Organogenesis,Shoot Buds,Skoog,Miller,Cultured Tissues,Tobacco Callus,Meristemoids

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Home >> Organogenesis and Embryogenesis >>Organogenesis

Organogenesis and Embryogenesis
Organogenesis

	Organogenesis - Organogenesis is an outcome of the process of dedifferentiation followed by redifferentiation of cells. Dedifferentiation favors unorganized cell growth and the resultant developed callus has meristems randomly divided. Most of these meristems, if provided appropriate in vitro conditions, would redifferentiate shoot buds and roots. As early as 1939 it was observed by White that by submerging callus from a tobacco hybrid in a stationary liquid medium, leafy buds and shoot like structures could be induced. He interpreted this to be the result of limited air supply. Shoot bud differentiation in cultured tissues is dependent on the auxin cytokinin ratio in the medium.
Skoog and his coworkers have done detailed and comprehensive work on this subject. Skoog and Miller (1957) rejected the concept of organ forming substances (rhizocaulines and caulocaulines) proposed by Went (1938) and instead suggested that organ formation is controlled by quantitative interaction (ratio rather than absolute concentration) of substances in growth and development. Cytokinin (adenine or kinetin) in the medium leads to the promotion of bud differentiation and development. Kinetin is 30,000 times more potent than adenine. A relatively high concentration of auxin favors cell proliferation and root differentiation while higher levels of cytokinin promote bud differentiation.
Thus root and shoot differentiation is a function of quantitative interaction between auxin and cytokinin. Casein hydrolysate or tyrosine also induces kinetin type bud formation even in the presence of higher levels of IAA in the medium. The requirement of exogenous auxin and cytokinin in the process of bud differentiation varies with the tissue system and apparently depends on the endogenous levels of the two hormones in the tissue. Polyamines also have been shown to be associated with induction of cell division, growth and differentiation of bacterial, animal and plant cells.
Other cytokinins which influence the induction of shoot-buds include BAP, 2-IP, 6-tetrahydropyraneadenine (SD 8339) and zeatin. Light intensity plays an important role in organogenesis. High light intensity has been shown to be inhibitory for shoot-bud formation in tobacco. The quality of light also influences organogenesis. Blue light promotes shoot bud differentiation in tobacco callus while red light stimulates rooting. In general, maintenance of callus under alternating light and dark periods (15-16 h) may prove satisfactory for differentiation of shoots.
	Temperature also affects the callus growth and differentiation. Increase in temperature up to 33°C may be associated with rise in the growth of tobacco callus but for shoot-bud differentiation a lower temperature (18°C may be optimal. The physical state of the medium also influences shoot-bud differentiation. A medium solidified with agar favors bud formation although there are some reports about the development of leafy shoot-buds on cultures grown in a liquid medium. The genome and physiological state of the explant are other factors accounting for differentiation in cultures. Under conditions favoring unorganized growth, the meristems in a callus are random and scattered Transfer of callus pieces to conditions supporting organized growth leads to the formation of meristemoids (also termed nodules or growth centres).

The meristemoids are localized clusters of cambium like cells which may become vascularized due to the appearance of tracheidal cells in the center. These are the sites for organ formation in the callus and can form roots or shoots. The origin of roots or shoots is endogenous.

Typical crown gall cells show a complete lack of organogenesis but have a capacity for unlimited growth Independent of exogenous hormones in cultures. The bacterium Agrobacterium tumefaciens infects some plant species and induces inside the host tissue a special type of tumor (teratoma) called a crown gall.

Cells of these tumors possess a pronounced capacity to differentiate shoot-buds and leaves when cultured in vitro for unlimited periods. However, shoots derived from a teratoma are abnormal in growth and morphology.

Braun (1959) used the nurse culture technique to clone single cells grown in shake cultures from pieces of a crown gall and obtained calli differentiating into shoots. Tips (3-5 mm long) excised from these abnormal teratoma shoots were grafted onto the cut end of the stems of normal tobacco plants from which axillary buds had been removed.

The successful grafts were also observed to have abnormal shoots. Braun and Wood (1976) continued to graft these shoot tips onto normal tobacco plants until after successive generations of grafting they obtained shoots which appeared structurally, histologically and functionally normal.

Thus whole plants were recovered in cultures from crown gall tissue, which under natural conditions grows as an unorganized mass of cells.

Early investigators in attempting to grow callus tried to eliminate any organized development; they considered organogenesis an impediment for analysis of differentiation and growth. They saw a callus as possessing fewer internal variables, thereby permitting more rigorous control of external variables for studying organogenesis.

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