micropropagation- a very useful technology in plant tissue culture.

2.  The culture of plant seeds, organs, tissues,
cells, or protoplasts on nutrient media under
sterile conditions.

3.  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

4.  Micropropagation
 Germplasm Preservation
 Somaclonal variation
 Embryo Culture
 Haploid Production
 In vitro Hybridization- Protoplast fusion

5.  In vitro Clonal Propagation.
 Micropropagation is the practice of rapidly
multiplying stock plant material to produce a
large number of progeny plants, using modern
plant tissue culture methods.

6.  Clone is a plant population derived from a
single individual by asexual reproduction.
 Clonal Propagation is the multiplication of
genetically identical individuals by asexual
reproduction.

7.  Clonal reproduction
 Multiplication stage can be recycled many
times to produce an unlimited number of
clones
 Easy to manipulate production cycles
 Disease-free plants can be produced

8. Rapid clonal in vitro propagation of plants:
• From cells, tissues or organs
• Cultured aseptically on defined media
• Contained in culture vessels
• Maintained under controlled conditions of
light and temperature

9. Commercialization of Micropropagation 1970s & 1980s
Murashige (1974)
Broad commercial application

10. Tip
bud
Leaf
Axillary
bud
Internode
Root
Starting material for
micropropagation

11.  Part of plant
 Genotype
 Physiological condition
 Season
 Position on plant
 Size of explant

12.  Minerals
 Sugar
 Organic ‘growth factors’
 Growth regulators
 Gelling agent
 Other additives

13.  Temperature
 Moisture
 Light

14. 1. Selection of plant material
2. Establish aseptic culture
3. Multiplication
4. Shoot elongation
5. Root induction / formation
6. Acclimatization

15.  Stage I –Establishment
◦ Selection of the explant plant
◦ Sterilization of the plant tissue takes place
◦ Establishment to growth medium
 Stage II - Proliferation
◦ Transfer to proliferation media
◦ Shoots can be constantly divided
 Stage III – Rooting & Hardening
◦ explant transferred to root media
◦ explant returned to soil

16.  Organogenesis
◦ Organogenesis via callus formation
◦ Direct adventitious organ formation
 Embryogenesis
◦ Direct embryogenesis
◦ Indirect embryogenesis
 Microcutting
◦ Meristem culture (Mericloning)
◦ Bud culture

17.  PGRs are prob. the most important factor affecting
organogenesis
◦ cytokinins tend to stimulate formation of shoots
◦ auxins tend to stimulate formation of roots
 The central dogma of organogenesis:
◦ a high cytokinin:auxin ratio promotes shoots and
inhibits roots
◦ a high auxin:cytokinin ratio promotes roots and/or
callus formation while inhibiting shoot formation

18. The process of initiation and development of a
structure that shows natural organ form and
function.
The ability of non-meristematic plant tissues to
form various organs de novo.
The production of roots, shoots or leaves.
These organs may arise out of pre-existing
meristems or out of differentiated cells.
This, like embryogenesis, may involve a callus
intermediate but often occurs without callus.

19. Tissue culture maintains the genetic of the cell
or tissue used as an explant.
Tissue culture conditions can be modified to
cause to somatic cells to reprogram into a
bipolar structure.
These bipolar structures behave like a true
embryo - called somatic embryos.

20.  An Embryo is made up of actively growing
cells and the term is normally used to describe
the early formation of tissue in the first stages
of growth.

21.  The process of initiation and development of
embryos or embryo-like structures from
somatic cells
 The production of embryos from somatic or
“non-germ” cells.
 Usually involves a callus intermediate stage
which can result in variation among seedlings

22.  Somatic embryogenesis is a useful
regeneration pathway for many monocots and
dicots, but is especially useful for the grasses
 Types of embryogenesis
◦ zygotic embryogenesis – the result of normal
pollination and fertilization
◦ somatic embryogenesis – embryos from
(cultured) sporophytic cells , that is embryos
arise indirectly

23.  The composition of the culture medium
controls the process-
◦ auxin (usually 2,4-D) added causes
induction, the formation of embrygogenic
clumps or proembryogenic masses (PEMs)
(induction medium)
◦ auxin is deleted and the clumps become
mature embryos (maturation medium)

24.  Stages of development
◦ early cell division doesn't follow a fixed
pattern, unlike with zygotic embryogenesis
◦ later stages are very similar to zygotic
embryos (dicot pattern)
 globular stage (multicellular)
 heart-shaped stage (bilateral symmetry) –
bipolarity
 torpedo-shaped stage – consists of initial
cells for the shoot/root meristem

26. Somatic Embryogenesis
Stimulation of callus or suspension cells to undergo a
developmental pathway that mimics the development of the
zygotic embryo.

27. • From one to many propagules rapidly.
• Multiplication in controlled lab conditions.
• Continuous propagation year round.
• Potential for disease-free propagules.
• Inexpensive per plant once established.

28. • Specialized equipment/facilities required.
• More technical expertise required.
• Protocols not optimized for all species.
• Plants produced may not fit industry
standards.
• Relatively expensive to set up.

29. • Equipment/facility intensive operation
• Technical expertise in management positions
• Protocols not optimized for all species
• Liners may not fit industry standard
• Propagules may be too expensive

30. • Rapid increase of stock of new varieties.
• Elimination of diseases.
• Cloning of plant types not easily propagated
by conventional methods.
• Propagules have enhanced growth features
(multibranched character;Ficus, Syngonium)