Sporulation of Diploids

— Ryla Cantergiani 2020/03/30 17:07

Sporulation is how you generate haploid yeast strains from diploid yeast strains. An example of this would be to generate haploid deletion strains from a diploid strain from the het dip (heterozygous diploid) deletion collection.

If this doesn't work, try GNA sporulation, which takes longer but is reported to be more reliable.

You'll need:

• Culture Tubes
• YEP + 2% glucose
• pipette tips
• diploid strains of interest
• 30°C shaker

1. Set up and label culture tubes. Label each tube with a piece of tape and tough spot.
2. Fill each culture tube with 3 mL of YEP + 2% glucose.
3. Inoculate each tube using a pipette tip (P10 or P20 work well) from a single colony of your diploid strain in question. Circle, initial, and date the colony used. Shake at 30°C overnight.
4. The next day, add 3ml of Sporulation Media (SPM) to a new culture tube for each strain (be sure to label your new tubes).
5. Using 1000μL tips and a P1000 pipettor, for each strain, aliquot 300μL of your diploid culture into the new tube containing 3mL of Sporulation Media. Shake at 30°C for 5-7 days before going to Tetrad Dissection