Difference between revisions of "Team:Paris Bettencourt/Notebook/Riboswitch"
Line 1,085: | Line 1,085: | ||
Change my restriction enzymes<br> | Change my restriction enzymes<br> | ||
Use DpnI<br> | Use DpnI<br> | ||
+ | |||
+ | <br><br> | ||
+ | <h1 class="date two">August 17th</h1> | ||
+ | |||
+ | <h2>Goal</h2> | ||
+ | Miniprep and digest colonie 6 and 5 for control | ||
+ | |||
+ | <h2>Procedure</h2> | ||
+ | Using Promega SV Miniprep Kit <br> | ||
+ | Nanodrop after<br> | ||
+ | 6 = 55 ng/uL<br> | ||
+ | 5 = 51 ng/uL<br> | ||
+ | Digestion : Hind III enzyme (1 digestion site in my construction, inside my insert) | ||
+ | <style type="text/css"> | ||
+ | .tg {border-collapse:collapse;border-spacing:0;} | ||
+ | .tg td{font-family:Arial, sans-serif;font-size:14px;padding:10px 5px;border-style:solid;border-width:1px;overflow:hidden;word-break:normal;} | ||
+ | .tg th{font-family:Arial, sans-serif;font-size:14px;font-weight:normal;padding:10px 5px;border-style:solid;border-width:1px;overflow:hidden;word-break:normal;} | ||
+ | </style> | ||
+ | <table class="tg"> | ||
+ | <tr> | ||
+ | <th class="tg-031e"></th> | ||
+ | <th class="tg-031e">5</th> | ||
+ | <th class="tg-031e">6</th> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">Hind III</td> | ||
+ | <td class="tg-031e">4 uL</td> | ||
+ | <td class="tg-031e">4 uL</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">FastAP</td> | ||
+ | <td class="tg-031e">4 uL</td> | ||
+ | <td class="tg-031e">4 uL</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">FD buffer</td> | ||
+ | <td class="tg-031e">8 uL</td> | ||
+ | <td class="tg-031e">8 uL</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">DNA (50ng/uL)</td> | ||
+ | <td class="tg-031e">20 uL</td> | ||
+ | <td class="tg-031e">20 uL</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">Water</td> | ||
+ | <td class="tg-031e">44 uL</td> | ||
+ | <td class="tg-031e">44 uL</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">TOTAL</td> | ||
+ | <td class="tg-031e">80 uL</td> | ||
+ | <td class="tg-031e">80 uL</td> | ||
+ | </tr> | ||
+ | </table> | ||
+ | Mix, Incubate 30 min 37°C | ||
+ | PCR purification | ||
+ | |||
+ | <br><br> | ||
+ | <h1 class="date two">August 18th</h1> | ||
+ | |||
+ | <h2>Goal1</h2> | ||
+ | New digestion | ||
+ | |||
+ | <h2>Procedure</h2> | ||
+ | |||
+ | <style type="text/css"> | ||
+ | .tg {border-collapse:collapse;border-spacing:0;} | ||
+ | .tg td{font-family:Arial, sans-serif;font-size:14px;padding:10px 5px;border-style:solid;border-width:1px;overflow:hidden;word-break:normal;} | ||
+ | .tg th{font-family:Arial, sans-serif;font-size:14px;font-weight:normal;padding:10px 5px;border-style:solid;border-width:1px;overflow:hidden;word-break:normal;} | ||
+ | </style> | ||
+ | <table class="tg"> | ||
+ | <tr> | ||
+ | <th class="tg-031e"></th> | ||
+ | <th class="tg-031e">F2d'</th> | ||
+ | <th class="tg-031e">F3d'</th> | ||
+ | <th class="tg-031e">LP1d'</th> | ||
+ | <th class="tg-031e">LP3d'</th> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">DNA</td> | ||
+ | <td class="tg-031e">55ng/uL = 20uL</td> | ||
+ | <td class="tg-031e">76 ng/uL = 15uL</td> | ||
+ | <td class="tg-031e">76 ng/uL = 15uL</td> | ||
+ | <td class="tg-031e">82ng/uL = 15uL</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">FastAP</td> | ||
+ | <td class="tg-031e"></td> | ||
+ | <td class="tg-031e"></td> | ||
+ | <td class="tg-031e">2uL</td> | ||
+ | <td class="tg-031e">2uL</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">EcoRI</td> | ||
+ | <td class="tg-031e">2uL</td> | ||
+ | <td class="tg-031e"></td> | ||
+ | <td class="tg-031e">2uL</td> | ||
+ | <td class="tg-031e"></td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">XbaI</td> | ||
+ | <td class="tg-031e"></td> | ||
+ | <td class="tg-031e">2uL</td> | ||
+ | <td class="tg-031e"></td> | ||
+ | <td class="tg-031e">2uL</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">SpeI</td> | ||
+ | <td class="tg-031e">2uL</td> | ||
+ | <td class="tg-031e">2uL</td> | ||
+ | <td class="tg-031e">2uL</td> | ||
+ | <td class="tg-031e">2uL</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">FD buffer</td> | ||
+ | <td class="tg-031e">4uL</td> | ||
+ | <td class="tg-031e">4uL</td> | ||
+ | <td class="tg-031e">6uL</td> | ||
+ | <td class="tg-031e">6uL</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">Water</td> | ||
+ | <td class="tg-031e">12uL</td> | ||
+ | <td class="tg-031e">17uL</td> | ||
+ | <td class="tg-031e">33uL</td> | ||
+ | <td class="tg-031e">33uL</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">TOTAL</td> | ||
+ | <td class="tg-031e">40uL</td> | ||
+ | <td class="tg-031e">40uL</td> | ||
+ | <td class="tg-031e">60uL</td> | ||
+ | <td class="tg-031e">60uL</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">Nanodrop after purify</td> | ||
+ | <td class="tg-031e">41 ng/uL</td> | ||
+ | <td class="tg-031e">21 ng/uL</td> | ||
+ | <td class="tg-031e">16 ng/uL</td> | ||
+ | <td class="tg-031e">68 ng/uL</td> | ||
+ | </tr> | ||
+ | </table> | ||
+ | 20 min at 37°C | ||
+ | |||
+ | <h2>Goal2</h2> | ||
+ | Ligation | ||
+ | <h2>Procedure</h2> |
Revision as of 10:31, 16 September 2015