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Purifying DNA from an enzymatic reaction
From 2008.igem.org
Revision as of 12:05, 19 September 2008 by Riachalder (Talk | contribs)
1) Sample Capture
- Add 500 μl Capture buffer type 2 to up to 100 μl sample.
Note: If sample volume is greater than 100 μl, divide the sample and purify using more than one GFX MicroSpin column.
- Mix thoroughly.
Note: If sample contains DNA greater than 5 kbp, do not vortex, as this may cause shearing of the DNA.
- For each purification that is to be performed, place one GFX MicroSpin column into one Collection tube.
2) Sample Binding
- Centrifuge Capture buffer type 2-sample mix briefly to collect the liquid at the bottom of the tube.
- Load the Capture buffer type 2-sample mix onto the assembled GFX MicroSpin column and Collection tube.
- Spin the assembled column and Collection tube at 16 000 × g for 30 seconds.
- Discard the flow through by emptying theCollection tube. Place the GFX MicroSpin column back inside the Collection tube.
3) Wash & Dry
- Add 500 μl Wash buffer type 1 to the GFX MicroSpin column.
- Spin the assembled column and Collection tube at 16 000 × g for 30 seconds.
- Discard the Collection tube and transfer the GFX MicroSpin column to a fresh DNase-free 1.5 ml microcentrifuge tube
4) Elution
- Add 10–50 μl Elution buffer type 4 OR type 6 to the center of the membrane in the assembled GFX MicroSpin column and sample Collection tube.
- Incubate the assembled GFX MicroSpin column and sample Collection tube at room temperature for 1 minute.
- Spin the assembled column and sample Collection tube at 16 000 × g for 1 minute to recover the purified DNA.
- Proceed to downstream application. Store the purified DNA at -20°C.
