Phenol-based Method for the Isolation of DNA Fragments
from Low-Melting Temperature Agarose
- Reference: Favre, D. 1992. Biotechniques vol. 13
- Cut out slice containing DNA, smallest size possible.
- Estimate volume and double with TE (10 mM Tris-HCl, pH 8.0/1 mM EDTA).
Melt at 65 C 5-10 min.
- Add 1 volume Tris-buffered phenol at room temperature, mix by inversion.
- Spin 3 min at 10-12k rpm and transfer aqueous phase to new tube. Phenol
- Spin as in (4) and transfer aqueous phase to new tube containing 0.1
volume 4 M LiCl. Mix by inversion; a white precipitate forms immediately.
Place tube on ice 2 min. Spin as above, 3 min.
- Transfer to new tube, leaving transparent pellet behind. Add 1 ul
carrier (glycogen) and precipitate with 2.5 volumes cold ethanol. Mix,
leave at -70 C 5-10 min, and spin as above, 10 min. Wash pellet with
1 ml 70% ethanol, dry under vacuum, and resuspend in 10-20 ul water