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
 
Page visits: 1888