Difference between revisions of "Team:Paris Bettencourt/Notebook/Riboswitch"
Line 1,432: | Line 1,432: | ||
PS : 23 is two bands now maybe because the digestion is now finished (it was on the bench for one hour)<br> | PS : 23 is two bands now maybe because the digestion is now finished (it was on the bench for one hour)<br> | ||
23 sent to sequencing | 23 sent to sequencing | ||
+ | |||
+ | <br><br> | ||
+ | <h1 class="date two">August 31th</h1> | ||
+ | |||
+ | <h2>Goal 1</h2> | ||
+ | PCR for my Gibson 1 = Replace the RBS and first codon of the RFP in the classical pSB1C3 associated with Part:BBa J04450 by the riboswitch sequence | ||
+ | <h2>Procedure</h2> | ||
+ | PCR | ||
+ | <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">Insert</th> | ||
+ | <th class="tg-031e"></th> | ||
+ | <th class="tg-031e">Backbone</th> | ||
+ | <th class="tg-031e"><br></th> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">Template</td> | ||
+ | <td class="tg-031e">1ng = 2uL</td> | ||
+ | <td class="tg-031e">1 uL</td> | ||
+ | <td class="tg-031e">1ng = 2uL</td> | ||
+ | <td class="tg-031e">1uL</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">Primers</td> | ||
+ | <td class="tg-031e">209-210<br>1uL</td> | ||
+ | <td class="tg-031e"></td> | ||
+ | <td class="tg-031e">210-211<br>1uL</td> | ||
+ | <td class="tg-031e"></td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">MasterMix</td> | ||
+ | <td class="tg-031e">25 uL</td> | ||
+ | <td class="tg-031e"></td> | ||
+ | <td class="tg-031e">25 uL</td> | ||
+ | <td class="tg-031e"></td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">Water</td> | ||
+ | <td class="tg-031e">23 uL</td> | ||
+ | <td class="tg-031e">24 uL</td> | ||
+ | <td class="tg-031e">23 uL</td> | ||
+ | <td class="tg-031e">24 uL</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">TOTAL</td> | ||
+ | <td class="tg-031e">50uL</td> | ||
+ | <td class="tg-031e"></td> | ||
+ | <td class="tg-031e">50uL</td> | ||
+ | <td class="tg-031e"></td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">Elongation time</td> | ||
+ | <td class="tg-031e">30 sec</td> | ||
+ | <td class="tg-031e"></td> | ||
+ | <td class="tg-031e">2 min</td> | ||
+ | <td class="tg-031e"></td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">Expected length</td> | ||
+ | <td class="tg-031e">243</td> | ||
+ | <td class="tg-031e"></td> | ||
+ | <td class="tg-031e">3149</td> | ||
+ | <td class="tg-031e"></td> | ||
+ | </tr> | ||
+ | </table> | ||
+ | <h2>Results</h2> | ||
+ | Gel order : GR 1kb, B1, B2, I1, I2<br> | ||
+ | <img src="https://static.igem.org/mediawiki/2015/3/37/8778.png" width="350px"><br> | ||
+ | Bad result for B1 , B2 | ||
+ | |||
+ | <h2>Goal 2</h2> | ||
+ | New PCR for the backbone | ||
+ | <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">B2</th> | ||
+ | <th class="tg-031e">B3</th> | ||
+ | <th class="tg-031e">B4</th> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">Template</td> | ||
+ | <td class="tg-031e">3uL (ng)</td> | ||
+ | <td class="tg-031e">3uL (ng)</td> | ||
+ | <td class="tg-031e">3uL (ng)</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">MasterMixPhusion</td> | ||
+ | <td class="tg-031e">25 uL</td> | ||
+ | <td class="tg-031e">25 uL</td> | ||
+ | <td class="tg-031e">25 uL</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">DMSO</td> | ||
+ | <td class="tg-031e">1uL</td> | ||
+ | <td class="tg-031e">1uL</td> | ||
+ | <td class="tg-031e">1uL</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">Primers</td> | ||
+ | <td class="tg-031e">1uL</td> | ||
+ | <td class="tg-031e">1uL</td> | ||
+ | <td class="tg-031e">1uL</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">Water</td> | ||
+ | <td class="tg-031e">20 uL</td> | ||
+ | <td class="tg-031e">20 uL</td> | ||
+ | <td class="tg-031e">20 uL</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td class="tg-031e">T°C annealing</td> | ||
+ | <td class="tg-031e">56°C</td> | ||
+ | <td class="tg-031e">58°C</td> | ||
+ | <td class="tg-031e">60°C</td> | ||
+ | </tr> | ||
+ | </table> | ||
+ | <h2>Result</h2> | ||
+ | <img src="https://static.igem.org/mediawiki/2015/1/11/7665.png" width="350px"><br> |
Revision as of 12:56, 16 September 2015