| − | <li class="level1"><div class="li"> resuspend the pellet in 1 ml DSM (Difco Sporulation Medium)<a href="/igem2016/doku.php?id=labprotocols:media" class="wikilink1" title="labprotocols:media">Culture medium</a></div>
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| − | </li>
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| − | <li class="level1"><div class="li"> let the cells grow (24h,37°C 200rpm)</div>
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| − | </li>
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| − | </ul>
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| − | | + | |
| − | <p>
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| − | 4. Lysozym treatment
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| − | </p>
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| − | <ul>
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| − | <li class="level1"><div class="li"> dilute the cells 6:1 in lysozym solution (15mg/μl)</div>
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| − | </li>
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| − | <li class="level1"><div class="li"> incubate for 1 hour at room temperature</div>
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| − | </li>
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| − | <li class="level1"><div class="li"> wash 6 times with 1x PBS</div>
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| − | </li>
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| − | </ul>
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| − | | + | |
| − | <p>
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| − | 5. Count the spores (Neubauer-counting chamber, usually dilute in BPS 1 to 100)<br/>
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| − | <a href="/igem2016/lib/exe/fetch.php?media=labprotocols:sorecountingb54.png" class="media" title="labprotocols:sorecountingb54.png"><img src="/igem2016/lib/exe/fetch.php?w=400&media=labprotocols:sorecountingb54.png" class="media" alt="" width="400" /></a><br/>
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| − | 6. Make aliquots (around 100million spores per aliquot)
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| − | </p>
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| | </div> | | </div> |
| − | <!-- EDIT1 SECTION "Sporulation" [1-1087] -->
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| − | <h1 class="sectionedit2"><a name="induktion_of_sporulation" id="induktion_of_sporulation">Induction of sporulation</a></h1>
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| − | <div class="level1">
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| − | <p>
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| − | <a href="https://www.researchgate.net/post/How_to_induce_sporulation_of_Bacillus_spp_by_heating_or_any_other_simple_technique" class="urlextern" target="_Blank" title="https://www.researchgate.net/post/How_to_induce_sporulation_of_Bacillus_spp_by_heating_or_any_other_simple_technique" rel="nofollow">https://www.researchgate.net/post/How_to_induce_sporulation_of_Bacillus_spp_by_heating_or_any_other_simple_technique</a><br/>
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| − | <a href="https://www.researchgate.net/post/How_to_sporulate_the_bacteria_or_how_to_induce_the_sporulation_in_bacteria_especially_for_Bacillus_spp" class="urlextern" target="_Blank" title="https://www.researchgate.net/post/How_to_sporulate_the_bacteria_or_how_to_induce_the_sporulation_in_bacteria_especially_for_Bacillus_spp" rel="nofollow">https://www.researchgate.net/post/How_to_sporulate_the_bacteria_or_how_to_induce_the_sporulation_in_bacteria_especially_for_Bacillus_spp</a><br/>
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| − |
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| − | </p>
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| − | <div class="tags"><span>
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| − | <a href="/igem2016/doku.php?id=tag:workwithbs_labprotocols" class="wikilink1" title="tag:workwithbs_labprotocols" rel="tag">WorkwithBS labprotocols</a>
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| − | </span></div>
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| − |
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| − | </div>
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| − | <!-- EDIT2 SECTION "Induktion of sporulation" [1088-] -->
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| | </html> | | </html> |
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