Oct 18, 2020
Bacterial Transformation
- 1South China University of Technology
Protocol Citation: Jiaxin Li 2020. Bacterial Transformation. protocols.io https://dx.doi.org/10.17504/protocols.io.bnifmcbn
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: October 16, 2020
Last Modified: October 18, 2020
Protocol Integer ID: 43303
Thaw a tube of DH5a Competent E. coli cells on ice for 5-10 minutes
Add 1µl of plasmid DNA to per 100µl cell mixture. Flick the tube 4-5 times to mix cells and DNA but do not vortex.
Place the mixture on ice for 30 minutes. Do not mix.
Heat shock at exactly 42°C for exactly 90 seconds. Immediately place on ice for 2-3 minutes. Do not mix.
Pipette 800 µl of 42°C LB liquid medium into the mixture.
Place at 37°C, 180-220 rpm for 45-60 minutes.
Centrifuge the bacterium at 3000r for 3 minutes.
Discard 300 μl supernatant and resuspend the sediment.
Spread 50-100 µl of each dilution onto a selection plate and incubate for 12 hours at 37°C.