Yeast Miniprep
from Robzyk and Kassir, Nucleic Acid Res. (1992) 20:3790
Materials:
• 10 ml overnight culture yeast in drop out media (must be very dense)
• STET (8% sucrose, 50 mM Tris pH 8, 50 mM EDTA, 5% Triton X-100)
• Glass beads
• 7.5 M NH4OAc
• ice cold EtOH
• 70% EtOH
Procedure:
1) Spin down yeast culture. Aspirate off liquid and resuspend in 100 µl STET. Transfer to screwtop eppendorf tubes.
2) Add 2 scoops of glass beads (approx 200 mg) and vortex for 5 min.
3) Add another 100 µl STET. Vortex briefly to resuspend.
4) Boil for 3 min in boiling water bath.
5) Cool briefly on ice and spin in microfuge for 10 min at 4¡.
6) Transfer 100 µl of supe to new tube.
7) Add 50 µl 7.5 M NH4OAc. Incubate at -20¡ for 1 hr.
8) Spin 10 min at 4¡. (This step gets rid of impurities that inhibit transformations).
9) Collect 100 µl supe and add to 200 µl ice cold EtOH.
10) Spin 10 min at 4¡.
11) Rinse pellet with 70% EtOH.
12) Resuspend pellet in 20 µl of water. Use 10 µl to transform bacteria. Can do a regular transformation.
