PLANT TISSUE CULTURE

CALLUS CULTURE

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 Rice seed

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 Callus Induction/Multiplication Medium
Murashige and Skoog (MS,1962)

3 gr/l Casein Hydrolysate

37.35 mg/l Fe-EDTA

10 µM 2,4-D

2 µM BAP

Dark Culture

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 Callus Type
1. Development/Embryogenic callus, i.e. callus which is
capable of developing into either somatic organogenesis
or somatic embryogenesis. This type of callus has
chlorophyll, green, hard, and fast growing.
2. Proliferative callus, it grows very fast and results in the
production of a massive number of small cells with less
protoplasm content
3. Senescence callus, a very slow growing callus or even
not showing any development symptoms. It has no
chlorophyll, creamy in color, watery, slow growth, and
has a polyhedral cell form.

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 Callus Type
1. Embryogenic
2. Proliferative
3. Senescence

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 Rice callus growth curve

3000

2500
Fresh weight (mg)

2000
A
B
1500 C
D
1000 E
500

0
0 3 6 9 12 15 18 21 24 27 30 33 36 39 42
Time of culture (day)
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 Rice callus relative growth curve (ln)

0.15

0.10 A
B
RGR (ln)

C
0.05 D
E

0.00
3 6 9 12 15 18 21 24 27 30 33 36 39 42
-0.05
Time of culture (day)

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 Callus regeneration

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 Medium
MS

30 gr/l Maltose

0.3 gr/l Casen Hydrolysate

30 M BAP

0.625 % Phytagel

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 Plant regeneration : the production of
plantlets in sterile culture

Embryo culture

Somatic
embryogenesis

Somatic
organogenesis
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 Somatic embryogenesis : the
prod
•is independent bipolar structure
• is not physically attached to the
tissue of origin

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Organogenesis : s generated from cultured
tips, and their subseqthe formation and
outgrowth of shoot from callus or initiation
and outgrowth of axillary buduent adventitious
rooting :
• shoot is unipolar structure
• physically connected to the tissue of origin
• occasionally roots may give rise to shoots

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 Direct The formation of an asexual
embryo from a single cell or group
of cells as a part of the explant
tissue without an intervening
callus phase

Somatic
Embryogenesis

Consists of establishing an explant

in culture, subsequent proliferation
of callus and initiation of proembryo
and their subsequent development
Indirect to form bipolar embryo

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 Somatic Embryo Development

• Embryogenic Calli
• Globular Stage
• Heart Shape
• Torpedo-like Structure
• Cotyledonary stage

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 Warna embrio berhubungan dengan perkembangan embrio
somatik, misalnya embrio berwarna putih pada karet
berkembang menjadi planlet, sedangkan embrio hijau akan
membentuk kalus dan menghasilkan kalus embriogenik lewat
embriogenesis sekunder

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 Factors :
Genotype

Explant Age

Position

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 Media Type Auxin

PGR’s Sitokinin

Media Carbon
GA3
Source

Type
Gelling Agent
Concentration

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 Light irradiance

Environmental Culture
Factors Periode/timing

Temperature

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 Shoot and Plantlet Development

• Low Phytagel

• High Cytokinin
• No Auxin

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 Rooting

• Liquid Medium
• Filter paper bridge
• High Auxin (NAA)
• Low Cytokinin
• Double-magenta-pots
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 Acclimatisation

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 1. Hydroponics system

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 2. Soil system

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