Quick Yeast Genomic DNA Prep
1) Grow 10 ml yeast cultures to saturation in YEPD.
2) Pour the culture into a 15 ml conical and spin at 3000 rpm for 3 min. Dump sup. Resuspend in 500 µl distilled water.
3) Transfer to microfuge tube. Spin at 13000 rpm for 1 min. Dump sup. Vortex briefly to resuspend the pellet in the residual liquid.
4) Add 200 µl detergent lysis buffer, 200 µl phenol chloroform (at 4°C, take bottom layer), and ~300 mg glass beads (1 scoop).
5) Vortex 3-4 minutes on multihead vortex.
6) Add 200 µl TE pH 8. Spin at 13000 rpm for 5 minutes. Transfer aqueous layer (top) to a fresh tube.
7) Add 1 ml of ice cold 100% EtOH. Mix by inversion.
8) Spin at 13000 rpm for 2 minutes at 4°C. Dump sup. Resuspend in 400 µl TE pH 8 and 3 µl RNase.
9) Incubate 5 minutes at 37°C.
10) Add 10 µl 4 M ammonium acetate plus 1 ml of ice cold 100% EtOH. Mix by inversion
11) Spin at 13000 rpm for 2 minutes at 4°C. Dump sup. Air-dry pellet.
12) Resuspend in 50 µl TE.
13) Store at -20°C.
Materials:
-Saturated culture
-Detergent lysis buffer:
2% Triton X 100
1% SDS
100 mM NaCl
10 mM Tris-Cl pH 8
1 mM EDTA
-PCI: Phenol, chloroform, IAA (25:24:1)
-acid-washed glass beads
-TE pH 8
-10 mg/ml boiled RNase
-4 M ammonium acetate
