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MOPS Minimal Medium
Updated 5/5/03. Now this medium is available commercially.
Updated 6/14/02. Our protocol for preparing Neidhardt MOPS Minimal Medium.
Reference: F. C. Neidhardt, P. L. Bloch,
and D. F. Smith. 1974. Culture medium for enterobacteria. J Bacteriol
119(3): 736-747 [PubMed
Central].
MOPS Minimal Medium
10X MOPS mixture
100 ml
0.132 M K2HPO4
10 ml
milliQ H2O
880 ml
1mg/ml thiamine
0.1 ml
(optional - we do not use thiamine because it does not affect the growth rate of E. coli K-12 MG1655 in this medium.)
TOTAL
990 ml
1. Mix ingredients above and adjust the pH to 7.2 with approximately
300 microliters 10 M NaOH.
2. Filter sterilize. Can be stored at 4 degrees for up to 1 month.
3. Before use add 10 ml 100X carbon source (as appropriate - we typically
use a final concentration of 0.1% glucose).
10X MOPS Mixture
1. In a 1 L beaker with a stir bar, add the following to ~300 ml milliQ H2O:
Component FW grams MOPS
209.3
83.72
Tricine
179.2
7.17
2. Add 10 M KOH to a final pH of 7.4 (10 to 20 ml)
3. Bring total volume to 440 ml
4. Make fresh FeSO4 solution and add it to the MOPS/Tricine
solution:
Component FW grams H2O vol (ml) stock conc (M) FeSO4•7H2O 278
0.028
10 0.01
5. Add the following solutions to the MOPS/tricine/FeSO4 solution
(see below how to make each of these):
Mix in the order shown!
Component Volume 1.9 M NH4Cl
50 ml
0.276 M K2SO4
10 ml
0.02 M CaCl2•2H2O
0.25 ml
2.5 M MgCl2
2.1 ml
5 M NaCl
100 ml
Micronutrient stock
0.2 ml
Autoclaved milliQ H2O
387 ml
TOTAL
1000 ml
6. Filter sterilize with 1 L capacity 0.2 micron filter
7. Aliquot into sterile 100 or 200 ml plastic bottles and freeze at -20°.
Stocks used in 10X MOPS mixture
Make each separately, mixing the amount indicated into the specified volume. Store at room temp.
Component FW stock conc (M) grams vol (ml) note NH4Cl
53.49
1.9
50.82
500
K2SO4
174.3
0.276
4.8
100
CaCl2•2H2O
147
0.02
0.294
100
MgCl2
203.3
2.5
50.75
100
NaCl
58.44
5
292.2 1000
Micronutrient stock (100 ml)
Mix everything together in 40 ml autoclaved milliQ H2O, bring up total volume to 50 ml. Store at room temp.
Component Formula FW Grams for 50 ml ammonium molybdate
(NH4)6Mo7O24•4H2O
1235.9
0.009
boric acid
H3BO3
61.83
0.062
cobalt chloride
CoCl2
237.9
0.018
cupric sulfate
CuSO4
249.7
0.006
manganese chloride
MnCl2
197.9
0.040
zinc sulfate
ZnSO4
287.5
0.007
Potasium phosphate K2HPO4 Solution:
NOTE!! Make sure you use dibasic K2HPO4, and do NOT use monobasic KH2PO4 !!!!! Can be stored at room temperature after autoclaving.
Component FW stock conc (M) grams Vol (ml) note K2HPO4 173.2
0.132
23.0 1000 autoclave