Media Preparation

1.7 g Yeast nitrogen base without amino-acids without ammonium sulfate

5 g Ammonium sulfate

2 g drop-out powder mix

20g Bacto Agar

900 mls MilliQ water

Stir until all ingredients are dissolved

Autoclave at 121 C for 25 minutes.

20 g Glucose

100 mls MilliQ water

Stir until all ingredients are dissolved

Autoclave at 121 C for 25 minutes.

After autoclaving mix the two solutions

Stir slowly to avoid bubbles

Let cool down on the side for 45 minutes, then pour 1L= 2 sleeves

#3 Drop-out  plate medium (1 Liter)

1.7 g Yeast nitrogen base without amino-acids without ammonium sulfate

5 g Ammonium sulfate

2 g drop-out powder mix

20g Bacto Agar

900 mls MilliQ water

Stir until all ingredients are dissolved

Autoclave at 121 C for 25 minutes.

20 g Raffinose

100 mls MilliQ water

Stir until all ingredients are dissolved

Autoclave at 121 C for 25 minutes.

After autoclaving mix the two solutions

Stir slowly to avoid bubbles

Let cool down on the side for 45 minutes, then pour 1L= 2 sleeves

#4 Drop-out  plate medium (1 Liter)

1.7 g Yeast nitrogen base without amino-acids without ammonium sulfate

5 g Ammonium sulfate

2 g drop-out powder mix

20g Bacto Agar

900 mls MilliQ water

Stir until all ingredients are dissolved

Autoclave at 121 C for 25 minutes.

20 g Raffinose

20 g Galactose

100 mls MilliQ water

Stir, but ingridients will not dissovle

Autoclave at 121 C for 25 minutes.

After autoclaving mix the two solutions

Stir slowly to avoid bubbles

Let cool down on the side for 45 minutes, then pour 1L= 2 sleeves

YPD plate medium (1 Liter)

10 g yeast extract

20 g peptone

20g Bacto Agar

900 mls MilliQ water

Stir until all ingredients are dissolved

Autoclave at 121 C for 25 minutes.

20 g Glucose

100 mls MilliQ water

Stir, but ingridients will not dissovle

Autoclave at 121 C for 25 minutes.

For plates with antibiotics add the following:

4 ml G418 (50 mg/ml) = Kan or Geniticin

300 ul NAT (200 mg/ml) = clonNAT

4 ml Hygromycin B (50 mg/ml)

1 ml Chloramphenicol (40 mg/ml)

Let cool down on the side for 45 minutes

Add antibiotic and stir slowly to avoid bubbles

Pour plates right away 1L= 2 sleeves

Kac – sporulation plate media (1 Liter)

20 g Potassium acetate

20 g Bacto Agar

100 mls MilliQ water

Adjust  to pH 7.0

Stir until all ingredients are dissolved

Autoclave at 121 C for 25 minutes.

Let cool down on the side for 45 minutes, then pour 1L= 2 sleeves

5 FoA plate media (1 Liter)

3.35 g Yeast nitrogen base without amino-acids without ammonium sulfate

10 g glucose

1g         drop-out powder mix (Complete)

1000 mls MilliQ water

Adjust  to pH 7.0

Stir until all ingredients are dissolved

Autoclave at 121 C for 25 minutes.

Let cool down on the side for 45 minutes, then pour 1L= 2 sleeves

#2 Drop-out  liquid medium (1 Liter)

1.7 g Yeast nitrogen base without amino-acids without ammonium sulfate

5 g Ammonium sulfate

2 g drop-out powder mix

900 mls MilliQ water

Stir until all ingredients are dissolved

Autoclave at 121 C for 25 minutes.

20 g Glucose

100 mls MilliQ water

Stir until all ingredients are dissolved

Autoclave at 121 C for 25 minutes.

After autoclaving mix the two solutions

Stir slowly to avoid bubbles

Let cool down on the side for 45 minutes, then pour 1L= 2 sleeves

#3 Drop-out  liquid medium (1 Liter)

1.7 g Yeast nitrogen base without amino-acids without ammonium sulfate

5 g Ammonium sulfate

2 g drop-out powder mix

900 mls MilliQ water

Stir until all ingredients are dissolved

Autoclave at 121 C for 25 minutes.

20 g Raffinose

100 mls MilliQ water

Stir until all ingredients are dissolved

Autoclave at 121 C for 25 minutes.

After autoclaving mix the two solutions

Stir slowly to avoid bubbles

Let cool down on the side for 45 minutes, then pour 1L= 2 sleeves

#4 Drop-out  liquid medium (1 Liter)

1.7 g Yeast nitrogen base without amino-acids without ammonium sulfate

5 g Ammonium sulfate

2 g drop-out powder mix

900 mls MilliQ water

Stir until all ingredients are dissolved

Autoclave at 121 C for 25 minutes.

20 g Raffinose

20 g Galactose

100 mls MilliQ water

Stir, but ingridients will not dissovle

Autoclave at 121 C for 25 minutes.

After autoclaving mix the two solutions

Stir slowly to avoid bubbles

Let cool down on the side for 45 minutes, then pour 1L= 2 sleeves

YPD liquid medium (1 Liter)

10 g yeast extract

20 g peptone

20g                 Bacto Agar

900 mls     MilliQ water

Stir until all ingredients are dissolved

Autoclave at 121 C for 25 minutes.

20 g Glucose

100 mls MilliQ water

Stir, but ingridients will not dissovle

Autoclave at 121 C for 25 minutes.

LB  plate medium (1 Liter)

10 g Tryptone

5 g Yeast Extract

10 g Sodium Chloride

15 g Bacto Agar

1000 mls MilliQ water

Stir until all ingredients are dissolved

Autoclave at 121 C for 25 minutes.

For plates with antibiotics add the following, aliquots stored at -20 deg:

1 ml Carbenicillin (100 mg/ml) = Ampicilin

1 ml Kanamycin (50 mg/ml)

1 ml Chloramphenicol (40 mg/ml)

Let cool down on the side for 45 minutes

Add antibiotic and stir slowly to avoid bubbles

Pour plates right away 1L= 2 sleeves

LB liquid medium (1 Liter)

10 g Tryptone

5 g Yeast Extract

10 g             Sodium Chloride

1000 mls MilliQ water

Stir until all ingredients are dissolved

Autoclave at 121 C for 25 minutes.

TB  liquid medium (1 Liter)

47.6 g Terrific Broth powder (RPI)

6 ml Glycerol

1000 mls MilliQ water

Stir until all ingredients are dissolved

Autoclave at 121 C for 25 minutes.

Drop-out powder mix

• Weight all the powders except the one/ones that you do not need.

• Put the mix in a mortar and use the pestle to grind the powder.

• Put the mix in a clean plastic container and shake several times.

References to order all the ingredients

• D-glucose                 Sigma, ref G-8270

• D-Raffinose RPI, R20500-1000.0

• D-Galactose RPI, G33000-500.0

• Ammonium Sulfate Sigma, ref A-2939

• Bacto Agar               Difco, ref 2140-10

• Yeast nitrogen base w/o aa w/o ammonium sulfate Difco, ref 0335-15

• Bacto Triptone BD, 211705

• Bacto Peptone BD, DF0118-17-0

• Terrific Broth RPI, T15000-5000.