- Grow cells in ~4 mL LB until cloudy (OD600=0.5)
- Put on ice
- Transfer 1mL into an eppendorf tube on ice, let cool
- Centrifuge full speed for 30 sec, toss out supernatant
- Resuspend in 90uL of TSS solution
- Add 10uL KCM
Step 6.5: do the negative control before adding the DNA
- Add 1uL plasmid DNA
- Let sit on ice for 10min, heat shock 90 sec at 42, ice for a minute, rescue 1 hr, then incubate and/or plate
- Always do negative controls on your competent cell prep!
Sector a region of your plate and spread about 5uL of untransformed competent cells to confirm that they are free of contamination.