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Team:NTNU Trondheim/Protocols
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<div style="top:55px">Media</div> | <div style="top:55px">Media</div> | ||
</li> | </li> | ||
| - | <li id=" | + | <li id="week1" onclick="weekFilter(this)"> |
| - | <a class="nb-week" href="#1"> | + | <a class="nb-week" href="#1">Lysogeny Broth (LB)</a> |
</li> | </li> | ||
| - | <li id=" | + | <li id="week2" onclick="weekFilter(this)"> |
| - | <a class="nb-week" href="#2"> | + | <a class="nb-week" href="#2">SOC medium</a> |
</li> | </li> | ||
| - | <li id=" | + | <li id="week3" onclick="weekFilter(this)"> |
<a class="nb-week" href="#3">yB medium</a> | <a class="nb-week" href="#3">yB medium</a> | ||
</li> | </li> | ||
| - | <li id=" | + | <li id="week4" onclick="weekFilter(this)"> |
<a class="nb-week" href="#4"><i>Synechocystis</i> medium</a> | <a class="nb-week" href="#4"><i>Synechocystis</i> medium</a> | ||
</li> | </li> | ||
| - | <li id=" | + | <li id="week5" onclick="weekFilter(this)"> |
| - | <a class="nb-week" href="# | + | <a class="nb-week" href="#5"></a> |
</li> | </li> | ||
<li class="nb-month"> | <li class="nb-month"> | ||
<div style="top:140px;margin-left:-18px">Techniques</div> | <div style="top:140px;margin-left:-18px">Techniques</div> | ||
</li> | </li> | ||
| - | <li id=" | + | <li id="week6" onclick="weekFilter(this)"> |
| - | <a class="nb-week" href="# | + | <a class="nb-week" href="#6">Gibson assembly</a> |
</li> | </li> | ||
| - | <li id=" | + | <li id="week7" onclick="weekFilter(this)"> |
| - | <a class="nb-week" href="# | + | <a class="nb-week" href="#7">DNA isolation and cleaning</a> |
</li> | </li> | ||
| - | <li id=" | + | <li id="week8" onclick="weekFilter(this)"> |
| - | <a class="nb-week" href="# | + | <a class="nb-week" href="#8">DNA digestion</a> |
</li> | </li> | ||
| - | <li id=" | + | <li id="week9" onclick="weekFilter(this)"> |
| - | <a class="nb-week" href="# | + | <a class="nb-week" href="#9">PCR</a> |
</li> | </li> | ||
| - | <li id=" | + | <li id="week10" onclick="weekFilter(this)"> |
| - | <a class="nb-week" href="# | + | <a class="nb-week" href="#10">NanoDrop</a> |
</li> | </li> | ||
| - | <li id=" | + | <li id="week11" onclick="weekFilter(this)"> |
| - | <a class="nb-week" href="# | + | <a class="nb-week" href="#11">3A assembly</a> |
</li> | </li> | ||
| - | <li id=" | + | <li id="week12" onclick="weekFilter(this)"> |
| - | <a class="nb-week" href="# | + | <a class="nb-week" href="#12">Ligation</a> |
</li> | </li> | ||
| - | <li id=" | + | <li id="week13" onclick="weekFilter(this)"> |
| - | <a class="nb-week" href="# | + | <a class="nb-week" href="#13">Transformation (<i>Escherichia coli</i>)</a> |
</li> | </li> | ||
| - | <li id=" | + | <li id="week14" onclick="weekFilter(this)"> |
| - | <a class="nb-week" href="# | + | <a class="nb-week" href="#14">Transformation (<i>Synechocystis sp. PCC 6803</i>)</a> |
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</li> | </li> | ||
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<li class="nb-month"> | <li class="nb-month"> | ||
<div style="top:52px;margin-left:-9px">Plasmids</div> | <div style="top:52px;margin-left:-9px">Plasmids</div> | ||
</li> | </li> | ||
| - | <li id=" | + | <li id="week16" onclick="weekFilter(this)"> |
| - | <a class="nb-week" href="# | + | <a class="nb-week" href="#16">Right flank</a> |
</li> | </li> | ||
| - | <li id=" | + | <li id="week17" onclick="weekFilter(this)"> |
