IGEM:Brown/2007/Lab Protocols/10min E coli Miniprep

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

1. Spin 1.5 ml o/n culture E coli in an eppendorf for 10 s
2. Decant s/n and resuspend in the remaining volume
3. Add 300 microliters TENS. Vortex to mix completely
4. Add 150 microliters 3 M NaOAC. Mix by inversion
5. Spin 10 min room temperature (debris, genomic DNA)
6. Transfer s/n to fresh tube
7. Add 900 microliters ice cold EtOH. Mix thoroughly
8. Sping 15 min room temperature
9. Wash pellet 1x 70% EtOH (optional)
10. Dry pellet speed-vac
11. Resuspend in 100 microliter TE. Use 1.5-3 microliters for digest
12. Add 1 microliter1 1mg/ml RNAase to 10 microliter digest