Berkeley LBL/Miniprep


Miniprep Protocol using QlAprep Spin Miniprep Kit(1mL-5mL):

1. Palette cells by centifuging from liquid culture and discard the media, keeping the pellet.

2. Resuspend the pelleted bacterial cell in 250 uL Buffer P1 w/ RNase.

3. Add 250 uL Buffer P2 and gently invert the tube 4-6 times to lyse cell (do not allow the lysis reaction to go on for more than 5 minutes)

4. Immediately add 350 uL Buffer N3 and invert tube 4-6 times gently.

5. Centrifuge for 10 min. at maximum speed.

6. Transfer the supernatant retrieved from step 5 to a QlAprep Spin Column.

7. Centrifuge column for 1 min. and discard the flow-through.

8. Wash with 500 uL Buffer PB, centrifuge for 1 min. and discard the flow-through.

9. Wash with 750 uL Buffer PE w/ ethanol, centrifuge for 1 min and discard flow-through.

10. Centrifuge column for an additional minute to remove excess Buffer PE.

11. Transfer column to an eppendorf tube and elute with 50 uL 20% EB and let sit for 1 min.

12. Centrifuge for 1 min., and discard column.