Miniprep

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iGEM 2006 Lab Training Session II: Small Scale Plasmid Preparation (Miniprep)

Key Words: [http://en.wikipedia.org/wiki/Supernatant supernatant and pellet]

Procedure:

Preparation: overnight (o/n): 2.5mL LB broth, appropriate antibiotics, cells needed from single colony, incubate overnight

  • put 1.5mL o/n in a centrifuge tube
  • centrifuge for 1min
  • suck away supernatant
  • add 250uL Resuspension Solution (red)
  • vortex
  • add 250uL Lysis Solution (green)
  • shake with hands
  • sit for 3-5min
  • add 350uL Neutralization Solution (blue)
  • invert tube 4-6 times
  • centrifuge for 5min
  • pour supernatant in a column tube
  • centrifuge for 1min
  • throw away supernatant in a beaker
  • add 500uL Wash Solution
  • centrifuge for 1min
  • throw away supernatant
  • add 500uL Wash Solution
  • centrifuge for 1min
  • throw away supernatant
  • centrifuge for 1min
  • put blue column in a fresh centrifuge tube
  • add 50uL Elution Buffer
  • incubate for 2min
  • centrifuge for 2min
  • throw away blue column
  • store DNA in -20C freezer

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