Miniprep
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Revision as of 16:36, 28 June 2006
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
