Isolating Yeast Genomic DNA Using Qiagen's DNeasy Kit

Grow yeast cells in YPD medium at 30° C to mid-log phase.

Pellet cells in 50 mL in blue cap tubes and spin at 3500 rpm for 5 minutes at RT° (room temperature). Use the Labofuge 400 with swinging bucket rotor.

Resuspend in 1.7 mL potassium phosphate/sorbitol buffer (10 mM potassium phosphate/ 1.2M sorbitol pH 7.2)

Add 5.1 µL b-mercaptoethanol (bME) and 544 µL lyticase (1 mg/mL stock solution made in water).

Place in shaker for 15 minutes at 30° C.

Pellet cells by spinning at 5000 rpm in the Beckman J2-21 using the JA-20 rotor. RT°

Wash 1X in 2 mL potassium potassium phosphate/sorbitol buffer. Pellet cells at 5K rpm in the Beckman J2-21 using the JA-20 rotor. RT°

Remove supernatant, add 1 mL H2O but do not disturb the pellet. Spin 5K rpm in the Beckman J2-21 using the JA-20 rotor at RT°. Use a pasteur pipet to remove the liquid and save cells.

Add 500 µL H2O and resuspend cells until no chunks are visible.

Freeze thaw in -70° C three times in a row. (> 20 minutes each time)

Follow DNeasy kit beginning with step number 2 on page 15. Use 50 µL of the frozen cell gamish for your gDNA extraction.


Lab Schedule Outlined

Lab Schedule In Context of Research Project

Molecular Biology Main Page

Course Materials

Biology Main Page

College Home Page search


© Copyright 2002 Department of Biology, Davidson College, Davidson, NC 28036
Send comments, questions, and suggestions to: macampbell@davidson.edu