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In Vitro Culture Techniques : The Biotechnological Principles

 
     
 
Somatic Embryogenesis
Embryo production is a characteristic feature of the flowering plants. However, such structures (embryoids) have also been artificially induced in cultured plant tissues, besides zygote. Somatic embryogenesis was first induced in suspension culture (Stewart et al. 1958) and callus culture (Reinert, 1959) of carrot. More than 30 plant families are known so far where somatic embryoids have been induced (Raghavan, 1976; Ammirato, 1983).

  Events of somatic embryogenesis  
 

Fig. 8.4. Events of somatic embryogenesis

 

Somatic embryogenesis can be initiated in two ways : (i) by inducing embryogenic cells within the preformed callus, and (ii) directly from preembryonic determined cell, (without callus) which are ready to differentiate into embryoids (Sharp et al 1980). In the first case, ambryoids are initiated in callus from superficial cell aggregates where cells contain a large vacuole, dense cytoplasm, large starch granules and nucleus (Me Willian et al., 1974).

Two nutritional media of different composition are required to obtain embryoids. First medium contains auxin to initiate embryogenic cells. Second medium lacks auxin or reduced level of auxin is needed for subsequent development of the embryonic cells into embryoids and plantlets. In both the cases reduced amount of nitrogen is required (Ammirato, 1983). The embryogenic cells pass through 3 different stages e.g. globular, heart shaped and torpedo shaped, to form embryoids (Fig. 8.4).

These embryoids can be separated and isolated mechanically by using glassbeads. When embryoids reach torpedo stage they are transferred to filter paper bridge (a sterile and pluged culture tube containing about 10 ml MS liquid medium supplemented with Kinetin (0.2 mg/lit) and sucrose (2% W/V) on which Whatman No.1. Filter paper is placed to make a bridge (Dodds and Roberts, 1985). Some plants in which somatic embryogenesis has been induced in vitro are Atropa belladona, Brassica oleracea, Carica papaya, Coffea arabica, Citrus cinensis, Daucus carrota, Nicotiana tabacum, Pinus ponderosa and Saccharuni officinarum.

 

Content

Totipotency

Historical background

Requirements for cell and Tissue Cultures

 

A tissues culture laboratory

 

Nutrient media

 

 

Inorganic chemicals

 

 

Growth hormones

 

 

Organic constitutents

 

 

Vitamins

 

 

Amino acids

Culture of plant materials

 

Explant culture

 

Callus formation and its culture

 

Organogenesis

 

Root culture

 

Shoot culture and micropropagation

 

Cell culture

 

 

Benefits from cell culture

 

Somatic embryogenesis

 

Somaclonal variation

 

Protoplast culture

 

 

Isolation

 

 

Regeneration

 

Protoplast fusion and somatic hybridization

 

 

Fusion products

 

 

Method of somatic hybridization

 

Anther and pollen Culture

 

 

Culturing techniques

 

In vitro androgenesis (direct and indirect androgenesis)

 

Mentor pollen technology

 

Embryo culture

 

Embryo rescue

 

Protoplast fusion in fungi

 
     
 
 
     



     
 
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