🎋 Plant Biotechnology

• Plant biotechnology exploits plant cell or its constituents for generating useful products or services for the benefit to mankind.
• It includes two broad disciplines viz., Tissue culture and Genetic engineering.

Plant tissue culture or In-vitro culture

• It is multiplication of cell of large number of plants placed in appropriate environment conditions with required nutrients.
• The ability of the plant cells/tissue to develop into a complete plant is known as totipotency.
• It is the cultivation of plant organs, tissue, embryo, seeds, protoplasts or cells in the test tubes on artificial media.
• It reduces more than half of the time taken in conventional breeding programmes.
• Explant is an excised fragment of a tissue or an organ used to start tissue culture.

Techniques of Tissue Culture

• Micropropagation is in vitro multiplication of plants from a small tissue explant.
• Surface sterilization: The explant (the plant or plant part excised for the in vitro cultivation) is surface sterilized to eliminate contaminants using Sodium Hypochlorite [NaOCl]@ 1-2 per cent or Mercuric Chloride @ 0.1 per cent solution.
• Nutrient Medium: It is a medium containing salts, trace elements, vitamins, carbon sources, growth regulators 2,4-D @ 0.5-0.2 mg/ L is commonly used. Organic supplements like Coconut agent such as agar-agar is used. Generally B-5 mediuin (Gamborg et al.) or MS medium (Murashige and Skoog) are used.
• The medium commonly used carbon source is sucrose.
• Sub-culturing: Transferring of tissue to fresh/new media after a stipulated time. Usually sub-culturing is done after every 4-6 weeks. However, the suspension cultures are sub-cultured after every 3-14 days.
• Re-culture: Process by which a cell monolayer or a plant explant is transferred without subdivision into a fresh medium.
• Plant regeneration and transfer to soil: First the transfer of cultured plant is done in small pots and then to green house and finally in soil. Transfer is done when roots and shoots appears.

Classification of Tissue Culture Techniques

• Embryo culture: Young embryo is removed from developing seeds and planted on a suitable nutrient medium in vitro, with the goal of obtaining a viable seedlings/plants. It is applied in differentiation studies and soma clonal variations.
• Meristem culture: it is a culture of isolated mature or immature embryos. It is applied in micropropagation and production of virus free plants.
• Seed culture: Culture of seeds in vitro to generate seedlings/plants.
• Anther or Pollen culture: In this technique, haploid plants are obtained from pollen grains by replacing another or pollens into a suitable medium. This technique is used to obtained haploid plants.
• Callus culture: Callus is basically a more or less non-organized tumor tissue, which usually arises on wounds of differentiated tissue and organs. This culture is used in Cryopreservation of germplasm, in vitro cell selections for resistance to biotic and abiotic stresses.
• Nucellus culture: Nucellus culture has been utilized to study factors responsible for formation of adventive embryos.
• Cell culture: The growing of individual cells that have been obtained from an explant tissue or callus. It is applied in synthesis of new chemical substances production of useful metabolites, synthesis of new chemical substances.
• Protoplast culture: Protoplasts are the cells without cell wall. Culture of protoplasts and fusion of protoplasts from different strains.
• Organ culture: Culture of an organ in vitro in a way that allows development and/or preservation of the originally isolated organ.

Totipotency

• Totipotency is capability of an isolated single cell to multiply and differentiate into multicellular organism.
• Tissue culture is purely based on the totipotency of cells.
• Haberlandt first demonstrated the totipotency of cells in 1902.

Genetic Engineering

• Change in the genetic constitution of cells by introduction or elimination of specific genes using molecular biology techniques is known as Genetic engineering. This is a non-sexual method of gene transfer.
• Genetic engineering is the artificial manipulation, modification and recombination of DNA or other nucleic acid molecules in order to modify an organism or population of organisms. Genetic engineering works through recombinant DNA technology.
• Genetic engineering allows the use of several desirable genes in a single event and reduces the time to introgress novel genes into elite background.
• Biotechnology has provided several unique opportunities and develop transgenic plants with novel genes that include:
  • access to novel molecules
  • ability to change the level of gene expression
  • capability to change the expression pattern of genes
  • develop transgenic plants with novel genes
• Paul Bergh (Father of genetic engineering).
  • He transferred gene of SV-40 virus (Simian Virus) in to E. coli with the help of γ- phage (Nobel prize - 1980).
• The concept of genetic engineering was the outcome of two very significant discoveries made in bacterial research. These were:
  • Presence of extra-chromosomal DNA fragments called plasmids in the bacterial cell which replicate along with chromosomal DNA of the bacterium.
  • Presence of enzymes restriction endonucleases which cut DNA at specific sites. These enzymes are, therefore, called ‘molecular scissors’.
• It involves the following steps:
  1. Identification and isolation of desired genes
  2. Insertion of desired gene into a DNA of suitable vector
  3. Introduction of the recombinant DNA into suitable organism for multiplication
  4. Selection of host cells carrying the desired recombinant vector
  5. Multiplication/Expression/Integration of desired gene in the host
  6. Insertion of recombinant gene in to the organism concerned

Applications of Genetic Engineering

• Development of insect resistance in trees.
• Development of disease resistance in trees.
• Development of herbicide resistance in plants.
• Improvement of seed storage proteins in tree.
• Production of novel chemicals.
• Production of edible vaccine (antithrombin production — Hirudin by B. napus)