2. An Introduction
Plant tissue culture is:
A tool in biotechnology, used in
micropropagation, secondary metabolic
production(taxol), hybridisation etc.
It refers to the growth of plant
parts(organs, tissues etc.) in a sterile
environment in a nutrient medium.
3. Why Plant T.C.??
Saves time & labour in comparison to
conventional hybridization techniques.
Useful when other techniques are not
useful.
Eg.: Apical meristems which are free of
viruses(present in shoot tips) can be
harvested & nurtured to grow into virus-free
plants in times of viral attack.
4. Uses
• To save species from extinction
• To isolate disease from plants
• To produce plants with enhanced
stress or pest resistance
• To create new plant varieties
• To make money
6. Totipotency
Plant tissue culture is based on
totipotency
Plant cells have the ability to produce the
whole plant from single cells. This is called
totipotency.
Plants have the ability to reproduce
asexually through a type of cell division-
mitosis.
8. Beginning of T.C.
In 1902, a German physiologist, Gottlieb
Haberlandt proposed that single plant
cells could be cultured.
He isolated single fully differentiated
individual plant cells from different plant
species and was first to culture them in
Knop’s salt solution enriched with glucose.
9. History-1930s
In the 1930s, importance of vitamins was
determined for shoot and root culturing
Indole-Acetic Acid(IAA), a natural auxin,
and the most important, was discovered
during the 1930s.
Many more synthetic auxins were
discovered. Eg.: 2,4d &
NAA(Napthaleneacetic acid)
10. History-1957-58
Miller and Skoog discovered kinetin, a
synthetic cytokinin, in the University of
Wisconsin – Madison.
Kinetin plays active role in organogenesis.
Steward developed somatic embryo from
carrot cells
11. History-1958-60
Morel cultured orchids and dahlias & freed them
from a viral disease
History-1962
• Murashige and Skoog published recipe for MS
Medium for T.C.
12. T.C. Medium
Functions: provide H2O, provide mineral &
vitamin nutritional needs.
H2O is usually distilled
minerals must provide 17 essential
elements
energy source - sucrose is preferred
provide access to atmosphere for gas
exchange
serve as a dumping ground for plant
metabolites
13. T.C. medium
Macronutrients-eg: K, Ca, Mg,
N, P, O etc.
Micronutrients-eg: Fe, Mn, Mo,
etc.
Vitamins & Amino
acids:thiamine, pyridoxin,
nicotinic acid, biotin, citric
acid, ascorbic acid, Inositol,
Glycine etc.
Growth Regulators: auxins and
cytokinins
Carbon Source:Sucrose
pH usually 5.0-5.7
14. T.C. Steps
1. Explant collection
2. Callus generation
3. Organogenesis (shoot and root
induction)
4. Hardening
Micropropagation does not involve the
second step. There is direct
organogenesis.
15.
16. Explant collection
portion of plant removed
and used for T.C.
Important features
size
source - some tissues are
better than others
species dependent
physiological age -
young portions of plant
are most successful
17. Explant collection
degree of contamination
external infestation - soak plant
in sodium hypochlorite solution
internal infection - isolate cell
that is not infected
roots - especially difficult
because of soil contact
herbaceous plants
soft stem
easier to culture than woody
plants
18. Callus Generation
• The dedifferentiation of a plant
cell(explant) produces callus.
• Callus is then expanded into a larger
mass of undifferentiated cells.
• Callus is then activated, by selective
use of plant hormones to
redifferentiate to produce, shoots,
roots and ultimately, plantlets
19. Organogenesis
Usually induced by changes in hormonal
environment
Shooting: Higher cytokinin concentration
& lower auxin concentration(cytokinins
promote shoot growth).
Rooting: lower cytokinin concentration
and increase auxin(auxins promote shoot
growth)
20. Hardening
Hardening involves the formation of the
waxy cuticle on the leaves of the plant.
Plants in T.C. do not have cuticle
usually done in greenhouse with high
relative humidity(RH).
gradually increase light intensity and
lower RH after rooting occurs
allows plants to harden and helps plants
form cuticle
21. CREDITS TO
The inventors of tissue culture
Google images
The internet’s information
Microsoft Office 2010
