BG-11 Medium 

 
Directions  Improved recipe as of March 2009
For 1 L Total
Liquid media:
1. To approximately 900 mL of dH2O add the first 9 components in the order specified while stirring
continuously.
2. Bring total volume to 1 L with dH2O.
3. Cover and autoclave medium.
4. Allow to cool then store at refrigerator temperature.
Agar media:
1. To approximately 400 mL of dH2O add the first 9 components in the order specified while stirring
continuously.
2. Bring total volume to 500 mL with dH2O.
3. In a separate container add 15 g of agar to 500 mL of dH2O.
4. Cover and autoclave both solutions.
5. In a water bath allow both solutions to cool to 45-50°C.
6. Add sterile Sodium Thiosulfate to agar solution and mix well.
7. Combine both agar and liquid solutions, mix well. Note the agar can solidify quickly.
8. Allow to cool then store at refrigerator temperature. 

# Component Amount Stock Solution Final Concentration

Concentration
NaNO3 (Fisher BP360-
1 10 mL/L 30 g/200 mL dH2O 17.6 mM
500)
2 K2HPO4 (Sigma P 3786) 10 mL/L 0.8 g/200 mL dH2O 0.22 mM
MgSO4·7H2O (Sigma
3 10 mL/L 0.15 g/200 mL dH2O 0.03 mM
230391)
CaCl2·2H2O (Sigma C-
4 10 mL/L 0.72 g/200 mL dH2O 0.2 mM
3881)
Citric Acid·H2O (Fisher
5 10 mL/L 0.12 g/200 mL dH2O 0.03 mM
A 104)
Ammonium Ferric
6 10 mL/L 0.12 g/200 mL dH2O 0.02 mM
Citrate
Na2EDTA·2H2O (Sigma
7 10 mL/L 0.02 g/200 mL dH2O 0.002 mM
ED255)
8 Na2CO3 (Baker 3604) 10 mL/L 0.4 g/200 mL dH2O 0.18 mM
BG-11 Trace Metals
9 1 mL/L    
Solution
Sodium Thiosulfate
10 Pentahydrate (agar media 24.8 g/100 mL   1 mM
only,sterile) (Baker 3946)

BG-11 Trace Metals Solution  

Directions 
For 1 L Total
1. To approximately 900 mL of dH2O add the components in the order specified while stirring
continuously.
2. Bring total volume to 1 L with dH2O.
3. Store at refrigerator temperature.

# Component Amount Stock Solution Final Concentration

Concentration
1 H3BO3 (Baker 0084) 2.86 g/L   46 µM
MnCl2·4H2O (Baker
2 1.81 g/L   9 µM
2540)
ZnSO4·7H2O (Sigma Z
3 0.22 g/L   0.77 µM
   0251)
Na2MoO4·2H2O (J.T.
4 0.39 g/L   1.6 µM
Baker 3764)
CuSO4·5H2O (MCIB
5 0.079 g/L   0.3 µM
3M11)
Co(NO3)2·6H2O
6 49.4 mg/L   0.17 µM
(Mallinckroft 4544)
Artificial Seawater Medium  

Directions 

Modification of Brand's ASP2 medium for 2268 Agmenellum quadruplicatum; suitable for
other axenic marine cultures (salinity = 30 ppt).

For 1 L Total
1. To approximately 850 mL of dH2O, add each of the components in the order specified
(except vitamins) while stirring continuously.
2. Adjust the pH to 8.1.
3. Bring the total volume to 1 L with dH2O.
*For 1.5% agar medium: add 15 g of agar to the flask; do not mix.
4. Cover and autoclave medium.
5. Allow to cool and add vitamin B12.
*For agar medium: add vitamin, mix, and dispense before agar solidifies.
6. Store at refrigerator temperature.

