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Team:Alberta NINT/Protocols
From 2008.igem.org
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Swirl loop through LB + amp broth. | Swirl loop through LB + amp broth. | ||
Incubate overnight at 37 C. | Incubate overnight at 37 C. | ||
| + | |||
| + | '''Minipreps''' | ||
| + | QIAprep Spin Miniprep Protocol | ||
| + | |||
| + | Isolating DNA from LB culture tubes: | ||
| + | Add 2X 750ul broth from overnights into a 1.5ml tube. Centrifuge for 1 min to pellet and aspirate liquid. | ||
| + | Add 250ul Buffer P1 and resuspend pelleted cells. | ||
| + | Add 250ul Buffer P2 and invert tube 4-6 times. | ||
| + | Add 350ul Buffer N3 and invert tube immediately 4-6 times. | ||
| + | Centrifuge for 10 min. | ||
| + | Apply supernatant to QIAprep spin column. | ||
| + | Centrifuge for 1 min and discard flow through. | ||
| + | Add 500ul Buffer PB, centrifuge for 1 min and discard flow through (only if high nuclease activity). | ||
| + | Add 750ul Buffer PE, centrifuge for 1 min and discard flow through. | ||
| + | Centrifuge for an additional 1 min. | ||
| + | Place spin column in a clean 1.5ml tube. Add 50ul Buffer EB, let stand for 1 min and centrifuge for 1 min. | ||
Revision as of 17:52, 17 June 2008
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Lab Protocols
Coming Soon!
Overnights
Inoculating LB culture tubes:
Add 5ul Amp50 or Amp100 to 200ml LB broth in tube. Using sterile inoculating loop, touch an isolated colony. Swirl loop through LB + amp broth. Incubate overnight at 37 C.
Minipreps QIAprep Spin Miniprep Protocol
Isolating DNA from LB culture tubes:
Add 2X 750ul broth from overnights into a 1.5ml tube. Centrifuge for 1 min to pellet and aspirate liquid. Add 250ul Buffer P1 and resuspend pelleted cells. Add 250ul Buffer P2 and invert tube 4-6 times. Add 350ul Buffer N3 and invert tube immediately 4-6 times. Centrifuge for 10 min. Apply supernatant to QIAprep spin column. Centrifuge for 1 min and discard flow through. Add 500ul Buffer PB, centrifuge for 1 min and discard flow through (only if high nuclease activity). Add 750ul Buffer PE, centrifuge for 1 min and discard flow through. Centrifuge for an additional 1 min. Place spin column in a clean 1.5ml tube. Add 50ul Buffer EB, let stand for 1 min and centrifuge for 1 min.
