CH5. Types of Culture

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CH5. Types of Culture
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In vitro of explant

• The non-dividing, differentiated, quiescent cells
  of the explant when grown on a nutrient medium
  first undergo changes to achieve the meristematic
  state.

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Callus

• An amorphous mass of loosely arranged thin-walled
  parenchyma cells arising from the proliferating
  cells of the parent tissue cultured on agar
  medium.

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Cellular totipotency

• Cytodifferentiation
• Cell differentiation, mainly emphasis on vascular
  differentiation, tracheary element
  differentiation, etc.
• Dedifferentiation
• The phenomenon of mature cells reverting to a
  meristematic state and forming undifferentiated
  callus tissue.
• Redifferentiation
• The ability of the component cells of the callus
  to differentiate into a whole plant or organ.

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Types of culture
Embryo culture
Seed culture
Meristem culture
Cell culture
Plant tissue culture
Protoplast culture
Organ culture
Bud culture
Callus culture
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Seed Culture

• Seed culture is an important technique when
  explants are taken from in vitro-derived plants
  and in propagation of orchids.
• In nature germination of orchid seedlings id
  dependent on a symbiotic relationship with a
  fungus.
• In vitro, it is possible to independent on
  asymbiotic germination.

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Embryo culture

• Mature embryo culture
• Seed dormancy ex. ripe seeds
• Immature embryo culture / embryo rescue
• To avoid embryo abortion
• Ex. Hybrid embryo.

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Application of embryo culture

• Prevention of embryo abortion in wide crosses.
• Production of haploids
• Overcoming seed dormancy
• Shortening of breeding cycle
• Prevention of embryo abortion with early ripening
  stone fruit
• In vitro clonal propagation

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Callus formation
De-differentiation
Re-differentiation
Explants Callus    
Protoplasts   Development  
Suspension cells   Organs
  (leaves, roots, shoots, flowers,...)  
?
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Callus formation
Stimuli In vivo wound, microorganisms,
insect feeding   In vitro Phytohormones
  1. Auxin   2. Cytokinin   3.
Auxin and cytokinin   4. Complex natural
extracts    
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Callus formation
Stimuli In vitro Phytohormones   1.
Auxin   2. Cytokinin   3. Auxin and
cytokinin   4. Complex natural extracts    
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  Auxins 0.01 - 10 mg / L
Activity   IAA   IBA
  NAA   2,4 - D   2,4,6 - T   Dicamba
Picloram   
Cytokinins 0.01 - 10 mg / L    TDZ
BA (BAP) Kinetin    2 -
ip   Zeatin   
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Callus formation
Stimuli In vitro Phytohormones   1.
Auxin  
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Callus formation
Stimuli In vitro Phytohormones   1.
Auxin   2. Cytokinin   3. Auxin and
cytokinin   4. Complex natural extracts    
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Callus formation
Stimuli In vitro Phytohormones   1.
Auxin   2. Cytokinin   3. Auxin and
cytokinin   4. Complex natural extracts    
Coconut endosperm ( milk / water ) Yeast extract
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Callus growth   Growth characteristics of callus
    friable cultures   lignified and hard
cultures   
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Callus growth   Growth characteristics of callus
    pigmentation yellowish,   white,
  green,   pigmented, 
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Maintenance Subculture sub-division of a
culture for transfer to fresh medium.  
?
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Q How?
Q Where?
Medium
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Q How?
Q Where?
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Q When?
growth
time
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Q What happened in long-term cultures?
Physiological condition