155
APPENDIX 1
MEDIA PREPARATION
1
2
Algal culture medium
Reagents
Per Liter
NaNO3
1.00g
K2HPO4
0.25g
KgSO4
0.51g
NH4 Cl
0.05 g
CaCl2
0.01 g
FeCl3
0.003 g
Distilled water
1000ml
Algal proteose medium
Reagents
Per Liter
Proteose peptone
1g
Bristol’s solution
1 Lit.
Bristol’s solution (for 1000 ml)
NaNO3-
10g
K2HPO4
3g
KH2PO4
7g
156
CaCl2
1g
MgSO4.7H2O
3g
FeCl3 solution
0.1ml
FeCl3 solution (for 100 ml)
FeCl3
3
1g
BG-11 Medium
Component
Stock Solution
Quantity Used
(g · L-1 dH2O)
Fe Citrate solution
1 mL
Citric acid
6
1 mL
Ferric ammonium citrate
6
1 mL
NaNO3
—
1.5 g
K2HPO4·3H2O
40
1 mL
MgSO4·7H2O
75
1 mL
CaCl2·2H2O
36
1 mL
Na2CO3
20
1 mL
MgNa2EDTA·H2O
1.0
1 mL
__
1 mL
H3BO3
—
2.860 g
MnCl2·4H2O
—
1.810 g
ZnSO4·7H2O
—
0.220 g
CuSO4·5H2O
79.0
1 mL
Na2MoO4·2H2O
—
0.391 g
Co(NO3)2·6H2O
49.4
1 mL
Trace metals solution
157
4.
Bristol’s medium
Reagents
Per Liter
Macro nutrients
NaNO3-
250 mg
K2HPO4
75 mg
KH2PO4
175 mg
CaCl2
25 mg
NaCl
25 mg
MgSO4.7H2O
75 mg
Micro nutrients
5
FeCl3
0.3 mg
MnSO4.4H2O
0.3 mg
ZnSO4.7H2O
0.2 mg
H3BO3
0.2mg
CuSO4.5H2O
0.06 mg
George’s medium
Reagents
Per Liter
Peptone
15. 0 g
KNO3
1.00 g
KHPO4
0.20g
K2HPO4
0.02g
MgSO4 7H2O
0.02g
FeCl3
0.04 g
EDTA Mg or Ca
0.20 g
158
Oligo elements
6
H3BO3
2.9 mg
MnCl2. 2H2O
1.8 mg
ZnSO4. 7 H2O
0.2 mg
Na2 MoO4. 2 H2O
0.4 mg
CuSO4. 5 H2O
0.08 mg
Co (NO3)2. 6 H2O
0.05 mg
Distilled water
1000 ml
GO algal medium
Reagents
Per Liter
Soil water
100 ml
Peptone
15. 0 g
KNO3
2.05 g
KHPO4
0.25g
K2HPO4
0.07g
MgSO4 7H2O
0.02g
FeCl3
0.14 g
Soil water preparation
20 g of loamy soil was air dried and dissolved in 1000ml of distilled
water along with a pinch of CaCO3 was added to soil water prior to
sterilization.
The above contents were mix well and kept in a steaming condition
for 3 hours for two successive days. Collect the extract by filtering through
159
Whatman No.1 filter paper and sterilize the extract under 15 psi at 121°C
minutes in an autoclave for further use.
7
Potato glucose agar
Reagents
Per Liter
Potato
300.0g
Glucose
20.0g
Agar
15.0 g
Potatoes were sliced and kept in 500ml of boiling water for 30
minutes. These boiled potatoes were then strained through cheese cloth. The
volume was adjusted to one litre with distilled water and to this; 20 g glucose
and 15 g agar were added.
8
Trebouxia medium
Soil water
140 ml
Protease peptone
10g
Glucose
20g
Bristol’s solution
860ml
Bristol’s solution
To 940 ml of distilled water the following contents were added.
ml
Stock Solution
g/400ml
10
NaNO3
10.0
10
CaCl2 2 H2O
1.0
10
MgSO4 7 H2O
3.0
160
10
K2HPO4
7.0
10
KH2PO4
1.0
10
NaCl
1.2
10
Citric Acid
1.2
10
PIV Metal Solution
1.3
PIV Metal solution
To 1000ml, of disodium salt of was added and to which the
following contents were added.
EDTA
0.750g
FeCl3. 6H2O
97 mg
MnCl2. 4 H2O
41 mg
ZnCl2
5mg
CaCl2. 6 H2O
2 mg
Na2Mo O4. 2 H2O
4 mg
Soil water preparation
20 g of loamy soil was air dried and dissolved in 1000ml of distilled
water along with a pinch of CaCO3 was added to soil water prior to
sterilization.
