Yeast boiling DNA miniprep

 Yeast boiling DNA miniprep (Robzyk & Kassir, Nucl. Acid. Res. 20: 3790, 1992)

Harvest 1.5 ml of an ON yeast culture grown under selective conditions (5K, 5 minutes).

Resuspend in 100 µl STET (8% sucrose, 50 mM Tris pH 8, 50 mM EDTA, 5% Triton X-100).

Add 0.2 g of 0.45 mm glass beads and vortex vigorously for 5 minutes.

Add another 100 µl of STET, vortex briefly and place in a boiling water bath for 3 minutes.

Cool briefly on ice and spin in a microfuge for 10 minutes at 4℃.

Transfer 120 µl of sup to a new tube containing 60 µl of 7.5 M ammonium acetate, incubate 1 hour at -20℃ and then centrifuge for 10 minutes at 4℃.

Add 120 µl of the sup to 240 µl of ice-cold ethanol, incubate 30 minutes at -20℃ and then centrifuge for 10 minutes at 4℃.

Wash with 70% ethanol and resuspend in 25 µl TE.

Check 5 µl on a gel, use about 10 µl for E. coli transformation

Good for recovery of plasmids into E.coli.