| - | <a class="nb-week" href="# | + | <a class="nb-week" href="#17">Left flank</a> |
</li> | </li> | ||
| - | <li id=" | + | <li id="week18" onclick="weekFilter(this)"> |
| - | <a class="nb-week" href="# | + | <a class="nb-week" href="#18">Kanamycin resistance</a> |
</li> | </li> | ||
| - | <li id=" | + | <li id="week19" onclick="weekFilter(this)"> |
| - | <a class="nb-week" href="# | + | <a class="nb-week" href="#19">Lac promoter</a> |
</li> | </li> | ||
| - | <li id=" | + | <li id="week20" onclick="weekFilter(this)"> |
| - | <a class="nb-week" href="# | + | <a class="nb-week" href="#20">Glucose oxidase</a> |
</li> | </li> | ||
| - | + | <li id="week21" onclick="weekFilter(this)"> | |
| - | + | <a class="nb-week" href="#21">Left flank + Kan + Right flank</a> | |
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| - | <a class="nb-week" href="# | + | |
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</li> | </li> | ||
<li class="nb-month"> | <li class="nb-month"> | ||
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<div style="top:55px;margin-left:-29px">Calculations</div> | <div style="top:55px;margin-left:-29px">Calculations</div> | ||
</li> | </li> | ||
| - | <li id=" | + | <li id="week24" onclick="weekFilter(this)"> |
| - | <a class="nb-week" href="# | + | <a class="nb-week" href="#24">DNA concentration</a> |
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</li> | </li> | ||
</ul> | </ul> | ||
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<div>Techniques</div> | <div>Techniques</div> | ||
<div>Plasmids</div> | <div>Plasmids</div> | ||
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<div>Calculations</div> | <div>Calculations</div> | ||
</div> | </div> | ||
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| - | <div id=" | + | <div id="techniques-only" class="nb-only" onclick="onlyFilter(this)">only</div> |
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<div class="two columns nb-filter"> | <div class="two columns nb-filter"> | ||
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<div class="filter-container" onclick="filter(this)" style="background-image:url(https://static.igem.org/mediawiki/2014/b/b8/NTNU_calculations_off.png);width=100px;height=100px"> | <div class="filter-container" onclick="filter(this)" style="background-image:url(https://static.igem.org/mediawiki/2014/b/b8/NTNU_calculations_off.png);width=100px;height=100px"> | ||
| - | <img id=" | + | <img id="calc" src="https://static.igem.org/mediawiki/2014/b/bb/NTNU_calculations_on.png";width="100px" height="100px"> |
</div> | </div> | ||
| - | <div id=" | + | <div id="calc-only" class="nb-only" onclick="onlyFilter(this)">only</div> |
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<!-- PLACEHOLDER --> <div class="two columns nb-filter" style="opacity:0.00;width:65px"> | <!-- PLACEHOLDER --> <div class="two columns nb-filter" style="opacity:0.00;width:65px"> | ||
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<!-- -------------------Protocol entries------------------- --> | <!-- -------------------Protocol entries------------------- --> | ||
| - | <div id=" | + | <div id="week1entry" class="nb-week" style="display: block"> |
<div class="twelve columns"> | <div class="twelve columns"> | ||
| - | <h3 class="centered"> | + | <h3 class="centered">Lysogeny Broth (LB)</h3> |
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<div class="entry pc-media"> | <div class="entry pc-media"> | ||
