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M9 mineral medium

More details here:https://wiki.epfl.ch/ptpsp/documents/recipe%20for%20protein%20labeling.docx

For 1 liter M9 mineral medium add to 867 ml sterile water:

100 mL	M9 salt solution (10X)	Na2HPO4 KH2PO4 NaCl NH4Cl	33.7 mM 22.0 mM 8.55 mM 9.35 mM
20 mL	20% glucose	glucose	0.4 %
1 mL	1 M MgSO4	MgSO4	1 mM
0.3 mL	1 M CaCl2	CaCl2	0.3 mM
1 mL	biotin (1 mg/mL)	biotin	1 ug
1 mL	thiamine (1 mg/mL)	thiamine	1 ug
	trace elements solution (500X) made by Luciano	trace elements	1X

For every 1L for labelling add:

1 g of NH4Cl (Ammonium Chloride)

4 g of Glucose

Stock solutions

M9 salt solution (10X)

Na2HPO4-2H2O

KH2PO4

NaCl

NH4Cl (EXCLUDE if you want to use a stock for both labelled and unlabelled)

75.2 g/L

30 g/L

5 g/L

Dissolve the salts in 800 ml water and adjust the pH to 7.2 with NaOH. Add water to a final volume of 1 L and autoclave for 15 min at 121°C.

20% Glucose

20% (w/v) glucose

200 g/L

For 500 ml stock solution add 100 g glucose to 440 ml water. Autoclave for 15 min at 121°C.

1 M MgSO4

1 M MgSO4-7H2O

24.65 g/100 L

For 100 ml stock solution dissolve 24.65 g MgSO4-7H2O in 87 ml water. Autoclave for 15 min at 121°C

1 M CaCl2

1 M CaCl2-2H2O

14.7 g/100 mL

For 100 ml stock solution dissolve 14,70 g CaCl2-2H2O in 94.5 ml water. Autoclave for 15 min at 121°C.

Biotin 1mg/mL

biotin

50 mg/50 mL

For 50 ml stock solution dissolve 50 mg biotin in 45 ml water. Add small aliquots of 1N NaOH until the biotin has dissolved. Measure pH, should be neutral slightly basic. Add water to a final volume of 50 ml. Sterilize the solution over a 0.22-μM filter. Prepare 1 ml aliquots and store at -20°C.

Thiamine 1mg/mL

thamine-HCl

50 mg/50 mL

For 50 ml stock solution dissolve 50 mg thiamin-HCl in 45 ml water. Add water to a final volume of 50 ml. Sterilize the solution over a 0.22-μm filter. Prepare 1 ml aliquots and store at -20°C.