"
Team:Evry/Notebook/Protocols/Transformation
From 2014.igem.org
(Difference between revisions)
| (6 intermediate revisions not shown) | |||
| Line 2: | Line 2: | ||
<div align="center"> | <div align="center"> | ||
| - | <FONT COLOR="blue"> | + | <FONT COLOR="blue"> |
| - | <h4>Transformation of <i>Pseudovibrio<i></h4> | + | <h4>Transformation of <i>Pseudovibrio</i></h4> |
| - | </FONT> | + | </FONT> |
</div> | </div> | ||
<div align="left"> | <div align="left"> | ||
| - | |||
| - | <li>Place electroporation tanks in ice for 10min< | + | <ul> |
| + | <li>Place electroporation tanks in ice for 10min</li> | ||
| + | <li>Take samples of plasmids (25ng-50ng/µL) /!\ Keep them in ice /!\ </li> | ||
| + | <li>Take sample of competent cells /!\ Keep them in ice /!\ </li> | ||
| + | <li>Place 1µL of plasmid in the sample of competent cells</li> | ||
| + | <li>Transfer the full volume obtained in the electroporation tank</li> | ||
| + | <li>Place in the electroporator and pulse at 2000V </li> | ||
| + | <b>NB:</b> The optimal pulse length is between 5 and 6ms.</li> | ||
| + | <li>Add 1mL of MB 1X in the 30 seconds following the transformation</li> | ||
| + | <li>Incubate between 2h and 3h at 30°C with shaking</li> | ||
| + | <li>Centrifuge to concentrate all cells in the pellet</li> | ||
| + | <li>Discard the supernatant</li> | ||
| + | <li>Sowed the pellet on selective plates of MB 1X </li> | ||
<br> | <br> | ||
| - | < | + | </ul> |
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
</div> | </div> | ||
| - | </ | + | </html> |
Latest revision as of 00:48, 18 October 2014
Transformation of Pseudovibrio
- Place electroporation tanks in ice for 10min
- Take samples of plasmids (25ng-50ng/µL) /!\ Keep them in ice /!\
- Take sample of competent cells /!\ Keep them in ice /!\
- Place 1µL of plasmid in the sample of competent cells
- Transfer the full volume obtained in the electroporation tank
- Place in the electroporator and pulse at 2000V NB: The optimal pulse length is between 5 and 6ms.
- Add 1mL of MB 1X in the 30 seconds following the transformation
- Incubate between 2h and 3h at 30°C with shaking
- Centrifuge to concentrate all cells in the pellet
- Discard the supernatant
- Sowed the pellet on selective plates of MB 1X
