Typically, calluses are induced by the placement of nicked tissue between a solid
(such as agar, carrageenan, or dampened filter paper) and air saturated with seawater vapor. Callus induction has been performed successfully for many seaweeds.
Table 6.4 shows some of the important varieties of red, brown, and green algae for
which tissue culturing has been reported.
In this section, a large brown alga will be used as an example for tissue culturing.
The following is the sequence followed by Sega et al. using the kelp Laminaria
angustata (Saga and Sakai 1982).
(1) Thoroughly rinse both ends of section from seaweed stalk and cut to lengths of
around 5 cm.
(2) Soak both ends of the fragment in 100% ethanol and heat slightly with alcohol
lamp before cutting to around 5 mm each.
(3) Bore with cork borer (approximate 4 mm in dimeter) and cut to thickness of
around 2 mm.
(4) Introduce the resulting disk-shaped fragments into ASP 12 NTA agar medium
(50 ml).
Table 6.1 Developing crude seawater solution for major seaweeds
ES
a
PES
b
PESI
b
ESS
c
Distilled water
–
100 ml
100 ml
60 ml
Seawater
100 ml
–
–
–
Sodium nitrate (NaNo 3 )
10 mg
350 mg
350 mg
350 mg
Na 2 Áglycerol phosphate
–
50 mg
50 mg
80 mg
Disodium phosphate (Na 2 HPO 4 .12 H 2 O)
2 mg
–
–
–
Iron (as EDTA, mole ratio 1:1)
–
2.5 mg
2.5 mg
–
Iron sequestrene
–
–
–
40 mg
Mixed metal solution PII
d
–
25 ml
25 ml
40 ml
Vitamin B 12
–
10 µg
–
–
Thiamine
0.5 mg
–
–
Biotin
5 µg
–
–
ESS vitamin mixed solution
e
–
–
–
1 ml
Soil extract
5 ml
–
–
–
Tris
–
500 mg
500 mg
–
HEPES
–
–
–
1 g
Potassium iodide (KI)
–
–
100 µg
–
pH
–
7.8
7.8
7.8
a ES solution should be diluted after sterilization
b
2 ml added to 100 ml of seawater after filtering of undiluted PES, PESI solution
c 1 ml added to 100 ml of seawater after filtering of undiluted ESS solution
d
See note on Table 6.2
e Mixing of ESS vitamin solution (in 1 ml): 10 lg vitamin B 12 , 10 lg biotin, 1 mg thiamine
hydrochloride, 1 mg nicotinic acid, 1 mg calcium pantothenate, 100 lg p-aminobenzoic acid,
10 mg inositol, 1 mg thymine
150
6 Seaweed Biotechnology
(such as agar, carrageenan, or dampened filter paper) and air saturated with seawater vapor. Callus induction has been performed successfully for many seaweeds.
Table 6.4 shows some of the important varieties of red, brown, and green algae for
which tissue culturing has been reported.
In this section, a large brown alga will be used as an example for tissue culturing.
The following is the sequence followed by Sega et al. using the kelp Laminaria
angustata (Saga and Sakai 1982).
(1) Thoroughly rinse both ends of section from seaweed stalk and cut to lengths of
around 5 cm.
(2) Soak both ends of the fragment in 100% ethanol and heat slightly with alcohol
lamp before cutting to around 5 mm each.
(3) Bore with cork borer (approximate 4 mm in dimeter) and cut to thickness of
around 2 mm.
(4) Introduce the resulting disk-shaped fragments into ASP 12 NTA agar medium
(50 ml).
Table 6.1 Developing crude seawater solution for major seaweeds
ES
a
PES
b
PESI
b
ESS
c
Distilled water
–
100 ml
100 ml
60 ml
Seawater
100 ml
–
–
–
Sodium nitrate (NaNo 3 )
10 mg
350 mg
350 mg
350 mg
Na 2 Áglycerol phosphate
–
50 mg
50 mg
80 mg
Disodium phosphate (Na 2 HPO 4 .12 H 2 O)
2 mg
–
–
–
Iron (as EDTA, mole ratio 1:1)
–
2.5 mg
2.5 mg
–
Iron sequestrene
–
–
–
40 mg
Mixed metal solution PII
d
–
25 ml
25 ml
40 ml
Vitamin B 12
–
10 µg
–
–
Thiamine
0.5 mg
–
–
Biotin
5 µg
–
–
ESS vitamin mixed solution
e
–
–
–
1 ml
Soil extract
5 ml
–
–
–
Tris
–
500 mg
500 mg
–
HEPES
–
–
–
1 g
Potassium iodide (KI)
–
–
100 µg
–
pH
–
7.8
7.8
7.8
a ES solution should be diluted after sterilization
b
2 ml added to 100 ml of seawater after filtering of undiluted PES, PESI solution
c 1 ml added to 100 ml of seawater after filtering of undiluted ESS solution
d
See note on Table 6.2
e Mixing of ESS vitamin solution (in 1 ml): 10 lg vitamin B 12 , 10 lg biotin, 1 mg thiamine
hydrochloride, 1 mg nicotinic acid, 1 mg calcium pantothenate, 100 lg p-aminobenzoic acid,
10 mg inositol, 1 mg thymine
150
6 Seaweed Biotechnology