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29. PFENNIG'S MEDIUM II (modified 1988, for green sulfur bacteria)

Culture medium recipe

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* Modified for Chlorobaculum thiosulfatiphilum DSM 249
beta

Anaerobic in light

Solution occurs in 2 other media (first defined in medium 29 )
Equipment needed: Ice bath Filter
# Compound Amount Unit Conc. [g/L] Conc. [mM]
2 Solution A 460 ml - -
3 Solution B 135 ml - -
4 Solution C 50 ml - -
5 Solution D 450 ml - -
6 Solution E 26 ml - -
7 Solution F 100 ml - -
Instructions:
  1. Mix solution D, C and E. Bubble with CO2 in an ice bath under sterile conditions.
  2. Fill 50 ml in each bottle of solution A. Before using add 4 ml solution B and 0.1 ml solution F.
  3. Adjust the pH with filter-sterilised 1M Na2CO3 to 6.8 -7.1.
  4. Fill in sterile, N2 gassed screw-cap tubes under N2 gas.
  5. During the first 24 h, the iron of the medium precipitates in the form of black flocks. No other sediment should arise in the otherwise clear medium. Feed periodically with neutralized 3% solution of sodium sulfide to replenish sulfide and with other supplement solutions (see Ref. 3365).
Solution occurs in 2 other media (first defined in medium 29 )
Equipment needed: Autoclave
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 0.25 g 0.543 3.697
2 0.25 g 0.543 -
3 460.00 ml 998.207 55408.8
4 , for marine or estuarine isolates 100.00 g 217.391 3719.75
Instructions:
  1. Fill 10x 46ml in 100 ml screw-cap bottles. Bubble with N2/CO2 and autoclave 121°C 15 min.
  2. (For marine or estuarine isolates add 100.0 g NaCl to this solution and increase the MgSO4 x 7 H2O to 15.0 g).
Solution occurs in 2 other media (first defined in medium 29 )
Equipment needed: Autoclave
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 2 g 14.815 61.682
2 135 ml 998.207 55408.8
Instructions:
  1. Prepare in a screw-cap bottle, bubble with N2 to replace air, close tightly and autoclave.
Solution occurs in 2 other media (first defined in medium 29 )
Equipment needed: Filter
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 1.5 g 30 357.115
2 50.0 ml 998.207 55408.8
Instructions:
  1. Bubble with CO2 and filter sterilize into sterile, gas-tight, 100 ml screw-cap bottle.
Solution occurs in 2 other media (first defined in medium 29 )
Equipment needed: Autoclave Ice bath
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1
(0,1%)
0.5 ml - -
2 450.0 ml 998.207 55408.8
Instructions:
  1. Autoclave in a cotton-stoppered Erlenmeyer flask with an outlet tube for medium, connected to a glass outlet at the bottom of the vessel and has, at the other end, a silicon rubber tube with a pinch cock and a bell for aseptic dispensing of the medium into bottles.
  2. Cool to room temperature under an atmosphere of N2/CO2 in an ice bath.
Solution occurs in 2 other media (first defined in medium 29 )
Equipment needed: Filter
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 0.35 g 13.462 251.659
2 0.25 g 9.615 124.741
3 0.25 g 9.615 87.378
4 0.25 g 9.615 53.373
5 0.50 g 19.231 78.023
6 0.35 g 13.462 180.568
7 0.35 g 13.462 98.92
9 Trace element solution SL-10 B 1.00 ml - -
10 25.00 ml 959.814 53277.7
Instructions:
  1. Filter sterilize into sterile, gas-tight, 100 ml screw-cap bottle.

Solution F #

1108
Solution occurs in 2 other media (first defined in medium 29 )
Equipment needed: Filter
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 0.01 g 0.009 0.006
2 100.00 ml 90.091 5000.8
Instructions:
  1. Filter sterilized
Solution occurs in 2 other media (first defined in medium 29 )
Equipment needed: Autoclave
# Compound Amount Unit Conc. [g/L] Conc. [mM]
1 100 ml 998.207 55408.8
2 3 g 30 124.905
Instructions:
  1. The sulfide solution is prepared in a 250 ml screw-capped bottle with a butyl rubber septum and a magnetic stirrer. The solution is bubbled with nitrogen gas, closed and autoclaved for 15 min. at 121°C. After cooling to room temperature the pH is adjusted to about 7.0 by adding of sterile 2 N H2SO4 drop-wise with a syringe without opening the bottle.
  2. Appearance of a yellow colour indicates the drop of pH to about 8. The solution should be stirred continuously to avoid precipitation of elemental sulfur. The final solution should be clear and is yellow in colour.
# Compound Amount Unit Conc. [g/L] Conc. [mM]
2 1000.0 ml 998.207 55408.8
3
(25%)
7.7 ml 2.441 66.946
4 1.5 g 1.5 5.395
5 70.0 ml 203.7 1494.55
6 100.0 ml - -
7 300.0 ml 640.8 10363.4
8 190.0 ml - -
9 2.0 ml - -
10 24.0 ml - -
11 36.0 ml - -

Metadata

Media status: Official DSMZ medium
Source: DSMZ
Taxonomic range:  Bacteria
Medium type: Complex medium
Final pH: unknown
Equipment needed: Autoclave, Filter, Ice bath

Associated strains

All strains for this medium: 11
Strains with modifications: 11
Chlorobaculum thiosulfatiphilum DSM 249

Molecular composition

Compound Conc. [g/L] Conc. [mM]
Distilled water 923.832 51280.4
NaCl 81.9 1401.38
Na2S x 9 H2O 1.638 6.82
NaHCO3 1.229 14.624
H3BO3 0.525 8.488
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Cultivation metadata from 11 strains BacDive

phyla 1 11 strains
nutrition phototroph 3 strains
oxygen anaerobe 11 strains
oxygen obligate anaerobe 3 strains
temperature mesophilic 10 strains
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