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Team:NYU Gallatin/Project/Cloning/Protocols/Transformation
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Cloning: Transformation Protocol
- Thaw competent cells on ice. Not in hands.
- Transfer 50 µl of competent cells to 1.5 ml microcentrifuge tube. The competent cells are fragile and sensitive to temperature change, cool the microcentrifuge in the ice for a few seconds first, then be as quick as possible while transferring the cells and return them to the ice.
- Add 2 µl of the chilled assembly product to competent cells. Mix gently by pipetting up and down or folcking the tube 4-5 times. Do not vortex.
- Place the mixture on ice for at least 30 minutes. Do not mix.
- Heat shock at 42° C for exactly 30 seconds. Do not mix.
- Add 950 µl of room temperature SOC media to the tube.
- Place the tube at 37° C for 60 minutes. Shake vigorously (250 rpm) or rotate.
- Warm selection plates to 37° C.
- Spread 100 µl of the cells onto the plates.
- Incubate overnight at 37° C.
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