<div> | <div> | ||
| - | <h6>Recipe | + | <h6>Recipe |
<div class="nb-onetech-i nohilite">Antibiotic additions</div> | <div class="nb-onetech-i nohilite">Antibiotic additions</div> | ||
</h6> | </h6> | ||
<div class="nb-tech"> | <div class="nb-tech"> | ||
| + | <div class="AB-table"> | ||
| + | <table> | ||
| + | <tr> | ||
| + | <th> Antibiotic</th> <th> Stock concentration</th> <th>Final concentration</th> <th>Dillution factor</th> <th>Solvent</th> <th>Storage temperature</th> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td>Ampicillin</td> <td>50 mg / mL</td> <td>50 μg / mL</td> <td>1000</td> <td>Filter sterilized H<sub>2</sub>O</td> <td> 4 °C</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td>Chloramphenicol</td><td>30 mg / mL</td><td>30 μg / mL</td> <td>1000</td><td>Ethanol</td><td>-20 °C</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td>Kanamycin</td><td>50 mg / mL</td><td>30 μg / mL</td><td>1000</td><td>Filter sterilized H<sub>2</sub>O</td><td>4 °C</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td>Spectinomycin</td><td>50 mg / mL</td><td>50 μg / mL</td><td>1000</td><td>Filter sterilized H<sub>2</sub>O</td><td>4 °C</td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | </div> | ||
</div> | </div> | ||
| - | <p> Ingredients: <br> <ul> <li>Tryptone (10g)</li> <li>NaCl (10g)</li> <li>Yeast Extract (5g)</li> </ul> <br> <p> <ol> <li>Fill with 1 L of distilled / filtered H<sub>2</sub>O.</li> <li>Autoclave at 121 °C for 20 minutes. </li> <li>Add antibiotics if needed, after the medium has cooled down.</li> </ol></p> | + | <p> |
| - | </p> | + | Ingredients: <br> <ul> <li>Tryptone (10g)</li> <li>NaCl (10g)</li> <li>Yeast Extract (5g)</li> </ul> <br> <p> <ol> <li>Fill with 1 L of distilled / filtered H<sub>2</sub>O.</li> <li>Autoclave at 121 °C for 20 minutes. </li> <li>Add antibiotics if needed, after the medium has cooled down.</li> |
| + | </ol></p> | ||
| + | </p> | ||
</div> | </div> | ||
</div> | </div> | ||
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</div> | </div> | ||
</div> | </div> | ||
| - | <div id=" | + | |
| + | <div id="week2entry" class="nb-week" style="display: block"> | ||
<div class="twelve columns"> | <div class="twelve columns"> | ||
| - | <h3 class="centered"> | + | <h3 class="centered">SOC medium</h3> |
| - | + | </p> | |
| + | <div class="entry pc-media"> | ||
| + | <div> | ||
| + | <h6>Recipe | ||
| + | <div class="nb-onetech-i nohilite">show technical details</div> | ||
| + | </h6> | ||
| + | <div class="nb-tech">{{{tech}}}</div> | ||
| + | <p> <p> Ingredients (1L): <br> <ul> <li>Bactotryptone (20 g)</li> <li>Yeast extract (5g)</li> <li>NaCl (0.584g)</li> <li>KCl (0.186g)</li> <li>Agar (20g ONLY IF MAKING PLATES)</li> </ul> <br> <p> <ol> <li>Fill with 0.5 L of distilled / filtered H<sub>2</sub>O.</li> <li>Autoclave at 121 °C for 20 minutes. </li> <li>Add 10 mL 1M MgCl<sub>2</sub>, 10 mL MgSO<sub>4</sub> and 20 mL 1M glucose (all should be sterile) prior to use </li> | ||
| + | </ol> | ||
| + | </p> | ||
| + | </p> | ||
| + | </div> | ||
| + | </div> | ||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | |||
| + | <div id="week3entry" class="nb-week" style="display: block"> | ||
| + | <div class="twelve columns"> | ||
| + | <h3 class="centered">yB medium</h3> | ||
</p> | </p> | ||
| + | <div class="entry pc-media"> | ||
| + | <div> | ||
| + | <h6>Recipe | ||
| + | |||
| + | </h6> | ||
| + | <div class="nb-tech">{{{tech}}}</div> | ||
