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Team:INSA Toulouse/contenu/lab practice/notebook/protocols/miniprep
From 2013.igem.org
Notebook
Protocols
DNA MINI Prep
Prinking out a colony on a petri Dish is the first step.
We use those buffers for extraction:
| Buffer 1 | Buffer 2 | Buffer 3 |
| TE 10:1 (mM) pH8 |
NaOH 0.2M SDS 1% |
AcOOK 3M AcOOH 15% (V/V) |
· Centrifugate of 2mL eppendorf (vortexing cells)
· +200µL TE 8.0 (mix become homogeneous)
· +400µL Buffer2 (wait 5 min in ice, invert tighltly)
· +300µL Buffer3 (no vortex)
· Centri. (10000 or + RPM) 4°C, 15’
· Keep the supernatant, on a 1.5mL eppendorf, add 600µL isopropanol
· Centri. (10000 or + RPM) 4°C, 15’
· Wash pellet with EtOH 70%
· Centri, discard supernatant
· Let EtOH dry
· Retake pellet with Buffer1 (pH 7.4)
