PLANT TRANSFORMATION TECHNIQUE USING AGROBACTERIUM
AGRObacterium-mediated technique is the most widely used for the transformation of plants and generation of transgenic plants. The important requirements for gene transfer in higher plants through agrobacterium mediation are listed as under.
- The explants of the plant must produce phenolic compounds (e.g. autosyringone) for activation of virulence genes.
- . Transformed cells/tissues should be capable to regenerate into whole plants.
Most of the Agrobacterium-mediated plant transformations have the following basic protocol:
- 1. Development of Agrobacterium carrying the cointegrate or binary vector with the desired gene.
- 2. Identification of a suitable explants e.g. cells, protoplasts, tissues, calluses, organs.
- 3. Co-culture of explants with Agrobacterium.
- 4. Killing of Agrobacterium with a suitable antibiotic without harming the plant tissue.
- 5. Selection of transformed plant cells.
- 6. Regeneration of whole plants.
ADVANTAGES OF AGROBACTERIUM MEDIATED TRANSFORMATION.
1. THIS is a natural method of gene transfer.
2. Agrobacterium can conveniently infect any explants (cells/tissues/organs).
3. Even large fragments of DNA can be efficiently transferred.
LIMITATIONS OF AGROBACTERIUM MEDIATED TRANSFORMATION.
1. There is a limitation of host plants for Agrobacterium, since many crop plants (monocotyledons e.g. cereals) are not infected by it. In recent years, virulent strains of Agrobacterium that can infect a wide range of plants have been developed.
2.The cells that regenerate more efficiently are often difficult to transform, e,g, embryonic cells lie in deep layers which are not easy targets for Agrobacterium.