Plant tissue culture

1. Micropropagation
Dr. Renu Jangid

2. Micropropagation
• Micropropagation is the process of in vitro clonal
propagation through tissue culture techniques.
• Clonal propagation refers to the process of asexual
reproduction by multiplication of genetically
identical copies of individual plants.
• Micropropagation is used to multiply plants such as
genetically modified and also used to provide a
sufficient number of plantlets for planting from a
stock plant which does not produce seeds, and does
not respond well to vegetative.

3. Stages of Micropropagation
Micropropagation is complicated process and mainly involved five stages:-
 Stage 0: Selection of source or elite plants
• Selection of explant
 Stage I: Establishment of cultures
• Sterilization of the plant tissue take place
• Establishment to grow medium
 Stage II: Proliferation
• Transfer to proliferation media
• Multiplication of the regenerated shoots
 Stage III: Rooting and Hardening
 In vitro germination of somatic embryos and rooting of shoots
 Stage IV: Transfer to soil and acclimatization
 Explant returned to soil and hardening under green house environment

5. Methods
 Explant culture
 Callus culture
 Organogenesis
 Root culture
 Shoot culture/ Meristem culture
 Cell culture
 Somatic embryogenesis
 Protoplast culture
 Anther culture
 Embryo culture

6.  Meristem culture
 Actively dividing meristems are present at the axillary buds and
shoot tips
 In Meristem culture, the meristem and a few subtending leaf
primordia are placed into a suitable growing media.
 where they are induced to form new meristem. These meristems
are then divided and further grown and multiplied.
 To produce plantlets the meristems are taken of from their
proliferation medium and put on a regeneration medium.
 When an elongated rooted plantlet is produced after some weeks,
it can be transferred to the soil .
 A disease free plant can be produced by this method.

8. Somatic embryogenesis
 The process of regeneration of embryos from somatic cells, tissues and organs
is regarded as somatic embryogenesis.
 Somatic embryo called embryoids are similar to zygotic embryos except they
originate from somatic cells and are larger in size.
 It can be initiated in two ways:- Indirect and direct
 1. Indirect:- By inducing embryogenic cells within the performed callus. In
this case, embryoids are initiated in callus from superficial cell aggregates.
 2. Direct:- From pre embryonic determined cell, ( without callus) which are
ready to differentiate into embryoids.
 This embryoids can be separated and isolated mechanically by using
glassbeads.

9. Events of somatic embryogenesis

10. Application of Micropropagation
1. Micropropagation is the best technique for the production of millions of clones
in one year.
2. Micropropagation facilitates the growth, storage, and maintenance of a large
number of plants in small spaces which makes it a cost-effective process.
3. Micropropagation is used for germplasm storage and the protection of
endangered species. Some plants are difficult or resistant to be grown on a
large scale by conventional techniques. Micropropagation is a good alternative
to cultivate these plants.
4. Micropropagation can be used to obtain disease or pathogen-free plants.
Meristem tip culture is used to achieve this goal.
5. Micropropagation is used to increase the vigor and yield of floriculture species
by multifold.
6. Micropropagation is popular for the production of synthetic seeds that analog
to true seeds. These seeds match with the morphology, physiology, and
biochemistry of the zygotic embryos.
7. Micropropagation is the best technique to transfer different species of plants to
different countries. This makes the international exchange processes easier and
reduces the risk of contamination during the transfer.

11. Disadvantages of micropropagation
1. Micropropagation requires trained manpower, sophisticated facilities, and
expensive materials which make it a pricey technique.
2. Contamination is the major problem of micropropagation. The cultures grown
in labs are very sensitive to any microorganisms.
3. Some forms of cultures have the problem of pronounced genetic variability.
When the plants are propagated through shoot tip cultures, the genetic
variability is low, and observed when they are grown using adventitious shoots.
The genetic variability can be due to aberrant cell division in the callus,
chimeral breakdown, or pre-existing genetic variability.
4. Some plants are difficult to maintain by micropropagation because they start
releasing growth inhibitory substances (like phenol) in the medium. These
compounds turn the medium into dark color (it’s called brewing of the
medium) and inhibit the growth of the tissues.
5. When cultures undergo repeated cycles of in vitro shoot multiplication, they
develop water-soaked, translucent leaves.