Yeast DAPI Staining#

Protocol taken from here

Cite using: Alan Cone 2016. Yeast DAPI Staining. protocols.io https://dx.doi.org/10.17504/protocols.io.eigbcbw

Stains the nucleus of budding or fission yeast with DAPI in order to see it with a fluorescence microscope.

  1. Grow up yeast in liquid medium overnight. The OD may not matter too much here, but something in the 0.8 - 2 range is probably ideal.

  2. Add 333 μL (or 1 volume) yeast culture to a 1.5 mL microcentrifuge tube.

  3. Add 666 μL (or 2 volume) of 100% Ethanol to the 1.5 mL microcentrifuge tube.

  4. Let the yeast ethanol mixture sit at room temperature for 30-60 minutes.

  5. Spin down yeast cells for 1 minute at 2500 RPM.

  6. Pour out the supernatant and resuspend the pellet in 1mL of 1 x PBS(1X PBS (Phosphate-buffered saline )), then centrifuge for 1 minute at 2500 RPM.

  7. Pour out the supernatant and resuspend the pellet in 200 μL of a 1 x PBS / 1:2000 Dilution DAPI mixture 1.

  8. Add one drop of the yeast suspended in the PBS / DAPI mixture onto a microscope slide, add a coverslip on top, and go observe the stained yeast. Make sure after you do this part to go look at it within a few hours of resuspending in PBS / DAPI. The sooner you can get to a microscope the better.

1

Add 1 mL 1 x PBS to a 1.5 mL microcentrifuge tube.Add 0.5 μL of a 2.5 mg/mL (or a 1:2000 dilution) of DAPI to the 1 x PBS.