| + | <p> Ingredients: <br> <ul> <li>Yeast extract (2.5g)</li> <li>Bactotryptone (10g)</li> <li>KCL (0.38g)</li> </ul> <br> <p> <ol> <li>Fill with 0.5 L of distilled / filtered H<sub>2</sub>O.</li> <li>Add KOH until the pH is 7.4, then autoclave at 121 °C for 20 minutes. </li> <li>Add 17 mL sterile 1M MgSO4 </li> | ||
| + | </ol> | ||
| + | </p> | ||
| + | </p> | ||
| + | </div> | ||
| + | </div> | ||
| + | </div> | ||
| + | </div> | ||
| + | <div id="week4entry" class="nb-week" style="display: block"> | ||
| + | <div class="twelve columns"> | ||
| + | <h3 class="centered">Synechocystis medium</h3> | ||
| + | </p> | ||
| + | <div class="entry pc-media"> | ||
| + | <div> | ||
| + | <h6>Recipe | ||
| + | |||
| + | </h6> | ||
| + | <div class="nb-tech">{{{tech}}}</div> | ||
| + | <p> To grow Synechocystis cultures, BG-11 medium was made according to the recipe found on the <a href="http://unitedscientists.org/labs/norway/NTNU/PhotoSynLab/wiki/table-of-contents-2/protocol-physiological/bg-11-media">PhotoSynLab wiki</a>. | ||
| + | </p> | ||
| + | </div> | ||
| + | </div> | ||
| + | </div> | ||
| + | </div> | ||
| + | <div id="week6entry" class="nb-week" style="display: block"> | ||
| + | <div class="twelve columns"> | ||
| + | <h3 class="centered">Gibson Assembly</h3> | ||
| + | <div class="entry pc-techniques"> | ||
| + | <div> | ||
| + | <h6> | ||
| + | </h6> | ||
| + | <div class="nb-tech">{{{tech}}}</div> | ||
| + | <p> | ||
| + | The Gibson assembly protocol can be found at <a href="https://www.neb.com/protocols/2012/12/11/gibson-assembly-protocol-e5510">New England Biolabs</a>. | ||
| + | </p> | ||
| + | </div> | ||
| + | </div> | ||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | |||
| + | |||
| + | <div id="week7entry" class="nb-week" style="display: block"> | ||
| + | <div class="twelve columns"> | ||
| + | <h3 class="centered">DNA isolation and cleaning</h3> | ||
| + | </p> | ||
| + | <div class="entry pc-techniques"> | ||
| + | <div> | ||
| + | |||
| + | |||
| + | |||
| + | <div class="nb-tech">{{{tech}}}</div> | ||
| + | <p> For plasmid isolation, the Promega Wizard Plus SV Minipreps DNA Purification System A1460 Miniprep protocol was used. For PCR product isolation, the QIAquick PCR Purification kit was used. | ||
| + | </p> | ||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | <div id="week8entry" class="nb-week" style="display: block"> | ||
| + | <div class="twelve columns"> | ||
| + | <h3 class="centered">DNA digestion</h3> | ||
| + | </p> | ||
| + | <div class="entry pc-techniques"> | ||
| + | <div> | ||
| + | |||
| + | |||
| + | |||
| + | <div class="nb-tech">{{{tech}}}</div> | ||
| + | <p> | ||
| + | |||
| + | DNA digests for both ligation and verification used the protocol in the <a href="http://unitedscientists.org/labs/norway/NTNU/PhotoSynLab/wiki/table-of-contents-2/protocol-molecular/dna-ligation/restriction-enzyme-digest">PhotoSynLab wiki</a>. | ||
| + | |||
| + | |||
| + | |||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | |||
| + | <div id="week9entry" class="nb-week" style="display: block"> | ||
| + | <div class="twelve columns"> | ||
| + | <h3 class="centered">PCR</h3> | ||
| + | </p> | ||
| + | <div class="entry pc-techniques"> | ||
| + | <div> | ||
| + | |||
| + | <div class="nb-tech">{{{tech}}}</div> | ||
| + | <p> The touchdown PCR procedure detailed on the <a href="http://unitedscientists.org/labs/norway/NTNU/PhotoSynLab/wiki/table-of-contents-2/protocol-molecular/pcr-touchdown-pcr">PhotoSynLab wiki</a> was used to amplify DNA. | ||
