Miniprep
A miniprep is done to purify plasmids from a strain
What you need:
- An overnight culture of your plasmid-containing cells
- Autoclaved 1.5mL centrifuge tubes
- Resuspension solution containing RNase A
- Lysis solution
- Neutralization solution
- Large Zymo-spin columns with collection tubes
- DNA wash buffer
- Autoclaved MilliQ water
Procedure:
1. To a 1.5 mL centrifuge tube, add 1.5 mL of overnight culture
2. Centrifuge at full speed for 2 minutes and discard supernatant
3. Resuspend the cells by pipetting in 250 uL resuspension solution
4. Add 250 uL lysis solution and mix by inversion (about 30 inversions)
5. Add 350 uL neutralization solution and mix by inversion
6. Centrifuge the mixture at full speed for 5 minutes
7. Transfer the supernatant to a Zymo-spin column with collection tube
8. Centrifuge for 1 minute and discard the flow-through in collection tube
9. Add 500 uL wash buffer to column
10. Centrifuge for 1 minute and discard the supernatant
11. Repeat steps 9-10
12. Spin the empty column for an additional minute
13. Place the column in a new 1.5 mL centrifuge tube
14. Add 50 uL autoclaved water directly to the column
15. Let the column incubate at room temperature for 2 minutes
16. Spin the column in the centrifuge for 2 minutes
17. Store the liquid at -20˚C until use
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