Generation of Glycerol Stocks (For long term storage)
1.	Use a sterilized loop to inoculate the strain into 50 ml tube containing 5 ml YES+G418 medium. Use YES media without G418 when growing the wild type strains.
2.	Incubate the cells at 25-32°C with shaking for overnight or until the culture is turbid.
3.	Add 5 ml sterile 50% stock solution and mix thoroughly.
4.	Aliquot the stock into ultra-low temperature vials and store at -70°C.
5.	Revive the yeast by transferring a small portion of the frozen sample onto a YES+G418 agar plate. Use YES agar without the G418 when reviving the wild type strains.
The cells can then be stored at -70°C and remain viable for several years at least. It is wise to make a duplicate each time and store it in a different freezer. When using cells requiring the active nmt promoter to survive, freeze them in supplemented minimal media containing final concentration of 25% glycerol.

For strains stored on glycerol at -70°C:
1.	With a sterile spatula scrape off a small amount of frozen glycerol stock and patch onto a YES plate, keeping the stock frozen.
2.	Incubate at 25-32°C for 1-4 days, depending on the strain.
3.	When growth is visible streak out to single colonies on YES plates and incubate at 25-32°C for 2-3 days.
Strains stored as slants or patches are streaked out onto YES plates directly, and incubated at 25-32°C as appropriate.

Handling of 96-well plates type

After complete thawing, resuspend the cells by pipetting several times before replication since yeast has a tendency to sink to the bottom of the well without agitation. Avoid the repeated freeze-thaw as it decreases cell viability.
- Wash the lids with ethanol before returning them to their 96-well plates.
- When re-sealing 96-well plates, freeze the plates. Seal them before returning them to the freezer.
- Remove the foil seal only when the plate is frozen and allow the plates to thaw with the lids removed under a hood.

1.	Allow the plates to thaw completely and resuspend the cells before replication as yeast has a tendency to settle to the bottom of the well during growth. Avoid repeated freeze-thaw as it decreases cell viability.
2.	Use sterilized 96 lids (or wash the lids with ethanol) to inoculate the strain into 96 plates containing 120 ul YES / G418 medium. Use YES media without G418 when growing the wild type strain.
3.	Incubate the cells at 30-32°C for 2 days or until the culture is turbid.
4.	Add 80 ul sterile 50% stock solution and mix thoroughly.
5.	Aliquot the stock into ultra-low temperature vials and store at -70°C.
6.	Revive the yeast by transferring a small portion of the frozen sample onto YES/G418 agar plate. Use YES agar without the G418 when reviving the wild type strains.

Procedures for Verifying Haploids

1.	Preparation of the genomic DNA of mutants
	Colony or liquid cultures can be used for genomic DNA extraction. Automatic preparation is possible with ExiprepTM 16. Perform the extraction according to the AccuPrep® Genomic DNA Extraction Kit (Bioneer, K-3032, K-3032-2) The other extraction kit may be applicable.
2.	Run PCR with a cycling protocol described below:
	Add 1 ul of extracted genomic DNA and 1 ul of S. pombe QC Primer (CP3 or CP5) to AccuPower® PCR PreMix (Bioneer, K-2012) and 18 ul of DW. Spin down and vortex and spin down. Thermal cycle : 94°C 5 min, 94°C 40 sec, 50°C 40 sec, 72°C 40 sec(30 cycles), 72°C 5 min.
3.	Run electrophoresis for verification.
	Load the PCR products and carry out electrophoresis. Check the amplified bands.