Micropropagation

Micropropagation is an advanced vegetative propagation technology for producing a large number of transplants in a limited time and space.
STAGES
Stage 0 — Mother Plant Selection:
Stage I — Establishment of Aseptic Culture:
Stage II — Multiplication of shoots:
Stage III — In Vitro Rooting:
Stage IV — Transplantation or Hardening:
APPLICAIONS

1. MICROPOPAGATION By: Purbasa Maiti MSc Microbiology

2. DEFINITION Micropropagation is an advanced vegetative propagation technology for producing a large number of transplants in a limited time and space. Clones of mother plant

3. STAGES Stage 0 — Mother Plant Selection : Stage I — Establishment of Aseptic Culture : Stage II — Multiplication of shoots : Stage III — In Vitro Rooting : Stage IV — Transplantation or Hardening:

4. Stage 0 — Mother Plant Selection : selection of suitable mother plant is crucial disease free mother explant is selected High potential to grow and divide Certain growth parameters of mother plant can be improved by pretreatment of mother explant before initiation of cultures.

5. Stage I — Establishment of Aseptic Culture The main steps involved are :- Surface sterilisation Establishment of explant on appropriate culture medium. Plant tissues are commonly associated with bacteria and fungus . Therefore, adequate sterili­zation methods are employed to eliminate microorganisms from explant

6. In tissue culture media, presence of sucrose as a carbon source encourages their growth.

7. The main objective of the stage II is the multiplication of shoot to give rise new individual plant . I nvolves three routinely used methods like: (a) Callus mediated multiplication (b) Adventitious shoots mediated multiplication (c) By apical or axillary shoots Stage II — Multiplication of shoots:

8. (a) Callus Mediated Shoot Multiplication: Innumerable number of plants can be grown by callus culture. Ratio of auxins to cytokinins could play a decisive role in re-differentiation of shoots from the callus. Callus mediated shoot multiplication involves certain drawbacks . There might be gradual decline or total loss of regeneration potential of callus cells. callus mediated regenerated plants generally exhibit genetic variations which reflects on variations of morphological characters.

9. (b) In vitro Multiplication of Adventitious Shoots: In this method, adventitious shoots arise directly from the tissues of the explant like from stem, tuber, bulb, leaf tissues other than leaf axils remains (organogenesis). It does not involve callus mediated regeneration.

10. (c) Axillary Shoot Proliferation : Axillary shoots developed from axillary buds present in the axils of each leaf. In leaf axils unsprouted status of axillary buds is due to apical dominance exhibited by growing shoot tip region at the top. Synthesis of auxin in apical shoot meristem is probably responsible for apical dominance. However, apical dominance can be reversed by synthesis of cytokinin in axillary buds or entry of cytokinin into the buds. The culture media are generally enriched with very high concentration of cytokinin by s ynthetic cytokinin like BAP and kinetin play a prominent role in releasing unsprouted axillary buds and its further proliferation.

11. Stage III — In Vitro Rooting : Shoots or plantlets obtained during stage not contain roots and fails to grow in soil. Therefore, adequate steps are taken to grow individual plantlets that can carry out photosynthesis and survive without external supply of carbohydrate. Therefore, in vitro grown shoots must be transferred to a rooting media. There is a clear distinction between rooting media from shooting media. In vitro rooting can be accomplished by adding auxins to the culture media. Supplying riboflavin and other adjuvants enhances root induction process.

12. Stage IV — Transplantation or Hardening : Process of transferring plantlets from aseptic culture medium to soil. Immediate transfer of tissue culture plants into soil is detrimental for survival of regenerated plants due to desiccation, infection, and light temperature shock . Excess of water loss was noticed from the leaves of plants immediately after transplantation. High rate of water loss due to lack of protective cuticle layer. In order to overcome these problems regenerated plants are subjected to hardening.

13. Steps for hardening:

14. A pplications: Small amounts of plant tissue are sufficient for the production of millions of clones in a year using micropropagation. Plants in large numbers can be produced in a short period. Large amounts of plants can be maintained in small spaces. This helps to save endangered species and the storage of germplasm . The micropropagation method produces plants free of diseases. Fast international exchange of plant material without the risk of disease introduction is provided . Through somatic embryogenesis production of synthetic artificial seeds is becoming popular nowadays.