Flow Cytometry in yeast (Sytox green)
Pellet 107 cells and resuspend in 300 µl H2O
Transfer cells to 15 ml tubes
Slowly add 700 µl EtOH while vortexing
Incubate for 1 hr at RT
Centrifuge, resuspend in sodium citrate buffer (50 mM, pH 7.0) containing 250 µg/ml RNAseA and 1 mg/ml proteinase K
Incubate overnight at 37C
Centrifuge, resuspend in sodium citrate buffer and transfer to Falcon® 12 X 75 polystyrene tubes (# 2052, 2054, or 2058)
Sonicate (30% for 3 sec)
Add Sytox green (Invitrogen) to a final concentration of 1 µM and incubate for 1 hr at RT (or at 4C for longer-term)
Analyze by flow cytometry
Transfer cells to 15 ml tubes
Slowly add 700 µl EtOH while vortexing
Incubate for 1 hr at RT
Centrifuge, resuspend in sodium citrate buffer (50 mM, pH 7.0) containing 250 µg/ml RNAseA and 1 mg/ml proteinase K
Incubate overnight at 37C
Centrifuge, resuspend in sodium citrate buffer and transfer to Falcon® 12 X 75 polystyrene tubes (# 2052, 2054, or 2058)
Sonicate (30% for 3 sec)
Add Sytox green (Invitrogen) to a final concentration of 1 µM and incubate for 1 hr at RT (or at 4C for longer-term)
Analyze by flow cytometry