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Team:Calgary/Notebook/Protocols/transformation
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<li> Incubate for 30 to 60 minutes with shaking at 37°C. (Note that for Kanamycin containing plasmids always use one hour)</li> | <li> Incubate for 30 to 60 minutes with shaking at 37°C. (Note that for Kanamycin containing plasmids always use one hour)</li> | ||
<li> Spin down to remove all supernatant except approximately 100 μL</li> | <li> Spin down to remove all supernatant except approximately 100 μL</li> | ||
| - | <li> Plate approximately | + | <li> Plate approximately 50 μL on each of two antibiotic plates </li> |
<li> <a href="https://2012.igem.org/Team:Calgary/Notebook/Protocols/onculture">Grow overnight</a> at 37°C </li> | <li> <a href="https://2012.igem.org/Team:Calgary/Notebook/Protocols/onculture">Grow overnight</a> at 37°C </li> | ||
</ol> | </ol> | ||
Latest revision as of 03:29, 4 October 2012
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Bacterial Transformation
- Thaw 100 μL of competent cells (per transformation) on ice just before they are needed
- Add DNA (max 20μl) thawed cells and mix by flicking the side of the tube. Leave on ice for 30 minutes
- Heat shock 5 minutes at 37 degrees Celsius
- Place on ice for 5 minutes
- Add 250ul SOC medium to each tube
- Incubate for 30 to 60 minutes with shaking at 37°C. (Note that for Kanamycin containing plasmids always use one hour)
- Spin down to remove all supernatant except approximately 100 μL
- Plate approximately 50 μL on each of two antibiotic plates
- Grow overnight at 37°C
For this protocol we used a couple of controls
- Positive Control - pBluescript in TOP10 cells on ampicillin plates
- Negative Control - TOP10 cells grown on ampcillin plates
