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Media Preparation

Author: Karen Betancourt Moreira, updated date: , view: 8, Q&A: 0
Tags: Yeast, Media and Recipes

Procedure

#2 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 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


Adenine

0.5g

Alanine

2g

Arginine

2g

Asparagine

2g

Aspartic acid

2g

Cysteine

2g

Glutamic acid

2g

Glutamine

2g

Glycine

2g

Histidine

2g

Myo-inositol

2g

Isoleucine

2g

Leucine

4g

Lysine

2g

Methionine

2g

para-Aminobenzoic acid

0.2g

Phenylalanine

2g

Proline

2g

Serine

2g

Threonine

2g

Tryptophan

2g

Tyrosine

2g

Uracil

2g

Valine

2g


  • 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



Amino Acids


Reference Sigma





Adenine


A-2786

5g

L-Alanine


A-7469

25g

L-Arginine


A-8094

25g

L-Asparagine


A-7074

25g

L-Aspartic acid


A-7219

100g

L-Cysteine


C-7477

25g

L-Glutamic acid


G-8415

100g

L-Glutamine


G-8540

25g

L-Glycine


G-8790

100g

L-Histidine


H-6034

25g

Myo-inositol


I-7508

50g

L-Isoleucine


I-7403

25g

L-Leucine


L-8912

25g

L-Lysine


L-9037

25g

L-Methionine


M-5308

25g

para-Aminobenzoic acid

A-9878

5g

L-Phenylalanine


P-5482

25g

L-Proline


P-5607

25g

L-Serine


S-4311

25g

L-Threonine


T-8441

25g

L-Tryptophan


T-8941

25g

L-Tyrosine


T-8566

25g

Uracil


U-1128

25g

L-Valine


V-0513

25g



  • 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.

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