| + | </p> | ||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | |||
| + | <div id="week10entry" class="nb-week" style="display: block"> | ||
| + | <div class="twelve columns"> | ||
| + | <h3 class="centered">Nanodrop</h3> | ||
| + | </p> | ||
| + | <div class="entry pc-techniques"> | ||
| + | <div> | ||
| + | |||
| + | <div class="nb-tech">{{{tech}}}</div> | ||
| + | <p> <a href="http://www.nanodrop.com/library/nd-1000-v3.7-users-manual-8.5x11.pdf">A NanoDrop ND-1000 Spectrophotometer</a> was used to determine DNA concentrations. | ||
| + | </p> | ||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | <div id="week11entry" class="nb-week" style="display: block"> | ||
| + | <div class="twelve columns"> | ||
| + | <h3 class="centered">3A assembly</h3> | ||
| + | </p> | ||
| + | <div class="entry pc-techniques"> | ||
| + | <div> | ||
| + | |||
| + | <div class="nb-tech">{{{tech}}}</div> | ||
| + | <p> 3A assembly was conducted according to the <a href="http://parts.igem.org/Help:Protocol/3A_Assembly">iGEM 3A assembly protocol</a>. | ||
| + | </p> | ||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | |||
| + | <div id="week12entry" class="nb-week" style="display: block"> | ||
| + | <div class="twelve columns"> | ||
| + | <h3 class="centered">Ligation</h3> | ||
| + | |||
| + | <div class="entry pc-techniques"> | ||
| + | <div> | ||
| + | |||
| + | <div class="nb-tech">{{{tech}}}</div> | ||
| + | <p> Ligation was performed according to the protocol outlined on the <a href="http://unitedscientists.org/labs/norway/NTNU/PhotoSynLab/wiki/table-of-contents-2/protocol-molecular/pcr-touchdown-pcr">PhotoSynLab wiki</a>. | ||
| + | </p> | ||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | |||
| + | <div id="week13entry" class="nb-week" style="display: block"> | ||
| + | <div class="twelve columns"> | ||
| + | <h3 class="centered">Transformation (Escherichia coli)</h3> | ||
| + | </p> | ||
| + | <div class="entry pc-techniques"> | ||
| + | <div> | ||
| + | |||
| + | <div class="nb-tech">{{{tech}}}</div> | ||
| + | <p> The protocols used for preparing competent DH5a cells, as well as the heat-shock transformation procedure employed can be found at the <a href="http://unitedscientists.org/labs/norway/NTNU/PhotoSynLab/wiki/table-of-contents-2/protocol-molecular/heat-shock-transformation-2">PhotoSynLab wiki</a>. | ||
| + | </p> | ||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | <div id="week14entry" class="nb-week" style="display: block"> | ||
| + | <div class="twelve columns"> | ||
| + | <h3 class="centered">Transformation (Synechocystis sp. PCC 6803)</h3> | ||
| + | </p> | ||
| + | <div class="entry pc-techniques"> | ||
| + | <div> | ||
| + | |||
| + | <div class="nb-tech">{{{tech}}}</div> | ||
| + | <p> The transformation procedure for transforming Synechocystis can be found at the <a href="http://unitedscientists.org/labs/norway/NTNU/PhotoSynLab/wiki/table-of-contents-2/protocol-molecular/synechocystis-transformation-r-hill-method">PhotoSynLab wiki</a>. | ||
| + | </p> | ||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | <div id="week16entry" class="nb-week" style="display: block"> | ||
| + | <div class="twelve columns"> | ||
| + | <h3 class="centered">Right flank</h3> | ||
| + | |||
| + | <div class="entry pc-plas"> | ||
| + | <div> | ||
| + | |||
| + | <div class="nb-tech">{{{tech}}}</div> | ||
