IGEM:IMPERIAL/2006/LabCalendar/2006-7-11

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Miniprep- Prepare Plasmid DNA for electrophoresis

Follow miniprep protocol to prepare plasmid DNA samples for gel electrophoresis. Protocol for bacterial preparation for digestion:

  • Transfer 1.5 mL of culture into eppendorf
  • Spin for 20 sec and remove supernatant
  • Add 100 uL of solution 1 and vortex to resuspend
  • Add 200 uL of solution 2
  • Add 150 uL of solution 3
  • Mix by inverting, white ppt appears
  • Spin for 3 minutes
  • Pour supernatant in new labeled tube, add 1 mL of ethanol and mix
  • Spin for another 3 minutes and remove supernatant
  • Spin to remove resitudal liquid and then resuspend in 50 uL of MiliQ (ultrapure water) (1 sec on vortex, don't resuspend pellet)
  • Spin for 1 minute
  • Read to digest!

Make-up of solutions required for digest preparation

Solution 1

  • 90 g glucose
  • 10 mL 0.5M EDTA
  • 6.25 mL 2M Tris pH 8
  • Make up to 500 mL

Solution 2

  • 50 mL 10% SDS
  • 20 mL 5M NaOH
  • Make up to 500 mL

Solution 3

  • 147 g Potassium acetate
  • 57.5 mL glacial acetic acid
  • Make up to 500 mL
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