IGEM:Brown/2007/Lab Protocols/10min E coli Miniprep
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Stocks
TENS (for 100ml) *1ml 10N NaOH *1ml 20% SDS *1ml 1M Tris pH 7.5 *200 microliters 0.5 M EDTA *96.8 mL water 3 M NaOAc pH 5.2
Procudure
- Spin 1.5 ml o/n culture E coli in an eppendorf for 10 s
- Decant s/n and resuspend in the remaining volume
- Add 300 microliters TENS. Vortex to mix completely
- Add 150 microliters 3 M NaOAC. Mix by inversion
- Spin 10 min room temperature (debris, genomic DNA)
- Transfer s/n to fresh tube
- Add 900 microliters ice cold EtOH. Mix thoroughly
- Sping 15 min room temperature
- Wash pellet 1x 70% EtOH (optional)
- Dry pellet speed-vac
- Resuspend in 100 microliter TE. Use 1.5-3 microliters for digest
- Add 1 microliter1 1mg/ml RNAase to 10 microliter digest