| + | <p> <a href="http://parts.igem.org/Part:BBa_K1424001">BBa_K1424001</a>. | ||
| + | </p> | ||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | <div id="week17entry" class="nb-week" style="display: block"> | ||
| + | <div class="twelve columns"> | ||
| + | <h3 class="centered">Left flank</h3> | ||
| + | |||
| + | <div class="entry pc-plas"> | ||
| + | <div> | ||
| + | |||
| + | <div class="nb-tech">{{{tech}}}</div> | ||
| + | <p> <a href="http://parts.igem.org/Part:BBa_K1424000">BBa_K1424000</a>. | ||
| + | </p> | ||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | <div id="week18entry" class="nb-week" style="display: block"> | ||
| + | <div class="twelve columns"> | ||
| + | <h3 class="centered">Kanamycin resistance</h3> | ||
| + | |||
| + | <div class="entry pc-plas"> | ||
| + | <div> | ||
| + | |||
| + | <div class="nb-tech">{{{tech}}}</div> | ||
| + | <p> <a href="http://parts.igem.org/Part:BBa_K1424003">BBa_K1424003</a>. | ||
| + | </p> | ||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | |||
| + | <div id="week19entry" class="nb-week" style="display: block"> | ||
| + | <div class="twelve columns"> | ||
| + | <h3 class="centered">Lac promotor</h3> | ||
| + | |||
| + | <div class="entry pc-plas"> | ||
| + | <div> | ||
| + | |||
| + | <div class="nb-tech">{{{tech}}}</div> | ||
| + | <p> <a href="http://parts.igem.org/Part:BBa_K1424002">BBa_K1424002</a>. | ||
| + | </p> | ||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | <div id="week20entry" class="nb-week" style="display: block"> | ||
| + | <div class="twelve columns"> | ||
| + | <h3 class="centered">Glucose oxidase</h3> | ||
| + | |||
| + | <div class="entry pc-plas"> | ||
| + | <div> | ||
| + | |||
| + | <div class="nb-tech">{{{tech}}}</div> | ||
| + | <p> <a href="http://parts.igem.org/Part:BBa_K1424004">BBa_K1424004</a>. | ||
| + | </p> | ||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | <div id="week21entry" class="nb-week" style="display: block"> | ||
| + | <div class="twelve columns"> | ||
| + | <h3 class="centered">Left flank + Kanamycin resistance + Right flank</h3> | ||
| + | |||
| + | <div class="entry pc-plas"> | ||
| + | <div> | ||
| + | |||
| + | <div class="nb-tech">{{{tech}}}</div> | ||
| + | <p> <a href="http://parts.igem.org/Part:BBa_K1424005">BBa_K1424005</a>. | ||
| + | </p> | ||
| + | </div> | ||
| + | </div> | ||
| + | |||
| + | </div> | ||
| + | </div> | ||
| - | |||
| - | |||
| - | |||
<div id="week24entry" class="nb-week" style="display: block"> | <div id="week24entry" class="nb-week" style="display: block"> | ||
<div class="twelve columns"> | <div class="twelve columns"> | ||
| - | <h3 class="centered"> | + | <h3 class="centered">DNA concentration</h3> |
| - | <div class="entry pc- | + | <div class="entry pc-calc"> |
<div> | <div> | ||
| - | + | ||
| - | + | ||
| - | + | ||
<div class="nb-tech">{{{tech}}}</div> | <div class="nb-tech">{{{tech}}}</div> | ||
| - | <p> | + | <p> After measuring the concentration of a sample of DNA in ng/ul with the NanoDrop method, the concentration in terms of pmolar could be determined with the equation |
| + | <img src="https://static.igem.org/mediawiki/2014/c/c5/Eqn3.gif"> | ||
</p> | </p> | ||
</div> | </div> | ||
| Line 648: | Line 887: | ||
</div> | </div> | ||
</div> | </div> | ||
| + | |||
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<div class="printfooter"> | <div class="printfooter"> | ||