   # Component Amount Stock Solution Final Concentration

Concentration
1 NaCl (Fisher S271-500) 18 g/L   0.31 M
MgSO4·7H2O (Sigma
2 10 mL/L 26 g/100 mL dH20 10.5 M
230391)
3 KCl (Fisher P 217) 10 mL/L 6 g/100 mL dH20 8M
NaNO3 (Fisher BP360-
4 10 mL/L 10 g/100 mL dH20 11.8 M
500)
 CaCl2·2H2O (Sigma C-
5 10 mL/L 3 g/100 mL dH20 2M
3881)
6 KH2PO4 (Sigma P 0662) 10 mL/L 0.5 g/100 mL dH20 0.29 M
Tricine (adjust to pH 8)
7 20 mL/L 22.4 g/100 mL dH20 25 M
(Sigma T-5816-256)
8 P-II Metal Solution 10 mL/L    
9 Chelated Iron Solution 1 mL/L    
NH4Cl (Fisher A 649-
10 1 mL/L 2.7 g/100 mL dH20 0.5 M
500)
11 Vitamin B12 1 mL/L    
P-II Metal Solution  

Directions 

For 100 mL Total

1. To approximately 70 mL of dH2O, add each of the components in the order specified while
stirring continuously.
2. Bring total volume to 100 mL with dH2O.
3. Store at refrigerator temperature.
Note: CoCL2 can be interchancable with CoSO4.

# Component Amount Stock Solution Final Concentration

Concentration
Na2EDTA·2H2O (Sigma
1 0.1 g/100 mL   0.27 mM
ED255)
2 H3BO3 (Baker 0084) 0.114 g/100 mL   1.8 mM
FeCl3·6H2O (Sigma
3 4.9 mg/100 mL   0.018 mM
   1513)
MnSO4·H2O (Sigma
4 16.4 mg/100 mL   0.097 mM
M8179)
ZnSO4·7H2O (Sigma Z
5 2.2 mg/100 mL   0.007 mM
0251)
CoCl2·6H2O (Sigma C-
6 0.48 mg/100 mL   0.002 mM
3169)
Chelated Iron Solution  

Directions 

This solution requires the combination of two liquid components:

Component A:
1. Pour approximately 450 mL dH2O into a beaker. Heat the beaker in the microwave oven to
near boiling.
2. Add 10.0 g of Na2EDTA to the stirring beaker.
3. After the EDTA fully dissolves, bring the total volume to 500 mL with dH2O.
Component B:
1. Pour approximately 450 mL 0.1 M HCL into a beaker.
2. Add 0.81 g of FeC3.6H2O to beaker.
 3. Bring the total volume to 500 mL with dH2O.
Slowly add Component B to the heated Component A while stirring.

# Component Amount Stock Solution Final Concentration

Concentration
Na2EDTA·2H2O (Sigma
1 10 g/500 mL   26 mM
   ED255)
2 0.1M HCL 450 mL   0.1 M
FeCl3·6H2O (Sigma
3 0.81 g/500 mL   3 mM
1513)
Vitamin B12  

Directions 

For 200 mL Total

1. Prepare 200 mL of HEPES buffer (50 mM).
2. Adjust the pH to 7.8.
3. Add Vitamin B12 (0.1 mM) wait until fully dissolved.
4. Sterilize by 0.45 µm Millipore filter. Store in dark at freezer temperature.
** The amount of vitamins added can vary from medium to medium so the final concentration
is not listed.

# Component Amount Stock Solution Final Concentration

Concentration
HEPES buffer pH 7.8
1 2.4 g/200 mL dH2O    
   (Sigma H-3375)
Vitamin B12
2 (cyanocobalamin, (Sigma 0.027 g/200 mL dH2O  
V-6629)
Bristol Medium  

Directions 

H.C. Bold's modification of Bristol's recipe (Bold 1949). General purpose freshwater medium
and as bristol's solution, an essential component of other media--see Bold 1NV, Bold 3N,
Bristol-NaCl, LDM, Proteose, Soil extract, and Trebouxia.

For 1 L Total
1. To approximately 900 mL of dH2O add each of the components in the order specified while
stirring continuously.
2. Bring total volume to 1 L with dH2O.
*For 1.5% agar medium add 15 g of agar into the flask; do not mix.
3. Cover and autoclave medium.
4. Store at refrigerator temperature.

