Yeast Growth Protocols

Find more protocols and selection guides in the Molecular Biology Guide.

Introduction

Yeasts are considered model systems for eukaryotic studies as they exhibit fast growth and have dispersed cells. Moreover, replica plating and mutant isolation of yeast cells can be done with relative ease and they have a well-defined genetic system. Most significantly, yeasts have a highly versatile DNA transformation system that can be utilized effectively for protein production.

Yeast Cultures

For general experimental purposes, yeasts are usually grown in YPD or synthetic media at 30°C. The following growth media and supplements are available from Sigma-Aldrich for yeast cultures:

Table 1: Yeast Culture Media
 

Catalog Number Name
Y1375 YPD Broth
Y1500 YPD Agar
Y0626 Yeast Nitrogen Base Without Amino Acids
Y1250 Yeast Nitrogen Base With Amino Acids

 

Table 2: Synthetic Drop-Out Medium Supplements (used along with Yeast Nitrogen Base without amino acids, Catalog Number Y0626)
 

Catalog Number Name
Y1501 Uracil
Y1751 Histidine
Y1896 Lysine
Y1376 Leucine
Y1876 Tryptophan
Y0750 Leucine and Tryptophan
Y1771 Uracil, Leucine, and Tryptophan
Y2146 Histidine, Leucine, and Tryptophan
Y2021 Histidine, Leucine, Tryptophan, and Adenine
Y2001 Histidine, Leucine, Tryptophan, and Uracil

 

Table 3: Other Supplements for Yeast Cultures
 

Catalog Number Name
T2559
EZMix™ Tryptone
G5516 Glycerol


For further details regarding yeast media, please refer to our Article, Introduction to Yeast Media.

Growth Protocols for Yeast

The protocols listed below can be used for growing and maintaining yeasts in liquid and solid media

Yeast Growth in YPD Broth

  1. Suspend 50g of the Product (Catalog Number Y1375) in 1L of distilled water.
  2. Autoclave for 15 minutes at 121°C.
  3. Inoculate yeast cultures (sourced from yeast broths/plates) in detergent-free tubes/flasks containing the prepared liquid medium, see steps 1 and 2 (the medium should not be more than a third of culture tube volume or a fifth of the total flask volume).
  4. Vortex the contents briefly to disperse cells.
  5. Grow the culture in a shaking incubator at 300 rpm (for flasks) or 350 rpm (for culture tubes).

Yeast Growth on YPD Agar

  1. Suspend 65g of the product (Catalog Number Y1500) in 1L of distilled water.
  2. Heat to boiling while stirring to dissolve all ingredients completely.
  3. Autoclave for 15 minutes at 121°C.
  4. Pour around 25-30mL of the agar medium on to sterile plates and let it set in a laminar flow chamber.
  5. Streak (yeast obtained from YPD plate)/spread (yeast sourced from broth cultures) cells on the agar plates and incubate at 30°C.

Note: Single yeast colonies may be observed after around 24 hours, but incubations over 48 hours are needed before they can be used for replica plating purposes. The growth rate of yeast cultures using synthetic drop-out medium supplements is ~50% slower.

Yeast Strain Preservation and Revival

Yeast cells grown using the above methods can be stored frozen in glycerol at -70°C for more than 3 years or in rich medium slants at 4°C for 6 months to a year. YPD plates with yeast cells can also be stored at 4°C for 2-3 months sealed with Parafilm®. The protocols for the preservation and revival of yeast cells are listed below:

Yeast Cells Preservation

  1. Prepare a sterile 30% (w/v) glycerol solution (Catalog Number G5516).
  2. Add 1.0mL of the glycerol solution into 4mL sterile screw-cap vials
  3. Take 1.0mL of early stationary/late log phase yeast broth
    OR Scrape a large inoculum from a freshly grown plate.
  4. Re-suspend in 1mL of sterile 30% glycerol solution.
  5. Mix, freeze on dry ice, and store at –70°C.
    Note:
    Yeast cells can also be stored similarly using 8% (v/v) dimethylsulfoxide (DMSO, Catalog Number D8418).

Yeast Strain Revival

  1. Scrape frozen stock with a sterile toothpick or bacteriological needle.
  2. Streak yeast cells on the appropriate plate and incubate at 30°C.
    Note: Avoid thawing the stock. In case the stock has thawed, vortex well before refreezing.

Materials

     

References

  1. Sherman, F., Getting started with yeast, Methods Enzymol., 350, 3-41 (2002).
  2. Methods in Molecular Biology, Vol. 177, Two-Hybrid Systems: Methods and Protocols, May 2001, Springer Protocols
  3. Current Protocols in Molecular Biology 13.2.1-13.2.12, April 2008, Wiley Interscience
  4. Schiestl and Gietz (1989) Curr. Genetics 16:339-346; Gietz, et al (1995) Yeast 11:355-360
  5. Li, B., and Fields, S., Identification of mutations in p53 that affect its binding to SV40 large T antigen by using the yeast two-hybrid system. The FASEB journal, 7(10), 957-963 (1993).

 

EZMix is a trademark of Sigma-Aldrich Co. LLC.

Parafilm is a registered trademark of Bemis Company, Inc.

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