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DSMZ

CCAP 89 . SBBM

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ml
Compound Amount Unit
Medium C14 970 ml
SE2 (Soil Extract 2) 30 ml
1 Used for some filamentous green algae e.g. Trentepohlia, can give better growth than 3N-BBM+V (Mail sol. C 14) or JM:SE (Mail sol. C 41). Make both media and sterilise, then mix aseptically in the ratio of 97% 3N-BBM+V and 3% SE2.

Mail sol. C14 #

1000
Compound Amount Unit
NaNO3
(75 g/l stock solution) 		10 ml
CaCl2 x 2 H2O
(2.5 g/l stock solution) 		10 ml
MgSO4 x 7 H2O
(7.5 g/l stock solution) 		10 ml
K2HPO4 x 3 H2O
(7.5 g/l stock solution) 		10 ml
KH2PO4
(17.5 g/l stock solution) 		10 ml
NaCl
(2.5 g/l stock solution) 		10 ml
Trace Elements (PIV) 6 ml
1 Once elements are dissolved autoclave at 15 psi for 15 minutes.
Vitamin B1 solution 1 ml
Vitamin B12 solution 1 ml
Agar , for solid medium 15 g
2 Make up to 1 litre with distilled water. For agar add 15 g per litre Bacterialogical Agar. Autoclave at 15 psi for 15 minutes.

Vitamin B1 solution #

100
Compound Amount Unit
Thiamine hydrochloride
(Vitamin B1) 		0.12 g
1 Filter sterile

Trace Elements (PIV) #

1000
1 Ensure elements are added in the following sequence:
Compound Amount Unit
Na2-EDTA 0.750 g
FeCl3 x 6 H2O 0.097 g
MnCl2 x 4 H2O 0.041 g
ZnCl2 0.005 g
CoCl2 x 6 H2O 0.002 g
Na2 MoO4 x 2 H2O 0.004 g
2 Once elements are dissolved autoclave at 15 psi for 15 minutes.

Vitamin B12 solution #

100
Compound Amount Unit
Cyanocobalamine
(Vitamin B12) 		0.1 g
1 Take 1 ml of this solution and add 99 ml Deionised water. Filter sterile.

SE2 (Soil Extract 2) #

660
Compound Amount Unit
Soil
(air­dried, sieved) 		105 g
Deionized water 660 ml
1 Site selection for a good soil is very important and for most purposes a soil from undisturbed deciduous woodland is best. Sites to avoid are those showing obvious signs of man's activity and particular care should be taken to avoid areas where fertilizers, crop sprays or other toxic chemicals may have been used.
2 A rich loam with good crumb structure should be sought. Stones, roots and larger invertebrates should be removed during an initial sieving through a 1 cm mesh. The sieved soil should be spread to air dry and hand picked for smaller invertebrates and roots. It should be turned periodically and picked over again. When dry it may be sieved through a finer mesh (2­4 mm) or stored as it is prior to use.
3 Soil is prepared as above. 105 g of air­dried sieved soil and 660 ml of deionized water are placed in a 1 litre bottle and autoclaved once at 15 psi for 15 minutes, then again after 24 hours. The contents of the bottle are left to settle (usually for at least a week) and then the supernatant is decanted and filtered. The final pH should be 7.0 - 8.0.

Mail sol. C41 #

1000
Compound Amount Unit
Ca(NO3)2 x 4 H2O
(4.0 g/200 ml stock solution) 		0.7 ml
KH2PO4
(2.48 g/200 ml stock solution) 		0.7 ml
MgSO4 x 7 H2O
(10.0 g/200 ml stock solution) 		0.7 ml
NaHCO3
(3.18 g/200 ml stock solution) 		0.7 ml
Solution 1 0.7 ml
Solution 2 0.7 ml
Solution 3 0.7 ml
NaNO3
(16.0 g/200 ml stock solution) 		0.7 ml
Na2HPO4 x 12 H2O
(7.2 g/200 ml stock solution) 		0.7 ml
SE2 (Soil Extract 2) 300.0 ml
Deionized water 1000.0 ml
Agar , for solid medium 15.0 g
1 This recipe is a 7:3 mixture of JM and SE2. Make up to 1 litre with deionized water. For agar, add 15.0 g per litre of Bacteriological Agar (Thermo Scientific™ Oxoid™ Agar No.1). Autoclave at 15 psi for 15 minutes.

Solution 1 #

200
Compound Amount Unit
FeNa-EDTA 0.45 g
Na2-EDTA 0.45 g

Solution 2 #

200
Compound Amount Unit
H3BO3 0.496 g
MnCl2 x 4 H2O 0.278 g
(NH4)6Mo7O24 x 4 H2O 0.200 g

Solution 3 #

200
Compound Amount Unit
Cyanocobalamine 0.008 g
Thiamine HCl 0.008 g
Biotine 0.008 g

SE2 (Soil Extract 2) #

1000
Compound Amount Unit
Soil
(air-dried) 		trace
Distilled water trace
1 Site selection for a good soil is very important and for most purposes a soil from undisturbed deciduous woodland is best. Sites to avoid are those showing obvious signs of man's activity and particular care should be taken to avoid areas where fertilizers, crop sprays or other toxic chemicals may have been used.
2 A rich loam with good crumb structure should be sought. Stones, roots and larger invertebrates should be removed during an initial sieving through a 1 cm mesh. The sieved soil should be spread to air dry and handpicked for smaller invertebrates and roots. It should be turned periodically and picked over again. When dry sieve through a finer mesh (2-4 mm) and store in an airtight container away from light and heat.
3 Soil is prepared as above. Air-dried soil and twice its volume of supernatant distilled water are autoclaved together at 15 psi for 2 hours and left to cool. The supernatant is then decanted and then distributed to containers in volumes suitable for making up batches of media. The aliquots and their containers are autoclaved for an appropriate length of time (e.g. 1 litre or less for 15 minutes) and are then kept in a cool place (e.g. a refrigerator) until required.