Team:Purdue/E. coli transformation

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Materials

  • buckets of ice
  • 5 ml SOC
  • X21 Gold Competent Cells
  • Kan Plates
  • hot water bath
  • shaker in 37 degree room

Procedure

  1. Took XL-21 gold from -80, added 12μl to transformation tube and moved immediately to ice bucket
  2. Added 1.5 μl luciferase plasmid DNA (with kanamycin resistance) to eppendorf tube
  3. Prepared a control without the plasmid
  4. Stored on ice for 30 minutes
  5. Heat shock in 42 degree water bath for 28 sec
  6. Moved to ice bucket for 2 minutes
  7. Added 200μl SOC carefully to the control and the transformant
  8. Moved the tubes to 37 degree shaker, set at 100 rpm, tilted at an angle for better mixing – for one hour
  9. Obtained three kan plates and plated 50 μl control, 50 μl transform, and 100 μl of transform – streaked with glass beads
  10. Stored plates in 37 degree room for 11 hours
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