Freimoser:Recipes:Yeast:SC Sfree: Difference between revisions

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====1000x trace element solution 1, 10 ml====
====1000x trace element solution 1, 10 ml====
*Boric acid (H<sub>3</sub>BO<sub>3</sub>): 0.05 g (8 uM)
*Boric acid (H<sub>3</sub>BO<sub>3</sub>): 0.05 g (8 μM)
*CuCl<sub>2</sub> x 2 H<sub>2</sub>O: 0.004 g (0.25 uM) (replaces CuSO<sub>4</sub>)
*CuCl<sub>2</sub> x 2 H<sub>2</sub>O: 0.004 g (0.25 μM) (replaces CuSO<sub>4</sub>)
*KI (potassium iodide): 0.01 g (0.6 uM)
*KI (potassium iodide): 0.01 g (0.6 μM)
*C<sub>4</sub>H<sub>10</sub>O<sub>6</sub>Zn (zinc acetate): 0.055 g (2 uM) (replaces ZnSO<sub>4</sub>)
*C<sub>4</sub>H<sub>10</sub>O<sub>6</sub>Zn (zinc acetate): 0.055 g (2 μM) (replaces ZnSO<sub>4</sub>)


====1000x MnCl<sub>2</sub> x 4 H<sub>2</sub>O (manganese chloride):====
====1000x MnCl<sub>2</sub> x 4 H<sub>2</sub>O (manganese chloride):====
*0.04 g in 10 ml (1000x) (2 uM) (replaces MnSO<sub>4</sub>)
*0.04 g in 10 ml (1000x) (2 μM) (replaces MnSO<sub>4</sub>)


====1000 x Na<sub>2</sub>MoO<sub>4</sub> (sodium molybdate):====
====1000 x Na<sub>2</sub>MoO<sub>4</sub> (sodium molybdate):====
*0.02 g in 10 ml (1000x) (1 uM)
*0.02 g in 10 ml (1000x) (1 μM)


====500x FeCl<sub>3</sub> x 6 H<sub>2</sub>O (ferric chloride):====
====500x FeCl<sub>3</sub> x 6 H<sub>2</sub>O (ferric chloride):====
*0.02 g in 10 ml (500x) (1.5 uM)
*0.02 g in 10 ml (500x) (1.5 μM)


====1000x YNB vitamin solution====
====1000x YNB vitamin solution====
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====Complete amino acid mixture====
====Complete amino acid mixture====
We use commercially available complete amino acid mixtures and make a 100x stock solution that is filter sterilized.
We use commercially available complete amino acid mixtures and make a 10x stock solution that is filter sterilized.


====10x MES buffer (2-(N-morpholino)ethanesulfonic acid):====
====10x MES buffer (2-(N-morpholino)ethanesulfonic acid):====
*0.5 M, pH 6 (10x)
*0.5 M, pH 6 (10x)




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==Mixing==
==Mixing==


Volumes for the preparation of 50 ml 1.1x SC-S medium. For 1 ml final medium, 900 ul of this 1.1x stock and 100 ul of any sulfur source are used.
Volumes for the preparation of 50 ml 1.1x SC-S medium. For 1 ml final medium, 900 μl of this 1.1x stock and 100 μl of any sulfur source are used.


{|
{|
|
|
|-
|-
|width="200pt"|Compound
|width="150pt"|'''Compound'''
|width="150pt"|Not buffered Vol. (ul)
|width="175pt"|'''Not buffered, vol. to add [μl]'''
|width="150pt"|Buffered Vol. (ul)
|width="175pt"|'''Buffered, vol. to add [μl]'''
|-
|-
|'''H<sub>2</sub>O''' ||33'200 || 28'200
|'''H<sub>2</sub>O''' ||33'200 || 28'200

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Sulfur free SC-medium (self mixed SC medium)

Remark:

This recipe describes the mixing of a sulfur free SC-medium. All sulfur sources in “normal” SC-medium are replaced by other salts, mostly chlorides. The changes are also indicated. The phosphate source is separate in order to allow mixing of media with different phosphate content. The 20x salt solution, 100x phosphate source and 40% glucose solution were autoclaved. All other solutions were filter sterilized. The described procedure works, but it is possible that there are simpler solutions. But it is not possible to make a stock solution of all trace elements for example. The concentrations in parentheses indicate approximate concentrations in the final 1x medium.

20x Salt solution, 500 ml

  • MgCl2: 16 g (4 mM)
  • NaCl: 1 g (1.7 mM)
  • CaCl2 x 2 H2O: 1 g (1 mM)

1000x trace element solution 1, 10 ml

  • Boric acid (H3BO3): 0.05 g (8 μM)
  • CuCl2 x 2 H2O: 0.004 g (0.25 μM) (replaces CuSO4)
  • KI (potassium iodide): 0.01 g (0.6 μM)
  • C4H10O6Zn (zinc acetate): 0.055 g (2 μM) (replaces ZnSO4)

1000x MnCl2 x 4 H2O (manganese chloride):

  • 0.04 g in 10 ml (1000x) (2 μM) (replaces MnSO4)

1000 x Na2MoO4 (sodium molybdate):

  • 0.02 g in 10 ml (1000x) (1 μM)

500x FeCl3 x 6 H2O (ferric chloride):

  • 0.02 g in 10 ml (500x) (1.5 μM)

1000x YNB vitamin solution

100x NH4Cl (ammonium chloride):

  • 2 g in 10 ml (37.8 mM)

100 x phosphate source, 100 ml

  • KH2PO4: 10 g (7.5 mM)

20x Glucose (C6H12O6):

  • 40% (20x)

Complete amino acid mixture

We use commercially available complete amino acid mixtures and make a 10x stock solution that is filter sterilized.

10x MES buffer (2-(N-morpholino)ethanesulfonic acid):

  • 0.5 M, pH 6 (10x)


Mixing

Volumes for the preparation of 50 ml 1.1x SC-S medium. For 1 ml final medium, 900 μl of this 1.1x stock and 100 μl of any sulfur source are used.

Compound Not buffered, vol. to add [μl] Buffered, vol. to add [μl]
H2O 33'200 28'200
Salt solution 2'500 2'500
Trace elements 50 50
MnCl2 50 50
Na2MoO4 50 50
FeCl3 100 100
Vitamins 50 50
NH4Cl 1'000 1'000
KH2PO4 500 500
Glucose 2'500 2'500
Amino acids 5'000 5'000
MES buffer - 5'000
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