Plant tissue culture is the process of growing plant cells, tissues or organs in sterile conditions on a nutrient medium. It has many applications like germplasm preservation of endangered plants, genetic improvement of crops, and production of secondary metabolites. The basic steps include selection of explant, initiation of culture on growth media, multiplication through cell division, and rooting and transfer to soil. Technologies such as micropropagation, somatic hybridization, and cryopreservation have been commercialized for mass propagation of crops. However, issues remain regarding genetic stability, selection of fused products, and regeneration efficiency.
HYBRIDIZATION & HAPLOID PRODUCTION
Introduction
WIDE HYBRIDIZATION
INTER-SPECIFIC HYBRIDIZATION
Barriers to distant hybridization
Techniques to overcome barriers
Haploids and Doubled Haploids in Plant
Production of haploids and doubled haploids
a) Induction of maternal haploids
Wide hybridization
3. In vitro induction of maternal haploids – gynogenesis
Induction of paternal haploids – Androgenesis
Production of Homozygous Diploid Plants
Application of Haploids in Plant Breeding
Importance and Implications of Anther and Pollen Culture
A process where an embryo is derived from a single somatic cell or group of somatic cells. Somatic embryos (SEs) are formed from plant cells that are not normally involved in embryo formation.
Embryos formed by somatic embryogenesis are called Embryoids.
The process was discovered for the first time in Daucas carota L. (carrot) by Steward (1958), Reinert (1959).
The isolation, culture and fusion of protoplasts is a fascinating field in plant research. Protoplast isolation and their cultures provide millions of single cells (comparable to microbial cells) for a variety of studies.
WHAT IS ARTIFICIAL SEED..?
Artificial seed can be defined as artificial encapsulation of somatic embryos, shoot bud or aggregates of cell of any tissues which has the ability to form a plant in in-vitro or ex-vivo condition.
Artificial seed have also been often referred to as synthetic seed.
HISTORY
Artificial seeds were first introduced in 1970’s as a novel analogue to the plant seeds.
The production of artificial seeds is useful for plants which do not produce viable seeds. It represents a method to propagate these plants.
Artificial seeds are small sized and these provides further advantages in storage, handling and shipping.
The term, “EMBLING” is used for the plants originated from synthetic seed.
• The use of synthetic varieties for commercial cultivation was first suggested in Maize (Hays & Garber, 1919).
HYBRIDIZATION & HAPLOID PRODUCTION
Introduction
WIDE HYBRIDIZATION
INTER-SPECIFIC HYBRIDIZATION
Barriers to distant hybridization
Techniques to overcome barriers
Haploids and Doubled Haploids in Plant
Production of haploids and doubled haploids
a) Induction of maternal haploids
Wide hybridization
3. In vitro induction of maternal haploids – gynogenesis
Induction of paternal haploids – Androgenesis
Production of Homozygous Diploid Plants
Application of Haploids in Plant Breeding
Importance and Implications of Anther and Pollen Culture
A process where an embryo is derived from a single somatic cell or group of somatic cells. Somatic embryos (SEs) are formed from plant cells that are not normally involved in embryo formation.
Embryos formed by somatic embryogenesis are called Embryoids.
The process was discovered for the first time in Daucas carota L. (carrot) by Steward (1958), Reinert (1959).
The isolation, culture and fusion of protoplasts is a fascinating field in plant research. Protoplast isolation and their cultures provide millions of single cells (comparable to microbial cells) for a variety of studies.
WHAT IS ARTIFICIAL SEED..?
Artificial seed can be defined as artificial encapsulation of somatic embryos, shoot bud or aggregates of cell of any tissues which has the ability to form a plant in in-vitro or ex-vivo condition.
Artificial seed have also been often referred to as synthetic seed.
HISTORY
Artificial seeds were first introduced in 1970’s as a novel analogue to the plant seeds.
The production of artificial seeds is useful for plants which do not produce viable seeds. It represents a method to propagate these plants.
Artificial seeds are small sized and these provides further advantages in storage, handling and shipping.
The term, “EMBLING” is used for the plants originated from synthetic seed.
• The use of synthetic varieties for commercial cultivation was first suggested in Maize (Hays & Garber, 1919).
Mass multiplication procedure for tissue culture and PTC requirementDr. Deepak Sharma
This presentation include basic Micropropagation protocol: Application and advantages of mass multiplication. Beside this the requirement of tissue culture are there (Nutrient, gelling agent, energy source, vitamins and PGRs) are also included.
I have discussed Applications of Plant Tissue Culture under the following subheadings,
1. Micro Propagation
2. Clonal Propagation
3. Production of Genetically Variable Plants
4. Production of Virus Free Plants
5. Plant Breeding
6. Production of Useful Biochemicals
7. Preservation of Plant Genetic Resources
8. Importance of Tissue Culture in Biotechnology
Plant biotechnology also known as green biotechnology is the use of biotechnology in plant or crop production. There are several techniques used such as ell culturing. Organ culture, explant culture, cell suspension culture are some culture types. This is a very useful technology in which have several applications like synthetic seed production, somaclonal variation, cybridization, hybridization.
