TITLE: Solid Growth Medium - Yeast
AUTHORS: Mathias Hammer, Ammeret Rossouw, Azra Lari, Ben Montpetit, David Grunwald

[DESCRIPTION]
This protocol describes the steps to prepare solid culture medium for Saccharomyces cerevisiae.

[BEFORE_START]
Have the following solutions premixed:

Glucose 20% 500 ml solution:
Concentration: 200 g/l
mix 100 g Glucose in 500 ml deionized water (ddH2O)

Sodium Hydroxide 1M solution 50 ml:
Concentration: 39.997 g/l
mix 1.99985 g NaOH into 50 ml ddH2O

Optional:
SC-xx 10x 100ml solution:
Concentration: 19.2 g/l
mix 1.92 g into 100 ml ddH2O

YNB 20x 100ml solution:
Concentration: 134.4 g/l
mix 13.44 g into 100 ml ddH2O

[STEPS]
SECTION: Preparation of ~20 agar plates of yeast growth medium
1. Compound medium for autoclave
SECTION: Preparation of ~20 agar plates of yeast growth medium
1.1. Fill a 500 ml flask with 449 mLddH2O. 
Add a magnetic stirring bar and place the flask on a stirring hot plate.
SECTION: Preparation of ~20 agar plates of yeast growth medium
1.2. Add 3.36 g Yeast Nitrogen Base with Ammonium Sulfate without Amino Acids (YNB).

SECTION: Preparation of ~20 agar plates of yeast growth medium
1.3. Add 0.86 g SC-XX powder.
 

SECTION: Preparation of ~20 agar plates of yeast growth medium
1.4. Add 10 g Agar.
SECTION: Preparation of ~20 agar plates of yeast growth medium
1.5. Add 1 mL NaOH 1 molar solution.
 

SECTION: Preparation of ~20 agar plates of yeast growth medium
2. Autoclave for 15 min at 121 °C.
 

SECTION: Preparation of ~20 agar plates of yeast growth medium
3. Cooling and platting
SECTION: Preparation of ~20 agar plates of yeast growth medium
3.1. Add a sterile stirring bar and thermometer. Let the medium stir to homogenize its temperature, while cooling.
SECTION: Preparation of ~20 agar plates of yeast growth medium
3.2. When the medium cooled down to around 80 °C add 50 mL sterile Glucose 20%.
SECTION: Preparation of ~20 agar plates of yeast growth medium
3.3. When the medium cooled down to 60 °C , plate 25 mL per  10 cmPetri dish (~20 dishes).
SECTION: Preparation of ~20 agar plates of yeast growth medium
3.4. Let the medium solidify and cool down to Room temperature.
SECTION: Preparation of ~20 agar plates of yeast growth medium
4. Seal the prepared dishes in a plastic bag to prevent condensation and store them in 4 °C fridge.
 

SECTION: Preparation of ~20 agar plates of yeast growth medium
1.1. Fill a 500 ml flask with 374 mLddH2O. 
Add a magnetic stirring bar and place the flask on a stirring hot plate.
SECTION: Preparation of ~20 agar plates of yeast growth medium
1.2. Add 25 mLYNB 20x solution (Yeast Nitrogen Base with Ammonium Sulfate without Amino Acids).

SECTION: Preparation of ~20 agar plates of yeast growth medium
1.3. Add 50 mL SC-XX 10x solution.
 

SECTION: Preparation of ~20 agar plates of yeast growth medium
1.4. Add 10 g Agar.
SECTION: Preparation of ~20 agar plates of yeast growth medium
1.5. Add 1 mL NaOH 1 molar solution.
 

SECTION: Preparation of ~20 agar plates of yeast growth medium
2. Autoclave for 15 min at 121 °C.
 

SECTION: Preparation of ~20 agar plates of yeast growth medium
3. Cooling and platting
SECTION: Preparation of ~20 agar plates of yeast growth medium
3.1. Add a sterile stirring bar and thermometer. Let the medium stir to homogenize its temperature, while cooling.
SECTION: Preparation of ~20 agar plates of yeast growth medium
3.2. When the medium cooled down to around 80 °C add 50 mL sterile Glucose 20%.
SECTION: Preparation of ~20 agar plates of yeast growth medium
3.3. When the medium cooled down to 60 °C , plate 25 mL per  10 cmPetri dish (~20 dishes).
SECTION: Preparation of ~20 agar plates of yeast growth medium
3.4. Let the medium solidify and cool down to Room temperature.
SECTION: Preparation of ~20 agar plates of yeast growth medium
4. Seal the prepared dishes in a plastic bag to prevent condensation and store them in 4 °C fridge.