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Team:Grenoble/Biology/Protocols/Transformation 2
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<li>Thaw TSS cells on ice.</li> | <li>Thaw TSS cells on ice.</li> | ||
<li>Add DNA, pipette gently to mix (1 µL of prepped plasmid is more than enough).</li> | <li>Add DNA, pipette gently to mix (1 µL of prepped plasmid is more than enough).</li> | ||
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resuspend in enough medium to spread on one plate and plate it all. | resuspend in enough medium to spread on one plate and plate it all. | ||
This way you will find even small numbers of transformants.</li> | This way you will find even small numbers of transformants.</li> | ||
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Revision as of 10:15, 24 August 2012
Transformation of E. coli competent cells (heat shock)
Goal
Transform E. coli competent cells whith foreign DNA.Protocol
Following are the instructions from the OpenWetWare website.- Thaw TSS cells on ice.
- Add DNA, pipette gently to mix (1 µL of prepped plasmid is more than enough).
- Let sit for 30 minutes on ice.
- Incubate cells for 30 seconds at 42°C.
- Incubate cells on ice for 2 min.
- Add 1 mL of LB medium at room temperature.
- Incubate for 1 hour at 37°C on shaker.
- Spread 100-300 µL onto a plate made with appropriate antibiotic.
- Grow overnight at 37°C.
- Save the rest of the transformants in liquid culture at 4°C. If nothing appears on your plate, you can spin this down, resuspend in enough medium to spread on one plate and plate it all. This way you will find even small numbers of transformants.
