Quick Competent E.coli/Transformation

Gregory Broussard

Revised: 09/22/08

(Please use, change and disseminate freely!)

This is for when you need to do a transformation TODAY and all you have is a fresh culture of your bacteria, but NO COMPETENT CELLS…and you just need one colony harboring your favorite plasmid.  This is NOT for transformation efficient cells, but can save you a day’s time!!!

1. Pellet 1 ml of cell culture
2. Add 100 µl of 50 mM COLD CaCl₂
3. Add 1 µl plasmid DNA
4. Incubate on ice for 30 min
5. Heat-shock at 42ºC for 45 seconds
6. Immediately place tube on ice for 2 min
7. Add 450 µl TSB and gently mix
8. Incubate at 37ºC for 1 hour (*not any longer)
9. Plate on LB agar containing the correct antibiotics
10. It is recommended to plate 100 µl on one plate and plate the rest on another plate to avoid having a lawn
11. Incubate overnight @ 37ºC