Mini-prep for yeast genomic DNA enriched plasmids from lysed yeast powder
(Modified version by JLD of protocol from Clarence Chan // Heath Balcer’s NB p. 63)
1. Weigh out 0.1-0.15g of the lysed yeast powder of your choice. Thaw the powder
(in your hand)
2. Resuspend the powder in 1M Sorbitol / 0.01M EDTA pH 8.0 by pipetting.
3. Spin for 1 min.
4. Resuspend cells in 0.4ml of 0.15M NaCl / 0.01M EDTA pH 8.0 by pipetting.
5. Add 25µl of 20% SDS. Vortex immediately.
6. Let sit at room temperature for 20 min.
7. Incubate at 65°C for 10 min., then add 50µl of 5M potassium acetate. Mix
immediately without vortexing. Continue at 65°C for 5 min.
8. Place tubes on ice for 30 min., then centrifuge for 15 min.
9. Take the supernatant, add 50µl of 3M sodium acetate and 1ml of EtOH.
Centrifuge at room temperature for 10 min.
10. Discard supernatant and drain, rinse pellet with 1ml 70% EtOH, centrifuge
10min., decant and repeat with 100% EtOH. Air dry.
11. Dissolve pellet in 60µl of TE (65°C aids dissolution)
12. Optional: If you are going to manipulate the DNA (i.e., digest, gap repair…),
treat the DNA with RNase for 1 hr. prior to manipulation.