The phenomenon of morphogenesis, morphogenetic factors for in vitro regeneration

The phenomenon of morphogenesis, morphogenetic factors for in vitro regeneration:-

The phenomenon of morphogenesis:-

Introduction:- Biological organization of any life coordinated with several events as though a craftsman was moulding it according to a plan. In this process, the individual parts do not develop independently but all are knit together into an organised system. The biological science concerned with this dynamic and casual aspect of organic form is called "Morphogenesis".

> The derivation of this word is obvious, the origin of form. It attempts to expose the effects of

various factors and how these factors manifest an organic form in toto. "Morphogenesis", a

distinctive aspect of organization of life, is the crossroad where all the highways of biological

exploration tend to converge", says Sinnott.

> More studies have been made to understand morphogenetic problems of animals rather

than plants. Recent developments in plant cells, tissues and organs of higher plants in

culture, are making the science of plant morphogenesis a fruitful one. 

> Working with plants has a number of advantages.

i. In plants embryonic regions like meristem and cambium are permanently available for the study of development.

ii. The determinate type of development and abundance of organs such as leaves; flowers and fruits make the study possible under a wide range of environmental conditions.

iii. The behaviour of individual cells during development differs in plants from animals. In animals, the individual cells are free to move whereas this mobility is absent in plants and the cells are almost always attached firmly to the neighbours so that morphogenetic movements have no part in the development. This makes the study of morphogenetic problems simple in plants.

vi. The lesser plasticity of plant cells, their stationary habit, susceptibility to changes under environmental influences, ability to maintain polarity and differentiation and generation potential favour the study relatively simple one.

Morphogenesis in vitro:- Various terms are used to define the phenomenon in in vitro studies. For example differentiation, de-differentiation, re-differentiation, regeneration and morphogenesis are terms with overlapping meanings. To give a clear-cut view for the usage of terms, the sharp differences among them exposed hereunder.

i. Differentiation:- The term differentiation is used in many different senses in biology. In broad sense, it is defined as the process by which meristematic cells are converted into two or more types of cells, tissues or organs which are qualitatively different from each other.

ii. De-differentiation:- The term is used to denote the process of formation of unorganised tissues from the highly organized tissues.

iii. Re-differentiation:- The process of differentiation occurring in an undifferentiated tissue.

iv. Regeneration:- It is defined as the structuring of any part, which has been removed or physiologically isolated from the organism. In other words, genesis of an entire plant from cultured explants directly or via callus indirectly is called regeneration.

v. Morphogenesis:- Attainment of biological organization or form is termed as morphogenesis. Under in vitro conditions this can be achieved by two routes: de novo origin of organs, either shoots or roots from the cultured tissues precisely termed as organogenesis and de novo origin of embryos with distinct root and shoot poles on opposite ends from the somatic cells or cells cultured in vitro, otherwise called as somatic embryogenesis

Morphogenetic factors for in vitro regeneration:- Morphogenesis in culture proceeds along a number of pathways. Of them, two are major pathways - organogenesis and somatic embryogenesis. Organogenesis includes direct genesis of adventitious shoots or roots and indirectly via callusing. Embryogenesis also possesses two pathways where the outcome differs in the form "bipolar somatic embryos" which in later stage form individual plantlets. Several factors influence the phenomenon of morphogenesis considerably during culture. 

1. Genotype:- In the plant kingdom, certain plant groups appeared to respond more readily in culture than others. Members of carrot family (Umbelliferae) are considered to be a group that can readily form somatic embryos in culture. However, differences in response were observed among the different species of a genus and different cultivars in a species. It is now well accepted that genetic factors contribute to the response of plant tissues in culture. Though there are reports of recalcitrance among plant species to culture, this problem can be successfully overcome by manipulation of explants, culture medium or culture environment.

2. Explant:- Although all cells in a plant are considered totipotent, there are striking differences from cell to cell and from organ to organ within a plant to regenerate plants. In general, embryonic, meristematic and reproductive tissues appear to have greater potential for growth and morphogenesis in culture. For woody species, it is possible to regenerate some types of organs only when embryos or young inflorescences are cultured. The inoculum must comprise actively dividing cells or juvenile cells. It is a well known fact that physiological stage of the mother plant, its nutritional and environmental conditions would also affect the explant for morphogenesis. So the mother plant should be grown in a well controlled environment to get reproducible results even though some changes in endogenous rhythm are not avoidable.

3. Growth regulators:- It is known that the control of morphogenesis in the majority of the cultures is largely a function of the exogenous auxin/cytokinin ratio. High concentrations of kinetin cause shoot initiation, whereas high levels of auxin favour rooting. In somatic embryogenesis, auxin is required for induction of embryonic cells and maintenance of proliferative growth. Embryo formation can be induced by transferring the callus to less auxin medium or a medium lacking auxin. Plant growth regulators other than auxins and cytokinins have been shown to play an important role in the induction and control of morphogenesis. Gibberellic acid has been used most successfully to obtain rapid growth

of shoot apices and somatic embryos into plants.

4. Nutrient medium:- Components of nutrient medium play critical roles in controlling morphogenesis in culture. Effects of many inorganic and organic nutrients have been studied extensively. One of the most important components of the medium in effecting morphogenesis is the source and concentration of nitrogen. Supply of high levels of reduced nitrogen appears suitable to shoot formation and essential to somatic embryogenesis. This is supplied in the form of ammonium nitrate and sometimes substituted with amino acids such as glutamine, glycine and alanine and their amides. Presence of potassium in the medium enhances embryogenesis.

5. Other additives:- Supplementation of medium with casein hydrolysate and coconut milk also favour the morphogenesis in vitro. Coconut milk has been employed extensively as a medium component for somatic embryogenesis.

6. Culture environment:- Temperature, photoperiod, light intensity and osmotic concentration are other factors that may have determining role in organogenesis and embryogenesis. The optimum temperature for culture is 24 ± 2oC. Low temperature treatment of explants prior to culture favours their regenerative ability. Light also exerts a strong morphogenetic effect on plants in culture. Usually cultures produce shoots but the period of lighting should be maintained according to the photoperiodism of normal environment. The blue region of the spectrum promotes shoot formation and red light favours rooting. In the light, the somatic embryos of carrot formed plants; in the absence of light etiolation occurred. Overall osmotic concentration of a medium can also exert a profound effect on morphogenesis. Increased osmotic levels in medium enhance shoot and somatic embryo formation. The osmotic level can be increased by adding additional sucrose.