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This Gasta posits a strategic approach to integrating AI into HEIs to prepare staff, students and the curriculum for an evolving world and workplace. We will highlight the advantages of working with these technologies beyond the realm of teaching, learning and assessment by considering prompt engineering skills, industry impact, curriculum changes, and the need for staff upskilling. In contrast, not engaging strategically with Generative AI poses risks, including falling behind peers, missed opportunities and failing to ensure our graduates remain employable. The rapid evolution of AI technologies necessitates a proactive and strategic approach if we are to remain relevant.
The French Revolution, which began in 1789, was a period of radical social and political upheaval in France. It marked the decline of absolute monarchies, the rise of secular and democratic republics, and the eventual rise of Napoleon Bonaparte. This revolutionary period is crucial in understanding the transition from feudalism to modernity in Europe.
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Operation “Blue Star” is the only event in the history of Independent India where the state went into war with its own people. Even after about 40 years it is not clear if it was culmination of states anger over people of the region, a political game of power or start of dictatorial chapter in the democratic setup.
The people of Punjab felt alienated from main stream due to denial of their just demands during a long democratic struggle since independence. As it happen all over the word, it led to militant struggle with great loss of lives of military, police and civilian personnel. Killing of Indira Gandhi and massacre of innocent Sikhs in Delhi and other India cities was also associated with this movement.
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1. Plant Tissue Culture And its
Applications in Crop
Improvements
Arun patel
M.Sc- Agriculture Biotechnolgy
2. Learning Objectives
Introduction to Plant Tissue Culture
History of Tissue Culture
Media For Tissue Culture
Various Techniques for Tissue Culture
Germplasm Preservation
Applications
Limitations
3. Introduction to Tissue Culture
Tissue Culture (also known as Micropropagation or
In vitro culture) is:
The growing of plant cells, tissues, organs, seeds or
other plant parts in a sterile environment on a
nutrient medium.
4. HISTORY OF PLANT TISSUE CULTURE
1838-39 cellular theory (Cell is
autonom and totipotent)
Schleiden-
Schwann
1902 First attempt of plant tissue
culture
Harberlandt
1939 Continuously growing callus
culture
White
1946 Whole plant developed from
shoot tip
Ball
1950 Organs regenerated on callus Ball
1954 Plant from single cell Muir
1960 Protoplast isolation Cocking
5. 1962 MS media Murashige -
Skoog
1964 Clonal propagation of orchids Morel
1964 Haploids from pollen Guha
1970 Fusion of protoplasts Power
1971 Plants from protoplasts Takebe
1981 Somaclonal variation Larkin
1967 Anther Culture Maheshwari
7. Nutrient medium and the role of growth
hormones?
The nutrient medium commonly contains
Macronutrient, micronutrient, vitamins, Ferron &
carban source
The optimum culture medium may vary with the
species, the genotype within the species, and the
origin and age of the cultured tissue.
The preferred physical state of the culture medium,
whether a liquid medium or a solid agar gel, may
vary with the species and the culture environment.
pH- 5.8
8. Hormones in the agar
Two Hormones Affect Plant Differentiation:
Auxin: Stimulates Root Development
Cytokinin: Stimulates Shoot Development
Generally, the ratio of these two hormones can
determine plant development:
Auxin ↓Cytokinin = Root Development
Cytokinin ↓Auxin = Shoot Development
Auxin = Cytokinin = Callus Development
10. Types of In Vitro Culture
Culture of intact plants (seed and seedling culture)
Embryo culture (immature embryo culture)
Organ culture
Callus culture
Cell suspension culture
Protoplast culture
Somatic Embryogenesis
Micropropagation
Somaclonal variation
11. Micropropagation
Embryogenesis
Direct embryogenesis
Indirect embryogenesis
Organogenesis
Organogenesis via callus formation
Direct adventitious organ formation
Microcutting
Meristem and shoot tip culture
Bud culture
12. Steps of Micropropagation
Stage 0 – Selection & preparation of the mother plant
sterilization of the plant tissue takes place
Stage I - Initiation of culture
explant placed into growth media
Stage II - Multiplication
explant transferred to shoot media; shoots can be constantly divided
Stage III - Rooting
explant transferred to root media
Stage IV - Transfer to soil
explant returned to soil; hardened off
13. Features of Micropropagation
Clonal reproduction.
Way of maintaining heterozygosity.
Multiplication Stage can be recycled many times to
produce an unlimited number of clones.
Routinely used commercially for many ornamental species,
some vegetatively propagated crops.
Easy to manipulate production cycles
Not limited by field seasons/environmental influences.
Disease-free plants can be produced
Has been used to eliminate viruses from donor plants.