   # Component Amount Stock Solution Final Concentration

Concentration
 NaNO3 (Fisher BP360-
1 10 mL/L 10 g/400mL dH2O 2.94 mM
500)
CaCl2·2H2O (Sigma C-
2 10 mL/L 1 g/400mL dH2O 0.17 mM
3881)
MgSO4·7H2O (Sigma
3 10 mL/L 3 g/400mL dH2O 0.3 mM
230391)
4 K2HPO4 (Sigma P 3786) 10 mL/L 3 g/400mL dH2O 0.43 mM
5 KH2PO4 (Sigma P 0662) 10 mL/L 7 g/400mL dH2O 1.29 mM
6 NaCl (Fisher S271-500) 10 mL/L 1 g/400mL dH2O 0.43 mM
A+ Medium  

Directions 

For 1 L Total

Liquid media:
1. To approximately 900 mL of dH2O add the first 8 components in the order specified while
stirring continuously.
2. Bring total volume to 1 L with dH2O.
3. Cover and autoclave medium.
4. Add filter sterilized A+ Trace Components.
5. Allow to cool then store at refrigerator temperature.

Agar media:
1. To approximately 390 mL of dH2O add the first 8 components in the order specified while
stirring continuously.
2. Bring total volume to 500 mL with dH2O.
3. In a separate container add 15 g of agar to 500 mL of dH2O (final 1.5% w/v).
4. Cover and autoclave both solutions.
5. In a water bath allow both solutions to cool to 45-50 °C.
6. Add sterile Sodium Thiosulfate to agar solution and mix well.
7. Add filter sterilized A+ Trace Components.
8. Combine both agar and liquid solutions, mix well. Note the agar can solidify quickly.
9. Allow to cool then store at refrigerator temperature.

   # Component Amount Stock Solution Final Concentration

Concentration
1 NaCl (Fisher S271-500) 18 g/L   0.308 M
MgSO4·7H2O (Sigma
2 5 g/L   0.02 M
230391)
Na2EDTA·2H2O (Sigma
3 10 mL/L 0.3 g/100 mL 80.6 µM
ED255)
4 KCl (Fisher P 217) 10 mL/L 6 g/100 mL 8.05 mM
CaCl2·2H2O (Sigma C-
5 10 mL/L 3.7 g/100 mL 3.33 mM
3881)
NaNO3 (Fisher BP360-
6 10 mL/L 10 g/100 mL 11.8 mM
500)
 7 KH2PO4 (Sigma P 0662) 10 mL/L 0.5 g/100 mL 0.36 mM
8 Trizma Base pH 8.2 10 mL/L 10 g/100 mL 8.26 mM
A+ Trace Components
9 10 mL/L    
(sterilize before adding)
Sodium Thiosulfate
10 Pentahydrate (agar media 1 mL/L 24.819 g/100 mL 1 µM
only,sterile) (Baker 3946)
A+ Trace Components (sterilize before adding)  

Directions 

For 1 L Total
1. To approximately 900 mL of dH2O add the components and stock solutions in the order
specified while stirring continuously.
2. Bring total volume to 1 L with dH2O at refrigerator temperature, away from bright light.

# Component Amount Stock Solution Final Concentration

Concentration
1 H3BO3 (Baker 0084) 3.43 g/L   0.55 mM
2 ZnCl2 (Sigma Z-0152) 0.315 mg/L   2.31 µM
3 MoO3(85%) 0.03 mg/L   0.17 µM
Vitamin B12
4 (cyanocobalamin, (Sigma 0.004 mg/L   2.95 nM
V-6629)
  
FeCl3·6H2O (Sigma
5 1 mL 3.89 g/100 mL 24 µM
1513)
MnCl2·4H2O (Baker
6 1 mL/L 4.3 g/100 mL 34.2 µM
2540)
CuSO4·5H2O (MCIB
7 1 mL/L 0.003 g/100 mL 18.8 nM
3M11)
CoCl2·6H2O (Sigma C-
8 1 mL/L 0.012 g/100 mL 9.4 nM
3169)