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This comprehensive overview covers plant tissue culture, including its definition, advantages, history, and various techniques like callus, flower, anther, pollen, embryo, seed, protoplast, leaf, and root culture. It discusses nutritional requirements, applications in conservation, genetic modification, micropropagation, and production of haploid, disease-free plants, and secondary metabolites. The document also introduces edible vaccines, their mechanism, advantages, disadvantages, and potential applications in treating diseases.
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Plant tissue culture is defined as culturing plant seeds, organs, explants, tissues, cells or protoplasts on a chemically defined synthetic nutrient media(liquid/suspension) under sterile and controlled conditions of light, temperature and humidity. Basically plant tissue culture is a method of biological research in which fragment of tissue from an animal or plant are transferred to an artificial environment in which they can continue to survive and function. The culturetissue may consists ofa single cell or tissue a population of cells or a whole part of a plant. ADVANTAGES (i) Desired medicinal plant can be produced (ii) Production of genetically modified plant (iii) Help to produce new plants from a single or small part of parent plant such as seed, embryo, root tip, shoot tip, pollen grains etc (iv) Produce disease free plant (v) Produce exact copies of plants (vi) Quickly produce mature plants (vii) Also helpful techniques for production of primary and secondary metabolites HISTORICAL DEVELOPMENT OF PLANT TISSUE CULTURE Firstly this technique is proposed in 1902 by Gottlielo Hoberlandt , the successfully carried out tissue culture of mesophyll tissue of a tissue. TYPES OF CULTURE (i)callus culture, (ii) flower culture (complete) ,(iii) anther and pollen culture, (iv) embryo culture,(v) seed culture,(vi) protoplast culture,(vii) leaves culture,(viii) root culture
(i) Callus Culture It is undifferentiated mass which produced from an explant of plant in nutrient medium under aseptic condition is known as callus Culture. Importance – (a) It is the source of tissue for plant regeneration (b) Several biochemical assay are performed from callus culture (c) It can be used for long term maintenance of the cell lines (d) For maintain the callus culture for growth , it is necessary to subculture the callus ® Plant growth in callus culture in four phase (sigmoid cruve):- Lag phase, log phase, stationary phase, death phase (ii) Flower Culture In which new plant is generate from a flower. Procedure— Flower bud or matures flowers are collected washed and dipped in 5% tcepol solution for 10min (sterilization) then rewashed in laminar air flow cabinet using sterilized forceps bud or mature flower to nutrient mediaincubate the culture for 10 hrs at 25°c (iii) Anther and pollen Culture Anther excised from flower sterilized and washed with double distilled water directly transferred in nutrient agar or liquid media the plantlet are formed after 4-5 weeks of inoculation(embryogenesis occurs)pollen grains are aseptically removed from the anther and cultured on liquid medium then incubated at 28°C in dark for 2-3 weeksobtain haploid embryo or callus (iv) Embryo Culture Embryo is obtained or excited /isolated from ovule , seed or fruittransfer into nutrient medium under aseptically (salt, sugar, agar etc)incubate at 25°C for 16 hrs sub-culturing is done in fresh medium in the interval of 3-4 weeks (v)Seed Culture Seeds are cultured in vitro to generate seedling or plants in aseptic conditions for raising the sterile seedlings explants plants. It increase the efficiency of germination of seed
It present the growth of microbes in plant tissue culture Ex Streptomycine, Kanomycine etc.. (E) pH:- The pH of nutritional medium is generally adjusted between 5-6.
Application of plant tissue culture (I) to conserve rare species or endangered plant species (II) production of genetically modified plants (III) micropropagation (regeneration of whole plant through tissue culture) (IV) production of haploid plants (contain single set of chromosomes ) for improving crops (V) production of microbes free plants and also disease free plants (VI) to study the respiration and metabolism of plants (VII) production of sensory metabolites (responsible for protection of plants) (VIII) production of phytoconstituents in increased amount (IX) production of plant clones with new characteristics (X) for the evolution of organ functions in plants for the improvement of crops VACCINES Vaccines typically contains an agent that is made of weakened, killed form of microbes. It’s toxins or surface proteins stimulates the immune response.. IDEAL VACCINE (1) it should not be toxic or pathogenic (2) it should have low level of side effect (3) should not contaminate the environment (4) should not cause problem in individual (5) technique of vaccination should be simple (6) it should be cheap EDIBLE VACCINE Vaccine that one can eat are called edible vaccine. This are transgenic plant and animal based preparation that content some agent which trigger or stimulate an immune response. Concept of Edible Vaccine Developed by ARNDZEN in 1990 Introduction of gene of interest into the plant (Transformation) gene expressed in the plant tissue edible part (Transgenic plant ) genes encode_____ protective vaccine antigens for viral, bacteria and parasitic pathogen that cause disease in human and animals ingestion of the edible part of the transgenic plant Mechanism of Action After the oral administration of edible vaccine the hard outer layer of plant cell which protect them from antigen released the antigens in transgenic plants are delivered to these process M cells intestinal living take up these antigens they pass onto macrophages onto other antigens M cells located in payres patch are source of immunoglobulin antigens activate the beta cells within the lymphoid follicles after the local lymphosyte activation serum, IGG, IGE response. IGA responses
on memory cells neutralize the effect of the infection immune response generated which cause active immune response against disease. Preparation of Edible Vaccine Prepared by three methods (1) Bombardment method (a) DNA sequence selected (b) Precipitate the DNA sequence on metal micro particles (c) Bombardment is done on plant cell with a gene gun at high speed (d) The microparticles precipitate the cell wall and release exogenous DNA inside cell where it integrates into plant genome (e) Gene encoding antigen from pathogen is inserted into bacterial vector (2) Plasmid Vector Carrier System Agrobacterium tumofacience is a naturally occurring soil bacterium which has the ebelity to enter the plants through wounds and scratches. Process— (a) Cut leaves and expose the leave to bacterial carrying an antigen gene and antibiotic resistant gene. Allow bacteria to deliver the gene into the leaf cell. (b) Expose leaf to an antibiotic to kill cells that lack the new cells/genes. wait for surviving (gene altered) cells to multiply and form a callus (c) Allow to generate roots and shoots (3) Electroporation (a) It is the method in which the DNA is inserted into the cell (b) They are exposed to high voltage electric pulse which produce the pores in the plasma (c) Electroporation causes weakening of cell wall by which entry of DNA into the cytoplasm is possible ADVANTAGE OF EDIBLE VACCINE ---- (1) Enhance patient compliance specially in children because of oral administration (2) Eliminate the need of trained medical practitioners (3) Production is highly efficient and be easily scaled up (4) Activate mucosal and systemic activity (5) Easy to store because refrigeration is not required (6) Cheap and deliver multiple antigen DISADANTAGES ---- (1) Develop of immune tolerance to vaccine peptide or proteins (2) Stability of vaccines in fruits is known known (3) Dosage form of vaccine may be variable (4) Consistency of dosage form fruit to fruit , generation to generation, plant to plant is not similar APPLICATION---- (a) Malaria , (b) Measles , (c) Diabetes