Joyce: Phenol extraction: Difference between revisions
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#Pipette off the ether phase (ether is on top), discard in a falcon tube containing activated charcoal in the fume hood | #Pipette off the ether phase (ether is on top), discard in a falcon tube containing activated charcoal in the fume hood | ||
#Repeat the ether wash again | #Repeat the ether wash again | ||
#[Joyce: DNA precipitation|Precipitate the DNA] | #[[Joyce: DNA precipitation|Precipitate the DNA]] | ||
Revision as of 22:20, 10 September 2009
Concept
- To remove contaminating protein that could inhibit downstream protocols, or affect the quality of your DNA in the long-term.
- Phenol completely denatures proteins and concentrates them in the nonpolar (phenol) phase, leaving DNA undamaged in the aqueous phase
Procedure
- In a fume hood and while wearing gloves, add an equal volume of TE-saturated phenol to your DNA sample WARNING: Wear gloves and do this in a fume hood; phenol is some nasty stuff
- Vortex for at least 1 minute
- Spin at 14K RPM for 2 minutes
- Pipette off the aqueous phase to a new tube
- Add an equal volume of water-saturated ether to the DNA solution (fume hood, wear gloves)
- Vortex for at least 1 minute
- Spin at 14K RPM for 2 minutes
- Pipette off the ether phase (ether is on top), discard in a falcon tube containing activated charcoal in the fume hood
- Repeat the ether wash again
- Precipitate the DNA
