Jonesma Lab:Protocols:Ecolitransformation

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E. coli transformation

  1. Take aliquot of E. coli (Top10) from -80°C storage
  2. Allow to thaw on ice
  3. Add DNA (1μL of a vector or 10μL of a ligation reaction)
  4. Chill mixture on ice for 30 min
  5. Turn on waterbath to 42°C
  6. Move bacteria to waterbath (at 42°C) for 30s
  7. Immediately put back on ice for 5 min to chill
  8. Add 1mL of LB
  9. Shake at 37°C for 1 hr
  10. Spin at 3000rpm for 15 min
  11. Take off most of excess liquid and resuspend in approx 100μL LB
  12. Plate on LB plates containing your antibiotic of choice O/N at 37°C