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Team:British Columbia/Notebook/Protocols/transformation
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Revision as of 01:06, 18 October 2014
Transformation
Supplies:
- Competent cells in 100μL aliquots.
- Water bath/heat block at 42°C.
- Ice
- LB Broth
- LB agar plates (with appropriate selection)
Steps:
- Remove competent cells (100uL aliquots) from -80°C and thaw on ice.
- Note, aliquots can be thawed GENTLY by hand if time is an issue*
- Add 1µL ligation mix to thawed cells and incubate for 30 minutes on ice.
- Heat shock in water bath/heat block for 60 seconds and put back on ice for two minutes.
- Remember to turn off the water bath.
- The following steps must be done near a flame.
- Don’t forget a growth control, especially if you aren’t screening using antibiotics.
- Add 400uL LB Broth and incubate at 37C for 2 hours.
- Note, 1 hour is also sufficient
- Spread plate entire 500µL.
- Turn off the flame
- Incubate overnight (<18hrs) at 37°C.
