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Team:Paris Saclay/electro
From 2013.igem.org
(Created page with "{{Team:Paris_Saclay/incl_debut_generique}} = '''Electro-Elution''' = 1. Cut the strip under the UV plate, cut the gel in little pieces 2. Fill the vat with TAE 0.5X 3. Close o...") |
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| - | = ''' | + | = '''Electroelution''' = |
| - | 1. | + | 1. Run samples on agarose gel as usual. When fragments are well-seperated, slide out bands of interest with a razor blade under the UV plate. Make slices as thin as possible (trim off excess agarose).Slices can be stored in microfuge tubes in the refregerator if necessary. |
| - | 2. Fill the vat with TAE 0.5X | + | 2. Fill the vat with TAE 0.5X buffer |
| - | 3. Close one of the | + | 3. Close one of the wells of the saline bridge with a semi-permeable membrane, put the DNA in the large well |
4. Apply a 400V current after closing the vat, during 30min | 4. Apply a 400V current after closing the vat, during 30min | ||
| - | 5. Reverse the polarity during 20s, | + | 5. Reverse the polarity during 20s, pipet the solution from the small well and put it in a clean 1.5 ml tube |
| - | 6. Add 200µL of water and 600µL of phenol chloroform | + | 6. Add 200µL of water and 600µL of phenol/chloroform/isoamyl alcohol ratio 25/24/1 |
| - | 7. Vortex during | + | 7. Vortex during 1 min |
| - | 8. | + | 8. Pipet the aqueous phase |
| - | 9. Precipitation with ethanol | + | 9. Precipitation the DNA with ethanol (see protocol) |
Latest revision as of 09:53, 4 October 2013
Electroelution
1. Run samples on agarose gel as usual. When fragments are well-seperated, slide out bands of interest with a razor blade under the UV plate. Make slices as thin as possible (trim off excess agarose).Slices can be stored in microfuge tubes in the refregerator if necessary.
2. Fill the vat with TAE 0.5X buffer
3. Close one of the wells of the saline bridge with a semi-permeable membrane, put the DNA in the large well
4. Apply a 400V current after closing the vat, during 30min
5. Reverse the polarity during 20s, pipet the solution from the small well and put it in a clean 1.5 ml tube
6. Add 200µL of water and 600µL of phenol/chloroform/isoamyl alcohol ratio 25/24/1
7. Vortex during 1 min
8. Pipet the aqueous phase
9. Precipitation the DNA with ethanol (see protocol)
