Tissue culture of plants
- 1. TISSUE CULTURE OF PLANTS BY CH.BHARGAVI DEPARTMENT OF FORESTRY
- 2. CONTENTS: • Definition • Principle • Stages • Steps • Factors effecting tissue culture • Importance
- 3. Technique of growing plant cells, tissues and organs in an artificial medium, under aseptic conditions
- 4. PRINCIPLE Plant tissue culture mainly depends on totipotency and plasticity. • Totipotency: ability of plant cells to regenerate into whole plant. • plasticity: ability of plants to alter their metabolism, growth and development to suit their environment. Genetic alterations- Ploidy level, chromosome rearrangements, point mutations, activation of transposable elements, or changes in chloroplast or mitochondrial genomes. Epigenetic alterations- alteration that persists throughout the lifetime of the regenerated plant, but is not passed on to the next generation. Eg: DNA methylation
- 5. STAGES Initiation stage Multiplication stage Rooting stage Acclimatization
- 6. STEPS • Selection of plant • Isolation of explant: Plant tissue cultures are generally initiated from multicellular tissue fragments called as explants. Explants can be originated from a wide range of plant tissues such as leaf, stem, root, cotyledon etc.
- 7. STERILIZATION: It is a process for removing or deactivating all forms of life like fungus, bacteria, spores etc. • All the materials used in culture work must be free from microbes. • It can be achieved through autoclaving, flame sterilization, dry heat treatment etc.
- 8. INOCULATION OF EXPLANT: • Explants are usually placed on culture medium which is generally composed of inorganic salts, nutrients, vitamins and harmones and gelling agents. • Types of medium: • CHU (N6 ) Medium • Gamborg B5 Medium • Murashige & Skoog Medium (MS) • Nitsch Medium • Orchid Medium • Schenk & Hidebrandt Medium (SH)
- 9. Incubation: Cultures are incubated in culture room where light, temperatire and humidity are controlled. Sub culturing: It includes transfer of cell or tissue from old culture medium to fresh cukture medium for providing sufficient space and nutrients to growing plantltt
- 10. Hardening: • Plantlets are exposed to natural conditions in hardening room . • Plants are maintained under high humidity for 10-20 days and then transferred to field so as to grow under natural conditions.
- 11. FACTORS EFFECTING TISSUE CULTURE: Plant species Genotype within species Source of cultured tissue Age and health of donor Nutrient medium
- 12. PLANT TISSUE CULTURE ENABLES THE FOLLOWING STUDIES: Totipotency, nutrition, metabolism, division, differentiation and preservation of plant cells. Morphogenesis and plant regeneration from individual cells or tissues through the process namely organogenesis or somatic embryogenesis. Variations generated through in vitro culture. Evolution of haploids through anther and pollen culture including ovule culture. Plant tissue culture enables the following studies Wide hybridization programmes through ovule, ovary and embryo cultures to overcome both pre zygotic and post zygotic sterility mechanisms
- 13. Micropropagation of plant materials In vitro selection of mutants tolerant to biotic and abiotic stresses. In vitro culture and secondary metabolite biosynthesis. Plant genetic engineering through in vitro culture methods and DNA transfer technique The technique is useful for making clones of one particular plant. Propagation of rare plant species. Certain special plant varieties that are hard to maintain by producing seed can be maintained for the long term in culture.
- 14. REFERENCE: Barcelo, P., Rasco- gaunt, s., Thorpe, c. and lazzeri,2001. Transformation and gene expression. P.R shewry, P.A. lazzeri and k j Edwards,pp Dodds , J. H and Roberts, L.W. 1995. Experiments in plant tissue culture.