Quick genomic DNA prep

Quick Yeast Genomic Prep for PCR

1. Scrape cells off a plate with a pipet tip or toothpick (pin head amount) and resuspend in 500ul H2O

2. Spin for 15 sec @ top speed to harvest cells

3. Resuspend pellet in 250ul of Lysis Buffer (0.1M Tris pH8, 0.25M NaCl, 50mM EDTA, 1% SDS), vortex

4. Add ~500ul of 0.5mm glass beads per sample and shake in Vivadent for 5 sec

5. Spin for 2min @ top speed to reduce foaming

6. Add 250ul of chloroform and shake in Vivadent for 2 sec

7. Spin for 5 min @ top speed to separate phases

8. Transfer ~200ul of upper phase to a new tube, add 500ul 100% EtOH, mix by inverting

9. Spin for 5 min @ top speed

10. Aspirate supernatant and wash pellet once with 70% EtOH

11. Air dry pellet and resuspend in ~300ul H2O

12. Spin DNA sample before setting up PCR to collect insoluble material on the bottom, use 1ul DNA per 10ul PCR reaction

Lysis Buffer: For 50ml:

0.1M Tris pH85ml 1M Tris

0.25M NaCl2.5ml 5M NaCl

50mM EDTA5ml 0.5M EDTA

1% SDS5ml 10% SDS