Chromatium Medium (ATCC Medium 2010)

[所属分类:培养基配方] [发布时间:2020-4-28] [发布人:] [阅读次数:] [返回]

山东拓普生物工程有限公司

Shandong Tuopu Biol-Engineering Co.,Ltd

Chromatium Medium
(ATCC Medium 2010)

培养基配方
Composition per 127.0mL:
Solution 1................................................................76.2mL
Solution 2 + Solution 3...................................................44.8mL
Solution 4.................................................................6.0mL
Solution 1:
Composition per 2.5L:
NaCl......................................................................125.0g
CaCl2 ......................................................................2.0g
Preparation of Solution 1: Add NaCl and CaCl2 to distilled/deion-
ized water and bring volume to 2.5L. Distribute in 76.2mL volumes into
127.0mL screw-capped bottles. Autoclave for 15 min at 15 psi pressure–
121°C.
Solution 2:
Composition per 100.0mL:
Sodium ascorbate............................................................2.4g
KC1.........................................................................1.0g
KH2PO4 .....................................................................1.0g
MgCl2 ·6H2O ...............................................................0.8g
NH4Cl ......................................................................0.8g
Heavy metal solution......................................................50.0mL
Vitamin solution..........................................................15.0mL
Vitamin B 12 solution......................................................3.0mL
Preparation of Solution 2: Add components to distilled/deionized
water and bring volume to 100.0mL. Mix thoroughly.
Solution 3:
Composition per 900.0mL:
NaHCO3 .....................................................................4.5g
Preparation of Solution 3:  Add NaHCO3 to distilled/deionized
water and bring volume to 900.0mL. Mix thoroughly. Bubble 100%
CO2 through the solution for 30 min. After CO2 saturation of solution
3, add solution 2 and immediately filter the mixture through a Seitz fil-
ter (or a Millipore) using positive CO2 pressure to push the liquid
through.
Solution 4:
Composition per 200.0mL:
Na2S·9H2O..................................................................3.0g
Preparation of Solution 4: Add Na2S·9H2O to distilled/deionized
water and bring volume to 200.0mL. Add a magnetic stir bar to the
flask. Autoclave for 15 min at 15 psi pressure–121°C. On a magnetic
stirrer, slowly add 2.0mL of sterile 2M H2SO4 . This partially neutraliz-
es the solution. The solution should turn yellow. H2S gas will be liber-
ated. Neutralization and distribution of the solution should be done as
rapidly as possible under adequate ventilation.
Heavy Metal Solution:
Composition per liter:
Ethylenediamine tetraacetate (EDTA)..........................................1.5g
FeSO4 ·7H2O.................................................................0.2g
ZnSO4 ·7H2O.................................................................0.1g
MnCl2 ·4H2O................................................................0.02g
Modified Hoagland trace elements solution...................................6.0mL
Preparation of Heavy Metal Solution: Dissolve EDTA in ap-
proximately 800.0mL of distilled/deionized water. Add remaining
components. Bring volume to 1.0L with distilled/deionized water. Mix
thoroughly.
Vitamin B 12  Solution:
Composition per 100.0mL:
Vitamin B 12  (cyanocobalamin) ..............................................2.0mg
Preparation of Vitamin B 12  Solution: Add vitamin B 12 to dis-
tilled/deionized water and bring volume to 100.0mL. Mix thoroughly.
Vitamin Solution:
Composition per 100.0mL:
Pyridoxamine·2HCl ...........................................................5.0mg
Nicotinic acid................................................................2.0mg
Thiamine......................................................................1.0mg
Pantothenic acid..............................................................0.5mg
Biotin .......................................................................0.2mg
p-Aminobenzoic acid...........................................................0.1mg
Preparation of Vitamin Solution:  Add components to distilled/
deionized water and bring volume to 100.0mL. Mix thoroughly.
Modified Hoagland Trace Elements Solution:
Composition  per 3.6L:
H3BO3 .........................................................................11.0g
MnCl2 ·4H2O....................................................................7.0g
AlCl3 ..........................................................................1.0g
CoCl2 ..........................................................................1.0g
CuCl2 ..........................................................................1.0g
KI..............................................................................1.0g
NiCl2 ..........................................................................1.0g
ZnCl2 ..........................................................................1.0g
BaCl2 ..........................................................................0.5g
KBr ............................................................................0.5g
LiCl............................................................................0.5g
Na2MoO4 ........................................................................0.5g
SeCl4 ..........................................................................0.5g
SnCl2 ·2H2O....................................................................0.5g
NaVO3 ·H2O.....................................................................0.1g
Preparation of Modified Hoagland Trace Elements Solu-
tion: Prepare each component as a separate solution. Dissolve each
salt in approximately 100.0mL of distilled/deionized water. Adjust the
pH of each solution to below 7.0. Combine all the salt solutions and
bring the volume to 3.6L with distilled/deionized water. Adjust the pH
to 3–4. A yellow precipitate may form after mixing. After a few days,
it will turn into a fine white precipitate. Mix the solution thoroughly be-
fore using.
Preparation of Medium: To the 80.0mL of sterile solution 1 in
screw-capped bottles, add combined solutions 2 and 3 immediately af-
ter filtration and fill bottles to capacity. Mix thoroughly. Aseptically re-
move 6.0mL of the medium from the bottles and replace it with 6.0mL
of neutralized solution 4. Let stand for 24 hr. The medium should form
a fine white precipitate before using. To inoculate, remove 6.0mL of
the completed medium from the bottles and replace it with 6.0mL of
inoculum.
Use: For the cultivation of Ectothiorhodospira mobilis and Ectothior-
hodospira marismortui.