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yeast plasmid prep
- inoculate 2 ml of Glucose medium and let grow over night @ 30°C
- spin down 1.5 ml of the over night culture and resuspend in0.5 ml S-buffer
- incubate 30 min, 37°C
- add 0.1 ml lysis solution, vortex
- incubate 30 min, 65°C
- add 0.166 ml potassium acetate and mix, chill on ice for 10 min
- spin down for 10 min, pour the SN into a new tube (2 ml!)
- DNA precipitation: add 0.8 ml EtOH, 10 min on ice, spin down for 10 min, pour off the SN
- wash DNA with 0.5 ml 70% EtOH, dry pellet
- dissolve DNA in 0.04 ml water
S-buffer:
- 10 mM K2HPO4; pH 7.2
- 10 mM EDTA
- 50 mM 2-Mercaptoethanol
- 50 µg/ml zymolase
lysis solution:
- 25 mM Tris-HCl; pH 7.5
- 25 mM EDTA
- 2.5% SDS
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