15. What is Callus development
A callus is a blob of tissue – (mostly undifferentiated
cells)
A callus is naturally developed on a plant as a result
of a wound
This callus can be left to develop or can be further
divided
16. Callus Culture
Equimolar amounts of auxin and cytokinin stimulate
cell division. Leads to a mass proliferation of an
unorganised mass of cells called a callus.
Requirement for support ensures that scale-up is
limited.
Callus Suspension Culture
When callus pieces are agitated in a liquid medium,
they tend to break up.
Suspensions are much easier to bulk up than callus
since there is no manual transfer or solid support.
17. Protoplast Isolation
Created by degrading the cell wall using enzymes.
Very fragile, can’t pipette.
The membranes are made to fuse.
osmotic shock, electrical current, virus
Regenerate the hybrid fusion product.
Contain genome from both organisms.
Very, very difficult .
18. Use of enzymes results
in a high yield of
uniform protoplasts
after removal of cellular
debris Protoplasts can
originate from different
sources: greenhouse or
field material,
micropropagated
plants, calli,
19. Protoplast Fusion Techniques
Protoplast fuse spontaneously during isolation process
mainly due to physical contact.
Induced Fusion.
Chemofusion- fusion induced by chemicals.
Types of fusogens
PEG
NaNo3
Ca 2+ ions
Polyvinyl alcohal
Mechanical Fusion- Physical fusion of protoplasts under
microscope by using micromanipulator and perfusion
micropipette.
22. Uses for Protoplast Fusion
Combine two complete genomes
Another way to create allopolyploids
Partial genome transfer
Exchange single or few traits between species
May or may not require ionizing radiation
Genetic engineering
Micro-injection, electroporation, Agrobacterium
Transfer of organelles
Unique to protoplast fusion
The transfer of mitochondria and/or chloroplasts between
species
23. Somaclonal Variation
Variation found in somatic cells dividing mitotically in culture
A general phenomenon of all plant regeneration systems that involve a
callus phase
Some mechanisms:
Karyotipic alteration
Sequence variation
Variation in DNA Methylation
Two general types of Somaclonal Variation:
Heritable, genetic changes (alter the DNA)
Stable, but non-heritable changes (alter gene expression, epigenetic)
24. Somaclonal Breeding Procedures
Use plant cultures as starting material
Idea is to target single cells in multi-cellular culture.
Usually suspension culture, but callus culture can work (want as much
contact with selective agent as possible).
Optional: apply physical or chemical mutagen.
Apply selection pressure to culture.
Target: very high kill rate, you want very few cells to survive, so long as
selection is effective.
Regenerate whole plants from surviving cells.
25. Advantages of somatic hybridization
Production of novel interspecific and intergenic hybrid
Pomato (Hybrid of potato and tomato).
Transfer gene for disease resistance, abiotic stress
resistance, herbicide resistance and many other quality
characters.
Production of heterozygous lines in the single species
which cannot be propagated by vegetative means.
Production of unique hybrids of nucleus and cytoplasm.
26. Plant germplasm preservation
In situ : Conservation in ‘normal’ habitat
rain forests, gardens, farms
Ex Situ :
Field collection, Botanical gardens
Seed collections
In vitro collection: Extension of micropropagation techniques
Normal growth (short term storage)
Slow growth (medium term storage)
Cryopreservation (long term storage
DNA Banks
27. Cryopreservation
Storage of living tissues at ultra-low temperatures (-196°C)
Conservation of plant germplasm
Vegetatively propagated species (root and tubers, ornamental,
fruit trees).
Conservation of tissue with specific characteristics
Medicinal and alcohol producing cell lines
Genetically transformed tissues.
Transformation/Mutagenesis competent tissues (ECSs).
Conservation of plant pathogens (fungi, nematodes)
28. Applications:
Study of Biochemical & Physiological activities.
The effect of various hormones.
Production of Secondary Metabolites.
To preserve the plant species which are on red-line.
Improve crop yield with regard to molecular
breeding & Genetic Engineering.
To make transgenic & cis-genic plants.
29. Commercial Applications of Clonal Propagation
Clonal propagation has the potential for propagation
of thousands of plantlets from a single genetic stock.
Examples:
orchids,
potato,
asparagus,
strawberry, and
various flowers or herbaceous ornamentals that set seed
poorly.
This may not be suitable for seeding field crops.
30. Problems in Tissue Culture
Application of protoplast technology requires efficient
plant regeneration system.
The lack of an efficient selection method for fused
product is sometimes a major problem.
The end-product after somatic hybridization is often
unbalanced.
Regeneration products after somatic hybridization are
often variable.
It is never certain that a particular characteristic will be
expressed.
Genetic stability.
Sexual reproduction of somatic hybrids.
31. 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.
PTC , G.E. and Molecular breeding these techniques
are used to transfer the gene of same species or from
different species.
32. References
Plant Tissue Culture, ELESIVISER Publishers
,Bhojwani & Rajdhan
H.S. Chawla
M. S. Shekhawat
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