MORPHOGENETIC POTENTIAL OF EXPLANT

 

MORPHOGENETIC POTENTIAL OF EXPLANT

 

Ø  Plant tissue culture is the in vitro aseptic culture of cells, tissues, organs, or whole plant under controlled nutritional and environmental conditions to produce the clones of plants.

Ø  The resultant clones are true to type of the selected genotype. The controlled condition provide the culture an environment conducive for their growth and multiplication.

Ø  These conditions include proper  supply of nutrients, pH medium , adequate temperature and proper gaseous and liquid environment.

Ø  Plant tissue culture technology is being widely used for large scale plant multiplication. Apart from their use as a tool of research, plant tissue culture techniques have in recent years, become of major industrial importance in the area of plant propagation , disease elimination, plant improvement by the production of secondary metabolites.

Ø  Small pieces of tissue can be used to produce hundreds and thousands of plants in a continuous process.

Ø  A single  explant can be multiplied into several thousand plants in relatively short time period and space under controlled condition, irrespective of the season and weather on a year round basis. Endangered , threatened and rare species have successfully been grown and conserved by micropropagation  because of high coefficient of multiplication and small demands on number of initial plants and space.

Ø  Certain type of callus culture give rise to clones that have inheritable characteristics different from those of parent plants due to development of commercially important improved varieties .

Ø  Higher yields have been obtained by culturing pathogen free germplasm in vitro.

     SELECTION OF EXPLANT

 

·         The explants is selected it is either haploid or diploid explants

·         The plant growth can be achieved in two ways:

1.      Shoots directly by appropriate media

2.      By somatic embryogenesis

      FACTORS AFFECTING EXPLANT’S TISSUE CULTURE

             plant material is extremely important for the success of tissue culture. Factors affecting explant’s tissue culture response are

·         Genotype

·         Phusiological stage of donor plant

·         Explant source

·         Explant age

·         Explant size

·         Explant position in donor plant

·         Explant density

 

 

1.EXPLANTS – VEGETATIVE PLANT PARTS

  

1.SHOOT PRIMORDIA (Apical meristem)

·         The apical meristem is the growth region in plant found within the root tips and the tips of the new shoots and leaves.

        ·         Meristem culture can difine as the tissue culture technique which makes the use of apical            meristem with 1-3 leaf primordial by which clones of a plant can develop by vegetative propagation. The  meristem can culture by isolating from the stem by applying V shaped cut. In this technique , by the culturing of shoot meristem , adventitious roots can regenerate.

·         By the meristem culture , virus free plants can grow. The meristem from a plant which produces heterozygous seeds can store in the vitro conditions.

·         Meristem culture also helps in the plant breeding technique where hybrid breeds of the plant can grow. The quarantine authority for the international exchange easily accepts the plants obtained by the meristem culture

·         It is also useful in the micropropagation technique which involves the vegetative or asexual propagation of the whole plant.

·         The germplansm or the seed can conserve in vitro or by cryopreservation method.

·         Meristem contains high auxin concentration  which promotes plant growth.

·         It is a very effective method in the field of agriculture and industrial science, by which clones of a plant can prepare and disease resistant plants can also develop e.g., Tobacco Mosaic virus has been eliminated from Petunia species of the plant , Cauliflower Mosaic virus has been eliminated from the Brassica oleracea plants.

 

2.SHOOT TIP

  The terminal bud of a plant , which consist of the apical meristem and the immediately surrounding leaf primordial and developing leaves, and adjacent stem.

·         Actively growing shoot tip is surface sterilized and is placed on a defined culture medium under sterile condition.

·         This media must contains growth hormones and will often form roots and develop into whole plants.

·         Some of the crop species that have been freed of viruses by this technique, they include soyabean , sweet potato , sugar cane and rhubarb.

·         This method is used with both monocot and dicot species.

 

 

3.LEAVES OR LEAVES PROMORDIA

·         Leaves may be detached from shoots, surface sterilized and place in healthy condition for long period.

