Team:Arizona State/Lab/Protocols/Extraction

Miniprep:

[http://www.qiagen.com/products/plasmid/qiaprepminiprepsystem/qiaprepspinminiprepkit.aspx QIAprep Spin Miniprep Kit using micro-centrifuge:]

1. Add 1.5 mL of liquid culture to microcentrifuge tube and centrifuge at 13,000 rpm for 1 minute. Empty out supernatant and repeat. Empty out excess supernatant afterwards.
2. Resuspend pelleted bacterial cells in 250 μL Buffer P1 and vortex.
3. Add 250 μL Bufer P2 and mix thoroughly by inverting the tube 4-6 times.
4. Add 350 μL Buffer N3 and mix immediately and thoroughly by inverting the tube 4-6 times.
5. Centrifuge for 10 min at 13,000 rpm.
6. Apply the supernatant from step 5 to the QIAprep spin column by decanting or pipetting.
7. Centrifuge for 30-60 seconds. Discard the flow-through.
8. Wash the QIAprep spin column by adding 0.5 ml Buffer PB and centrifuging for 30-60 seconds. Discard the flow-through.
9. Wash QIAprep column by adding 0.75 ml Buffer PE and centrifuging for 30-60 seconds.
10. Discard the flow through, and centrifuge for an additional 1 minute to remove residual wash buffer.
11. To elute DNA, place the QIAprep column in a clean 1.5 ml microcentrifuge tube. Add 50 μL PCR water to the center of each QIAprep spin column, let stand for 1 minute, and centrifuge for 1minute.