The above contents were mix well and kept in a steaming condition
for 3 hours for two successive days. Collect the extract by filtering through
Whatman No.1 filter paper and sterilize the extract under 15 psi at 121°C
minutes in an autoclave for further use.
161
9
Bold’s Basal Medium (BBM)
Component
Stock Solution
Quantity Used
(g · L-1 dH2O)
Macronutrients
NaNO3
25.00
10 mL
CaCl2 · 2H2O
2.50
10 mL
MgSO4 · 7H2O
7.50
10 mL
K2HPO4
7.50
10 mL
KH2PO4
17.50
10 mL
NaCl
2.50
10 mL
1 mL
Alkaline EDTA Solution
EDTA
50.00
KOH
31.00
1 mL
Acidified Iron Solution
FeSO4·7H2O
4.98
H2SO4
1 mL
Boron Solution
1 mL
H3BO3
11.42
162
1 mL
Trace Metals Solution
10
ZnSO·7HO
8.82
MnCl2·4H2O
1.44
MoO3
0.71
CuSO4·5H2O
1.57
Co(NO3)2·6H2O
0.49
Dy-III Medium
Component
Stock Solution
Quantity Used
(g · L-1 dH2O)
MES buffer
___
200 mg
CaCl2 · 2H2O
75.00
1 mL
Na2SiO3 · 9H2O
15.00
1 mL
MgSO4 · 7H2O
50.00
1 mL
NH4NO3
5.00
1 mL
NaNO3
20.00
1 mL
KCl
3.00
1 mL
Na2 β-glycerophosphate
10.00
1 mL
H3BO3
4.58
1 mL
Trace metals solution
___
1 mL
Vitamins solution
___
1 mL
163
Trace Metals Solution
Component
1° Stock Solution
Quantity used
(g · L-1 dH2O)
Na2EDTA (anhydrous)
—
8.000 g
Fe (iron wire)
__
0.700 g
MnCl2 · 4H2O
__
0.720 g
ZnSO4 · 7H2O
__
0.176 g
Na2MoO4 · 6H2O
50
1 mL
CoCl2 · 6H2O
29
1 mL
Vitamins Solution
Component
1° Stock Solution
Quantity used
(g · L-1 dH2O)
11
Thiamine · HCl
—
200 mg
Biotin
0.5
1 mL
Cyanocobalamin
0.5
1 mL
Aaronson Medium Composition
Reagents
Per Liter
NH4Cl
0.5 g
KH2PO4
0.3 g
MgSO4.7H2O
1.0 g
164
MgCO3
0.4 g
C6H9NO6 (Nitrilotriacetic acid)
0.2 g
CaCO3
0.05 g
C5H9NO4 (Glutamic acid)
3.0 g
Glucose
10.0 g
Arginine
0.4 g
Hystidine monochloride
0.4 g
Biotin (Vitamin H)
0.00001 g
Thiamine HCl (Vitamin B1)
0.001 g
Microelement stock solution
10 ml
Microelement Stock Solution
Fe(NH4)2 (SO4)2.6H2O
0.5 g
H3BO3
0.025 g
MnSO4.H2O
0.125 g
CoSO4.7H2O
0.025 g
CuSO4.5H2O
0.020 g
ZnSO4.H2O
0.25 g
(NH4)6Mo7O24.4H2O
0.0125 g
Na3VO4.16H2O
0.025 g
165
12
13.
Cramer and Myers Medium
Reagents
Per Liter
(NH4)2HPO4
1g
KH2PO4
1g
MgSO4.7H2O
0.2 g
CaCl2
0.02 g
Na3C6H5O7.2H2O
0.8 g
Fe2(SO4)3. nH2O
3mg
MnCl2.4H2O
1.8 mg
CoSO4.7H2O
1.5 mg
ZnSO4.7H2O
0.4 mg
Na2MoO4.2H2O
0.2 mg
CuSO4.5H2O
0.02 mg
H3BO3
2.48 mg
Thiamine HCl
0.5 mg
Cyanocobalamin
0.02 mg
Glucose
10 g
Beijerinck Medium
Reagents
Per Liter
Stock I
100 ml
Stock II
40 ml
166
Stock
III 60 ml
Micronutrients
1 ml
NH4NO3
1.5 g
K2HPO4
0.2 g
MgSO4 7H2O
0.2 g
CaCl2.2H2O
0.1 g
KH2PO4
9.07 g
K2HPO4
11.61 g
Stock I
Stock II
Stock III
Micronutrients
H3BO3
10 g
MnCl2.4H2O
5g
EDTA
50 g
CuSO4.5H2O
1.5 g
ZnSO4.H2O
22 g
CoCl2.6H2O
1.5 g
FeSO4.7H2O
5g
(NH4)6Mo7O24.4H2O
1g