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Team:Washington/Protocols/Transformation
From 2011.igem.org
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#Spread 100-300 μl onto a plate made with appropriate antibiotic. | #Spread 100-300 μl onto a plate made with appropriate antibiotic. | ||
#Grow overnight at 37°C. | #Grow overnight at 37°C. | ||
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| + | Adapted from [http://openwetware.org/wiki/Transforming_chemically_competent_cells_%28Inoue%29 OpenWetWare protocol]. | ||
Revision as of 01:35, 14 September 2011
General Transformation Protocol
- Thaw 25 - 200 μl TB buffer cells on ice. Do not use glass tubes, which adsorb DNA.
- Add DNA, pipette gently to mix (keep volume of DNA less than 5% of the cell volume)
- Incubate on ice for 30 minutes
- Note: If you are in a rush, you can shorten this incubation time to 5-10 min.
- Incubate cells for 30 seconds at 42°C.
- Incubate cells on ice for 2 min.
- Add 4 volumes of room temperature SOC (not critical)
- Incubate for 1 hour at 37°C on shaker.
- Note: Can also save some time here by reducing incubation to ~45 min.
- Note: Step can be eliminated if plating on Amp plates, but not most other antibiotics
- Spread 100-300 μl onto a plate made with appropriate antibiotic.
- Grow overnight at 37°C.
Adapted from [http://openwetware.org/wiki/Transforming_chemically_competent_cells_%28Inoue%29 OpenWetWare protocol].