| - | Retrieved from "<a href="https://2014.igem.org/Team:NTNU_Trondheim/ | + | Retrieved from "<a href="https://2014.igem.org/Team:NTNU_Trondheim/Protocols">https://2014.igem.org/Team:NTNU_Trondheim/Protocols</a>"</div> |
<div id="catlinks"><div id='catlinks' class='catlinks catlinks-allhidden'></div></div> <!-- end content --> | <div id="catlinks"><div id='catlinks' class='catlinks catlinks-allhidden'></div></div> <!-- end content --> | ||
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title='Recent changes'>Recent changes</a></li> | title='Recent changes'>Recent changes</a></li> | ||
| - | <li id="t-whatlinkshere"><a href="/Special:WhatLinksHere/Team:NTNU_Trondheim/ | + | <li id="t-whatlinkshere"><a href="/Special:WhatLinksHere/Team:NTNU_Trondheim/Protocols" |
title="List of all wiki pages that link here [j]" accesskey="j">What links here</a></li> | title="List of all wiki pages that link here [j]" accesskey="j">What links here</a></li> | ||
| - | <li id="t-recentchangeslinked"><a href="/Special:RecentChangesLinked/Team:NTNU_Trondheim/ | + | <li id="t-recentchangeslinked"><a href="/Special:RecentChangesLinked/Team:NTNU_Trondheim/Protocols" |
title="Recent changes in pages linked from this page [k]" accesskey="k">Related changes</a></li> | title="Recent changes in pages linked from this page [k]" accesskey="k">Related changes</a></li> | ||
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<ul id="f-list"> | <ul id="f-list"> | ||
| - | <li id="t-print"><a href="/wiki/index.php?title=Team:NTNU_Trondheim/ | + | <li id="t-print"><a href="/wiki/index.php?title=Team:NTNU_Trondheim/Protocols&printable=yes" |
title="Printable version of this page [p]" accesskey="p">Printable version</a> | title="Printable version of this page [p]" accesskey="p">Printable version</a> | ||
</li> | </li> | ||
| - | <li id="t-permalink"><a href="/wiki/index.php?title=Team:NTNU_Trondheim/ | + | <li id="t-permalink"><a href="/wiki/index.php?title=Team:NTNU_Trondheim/Protocols&oldid=57997" |
title="Permanent link to this revision of the page">Permanent link</a> | title="Permanent link to this revision of the page">Permanent link</a> | ||
</li> | </li> | ||
Latest revision as of 21:39, 17 October 2014
Team:NTNU Trondheim/Protocols
From 2014.igem.org
"
Team:NTNU_Trondheim/Protocols
From 2014.igem.org
Jump to:
-
Media
- Lysogeny Broth (LB)
- SOC medium
- yB medium
- Synechocystis medium
-
Techniques
- Gibson assembly
- DNA isolation and cleaning
- DNA digestion
- PCR
- NanoDrop
- 3A assembly
- Ligation
- Transformation (Escherichia coli)
- Transformation (Synechocystis sp. PCC 6803)
-
Plasmids
- Right flank
- Left flank
- Kanamycin resistance
- Lac promoter
- Glucose oxidase
- Left flank + Kan + Right flank
-
Calculations
- DNA concentration
Protocols
Filter by subteam:
show all categories
show technical details
"_"
Media
Techniques
Plasmids
Calculations
only
only
only
only
Lysogeny Broth (LB)
Recipe
Antibiotic additions
| Antibiotic | Stock concentration | Final concentration | Dillution factor | Solvent | Storage temperature |
|---|---|---|---|---|---|
| Ampicillin | 50 mg / mL | 50 μg / mL | 1000 | Filter sterilized H2O | 4 °C |
| Chloramphenicol | 30 mg / mL | 30 μg / mL | 1000 | Ethanol | -20 °C |
| Kanamycin | 50 mg / mL | 30 μg / mL | 1000 | Filter sterilized H2O | 4 °C |
| Spectinomycin | 50 mg / mL | 50 μg / mL | 1000 | Filter sterilized H2O | 4 °C |
Ingredients:
- Tryptone (10g)
- NaCl (10g)
- Yeast Extract (5g)
- Fill with 1 L of distilled / filtered H2O.