·         Growth rate in the culture depends on their stages of maturity at excision.

·         Young leaves have more growth potential then the nearly mature ones.

·         It is observed that explants from immature young leaves grow better than explants from older leaves.

·         It is believed that leaf culture depends upon the physiological state and the age of the leaf.

·         The shoot forming potentials differ in the leaf cultures as per the derivation of the explants and the hormonal factors involved.

4.ROOT TIP

  The terminal portion of a root or root branch usually including the root cap and the meristematic region behind it and often the regions of differentiation , elongation and root hair formation.

·         Root cultureis the culture of apical or lateral root tips to produce in vitro root system.

·         Root tip culture are maintained in an agitated liquid medium with appropriate auxin.

·         These are mainly based with the inoculation of agrobacterium rhizogenes to produce secondary metabolites.

·         The lateral roots continue to grow and provide several roots, which after seven days are used to experimental cultures.

·         By this culture method it is possible to study the nutritional requirement of roots, shoot and root growth, conditions required for the  development of secondary vascular tissues , lateral root and bud formation, nodulation etc.

 

EXPLANTS – REPRODUCTIVE PLANT PARTS

 

1.FLOWER OR FLORAL BUD

 

·         Bud , small lateral or terminal protuberance on the stem of a vascular plant that may develop into flower, leaf  or shoot. Buds arise form from meristem tissue .

·         In floral bud culture Flower two days after pollination are excised, sterilized by immersion in 5% calcium hypochlorite, repeatedly washed with sterilized water  and transferred to culture tubes containing an agar mediu

·         When cultured , such flowers produce fruits. Larger fruits are obtained on medium supplemented with growth hormones. Flowers excised before pollination do not produce fruits.

·         This culture system is useful in studying microclimates or nutritional effects on the vegetative and reproductive processes of the plant.

 

 

 

2.ANTHER

 

 

·         Androgenesis is the in vitro development of haploid plants originating from potent pollen grains through a series of cell division and differentiation. It is a diploid structure(2n).

·         Two techniques are used to produce androgenic haploids, viz. anther culture and isolated pollen culture.

·         Anther is the part of  a stamen that produces and contains pollen and is usually borne on a stalk.

·         Anther culture is a technically simple and efficient method, requiring minimum facilities.

·         Flower buds with pollen grains at the most responsive stage are surface sterilized and the anthers, excised under aseptic conditions, cultured on semi solid or in liquid medium. in some cases, where the flower buds are small, whole buds or inflorescence enclosing the anthers at the appropriate stage of pollen development are cultured.

·         The cultures are exposed to a suitable stress treatment before incubation under normal culture conditions in dark.

·         The anthers are generally cultured on a solid agar medium where they develop into embryoids for the anther culture under alternate light and dark period.

 

 

3.POLLEN

 

·         Pollen is a fine powdery substance, typically yellow, consisting of microscopic grains discharged from the male part of a flower or from male cone. Each grain contains a male gamete. It is a haploid (n) gamete .

·         Pollen culture is now possible to achieve androgenesis in the cultures of mechanically isolated pollen of several plants.eg,. tobacco ,Brassica species.

·         Besides the culture medium and pretreatment , the planting density is a critical factor for the induction of androgenesis in pollen culture.

·         Treatment of pollen derived embryos and pollen derived calli  to regenerate complete plants  is the same as in anther culture.

·         The nutritional requirements of the isolated pollen in culture are more complex than those of cultured anthers.

·         A homogenous preparation of pollen at the developmental stage most suitable for androgenesis can be obtained by gradient centrifugation

·         Isolated pollen can be genetically modified by mutagenesis or genetic engineering before culture, and a new genotype can be selected at an early stage of development.

·         Pollen culture considerably improves the efficiency of androgenesis.

 

 

 

4.OVULE 

·         The part of the ovary of seed plants that contains the female germ cell and after fertilization becomes the seed.

