Small Scale Electrocompetent Prep-Generic

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Small Scale Electrocompetent Prep - Generic=

  1. Grow up strain to saturation
  2. Innoculate 2YT with 1:30 of o/n.
  3. Grow uninterrupted for 2.5 hours @ 37oC.
    1. Josh: use 4hrs @ 30C for ubercompetency
  4. Cool tube/flask; transfer 1.5 mL to chilled epi tube.
  5. Spin down for 20 sec at 14.1 rpm, discard supe, place immediately on ice.
  6. Resuspend with .5 mL 10% ice cold glycerol.
  7. Spin down for 30 sec (14.1 rpm), discard supe, place immediately on ice.
  8. Resuspend with .5 mL 10% ice cold glycerol.
  9. Spin down for 30 sec (14.1 rpm), discard supe, place immediately on ice.
  10. Resuspend in a small volume (90 ul) 10% ice cold glycerol.
  11. Add ~2uL DNA to cells, pipette & stir to mix.
  12. Immediately add to chilled electroporation cuvette.
  13. Immediately shock
  14. Immediately add 200 uL 2YT, pipette up and down to recover cells, move to tube and keep warm
  15. Put in shaking 37oC incubator for 1 hr
  16. Plate everything on appropriate antibiotic plate (allow to dry

before putting in 37oC incubator)

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