- Autoclave at 121 °C for 20 minutes.
- Add antibiotics if needed, after the medium has cooled down.
SOC medium
Recipe
show technical details
{{{tech}}}
Ingredients (1L):
- Bactotryptone (20 g)
- Yeast extract (5g)
- NaCl (0.584g)
- KCl (0.186g)
- Agar (20g ONLY IF MAKING PLATES)
- Fill with 0.5 L of distilled / filtered H2O.
- Autoclave at 121 °C for 20 minutes.
- Add 10 mL 1M MgCl2, 10 mL MgSO4 and 20 mL 1M glucose (all should be sterile) prior to use
yB medium
Recipe
{{{tech}}}
Ingredients:
- Yeast extract (2.5g)
- Bactotryptone (10g)
- KCL (0.38g)
- Fill with 0.5 L of distilled / filtered H2O.
- Add KOH until the pH is 7.4, then autoclave at 121 °C for 20 minutes.
- Add 17 mL sterile 1M MgSO4
Synechocystis medium
Recipe
{{{tech}}}
To grow Synechocystis cultures, BG-11 medium was made according to the recipe found on the PhotoSynLab wiki.
Gibson Assembly
DNA isolation and cleaning
{{{tech}}}
For plasmid isolation, the Promega Wizard Plus SV Minipreps DNA Purification System A1460 Miniprep protocol was used. For PCR product isolation, the QIAquick PCR Purification kit was used.
DNA digestion
{{{tech}}}
DNA digests for both ligation and verification used the protocol in the PhotoSynLab wiki.
PCR
{{{tech}}}
The touchdown PCR procedure detailed on the PhotoSynLab wiki was used to amplify DNA.
Nanodrop
{{{tech}}}
A NanoDrop ND-1000 Spectrophotometer was used to determine DNA concentrations.
3A assembly
{{{tech}}}
3A assembly was conducted according to the iGEM 3A assembly protocol.
Ligation
{{{tech}}}
Ligation was performed according to the protocol outlined on the PhotoSynLab wiki.
Transformation (Escherichia coli)
{{{tech}}}
The protocols used for preparing competent DH5a cells, as well as the heat-shock transformation procedure employed can be found at the PhotoSynLab wiki.
Transformation (Synechocystis sp. PCC 6803)
{{{tech}}}
The transformation procedure for transforming Synechocystis can be found at the PhotoSynLab wiki.
Right flank
{{{tech}}}
Left flank
{{{tech}}}
Kanamycin resistance
{{{tech}}}
Lac promotor
{{{tech}}}
Glucose oxidase
{{{tech}}}
Left flank + Kanamycin resistance + Right flank
{{{tech}}}
DNA concentration
{{{tech}}}
After measuring the concentration of a sample of DNA in ng/ul with the NanoDrop method, the concentration in terms of pmolar could be determined with the equation