·         In vitro culture of unfertilized ovules , ovaries is another approach to trigger apogamey. This method of haploid production called In vitro gynogenesis provides an attractive alternative to produce haploid of plants where androgenesis is either not applicable or unsuccessful or is fraught with problems such as formation of non haploid and albinos with high frequencies.

·         To- date gynogenesis has been achieved in about 25 species , including some important crop plants.

·         In ovule culture mature embryos are excised from ripened ovule and cultured mainly to avoid inhibition in the seed for germination. very small globular embryos require a delicate balance of the hormones.

·         Embryo is dissected from the ovule and put into culture media.

·         This type of culture is relatively easy as the embryos require a simple nutrient medium containing salts, sugar and agar for growth and development.

·         Multicellular immature embryos are dissected out and cultured aseptically to obtain viable hybrids. Once the embryo is rescued , two genomes are needed to be combined together to produce a fertile plant.

·         By this method dormancy period of seeds can be shortened , as well as haploids can be produced.

·         By ovule culture , it is possible to grow , study various nutritional requirements and stages young embryos or zygote.

 

 

 

5.OVARY

·         It’s the female part of the plant which has ovule . it is a diploid(2n) structure.

·         Ovaries excised after pollination can produce fruits on a simple medium containing mineral salts, sugar and vitamins.

·         Ovary taken from un pollinated flowers fail to produce fruits on such a simples medium but can develop into seedless fruits on a medium supplemented with hormones.

·         Physiology of fruit development can be studied.

·         Haploid can be produced.

·         Rare hybrids can also be produced by ovary culture.

6.COTYLEDON

  Cotyledon is an embryonic leaf in seed bearing plants, one or more of which are the first leaves to appear from a germinating seed.

·         In cotyledon culture Immature cotyledon develops into somatic embryos , shoot buds and complete plants if cultured on a suitable nutrient medium.

 

7.EMBRYO

 An embryo refers to the early developmental stage of eukaryotic organisms following the fertilization of an egg by sperm as a method of sexual reproduction. In plants the process of embryogenesis extends from the time of fertilization until dormancy.

·         In vitro zygotic embryo culture plants require a source of exogenous energy for carrying out the photosynthesis. These results confirm the hypothesis that depending on the species and stage of development of embryos. Embryos younger typically require higher concentration of carbohydrates in the culture medium to sustain germination.

·         Somatic embryogenesis is an in vitro method of plant regeneration widely used as an important biotechnological tool for sustained clonal propagation.

·         It is a process by which somatic cells or tissues develop  into differentiated embryos.

·         These somatic embryos can develop into  whole plant without undergoing the process of sexual fertilization as done by zygotic embryo.

·         Somatic embryogenesis has been reported in many plants including trees and ornamental plants of different families.

·         Somatic embryogenesis differs from organogenesis in the embryo , being a bipolar structure rather than monopolar.

·         For some species any part of the plant body serves as an explants for embryogenesis eg,. Carrot. Whereas in some species only certain regions of the plant body may respond in culture eg,.cereals.

 

·         Direct embryogenesis: the embryo initiates directly from the explants tissue through pre embryonic determined cells.

·         Indirect embryogenesis: cell proliferation, i.e. callus from explant ,takes place from which embryo arises from induced embryogenic determined cells.

·         Embryo culture is a type of plant tissue culture that is used to grow the embryos from seeds and ovules in a nutrient medium

·         In embryo culture , the plant develops directly from the embryo or indirectly through the formation of callus and then subsequent formation of shoots and roots.

 

 

 

CONCLUSION:

·         PTC is the technique by which plant cells can be grown in vitro sexually & asexually. By the help of this we can study biochemical, physiological and hormones activity.

·         High yield , good quality of crops can be obtained. Plant tissue culture provides great hope not only to create new combinations of genes but also to ensure the quantity of various food commodities.