TRANSGENIC PLANTS
Techniques to produce Genetic Modified Crop
Transformation with Agrobacterium Tumefaciens
Agrobacterium tumefaciens bacterium, which is a soil pathogen, acts as the vector for the antibody- or cytokine-producing gene. The genetic material that is introduced is called T DNA (transferred DNA) which is located on a Ti plasmid. A Ti plasmid is a circular piece of DNA found in almost all bacteria. This natural ability to alter the plant’s genetic makeup was the foundation of plant transformation using Agrobacterium. Agrobacterium tumefaciens is a valuable gene delivery system because it is able to carry any gene of interest within the T-complex, and insert the gene into the target plants DNA with a high degree of success. Therefore the T-DNA strand can be replaced by a gene of interest which will be inserted automatically into the host plant nucleus with a high degree of success and with little human intervention. Thus, A. tumefaciens is an ideal vehicle for transferring DNA into eukaryotic cells. It is widely used in transforming dicotyledonous plants such as tomatoes, potatoes, soybeans, apples, Arabidopsis and tobacco.
Currently, Agrobacterium-mediated transformation is the most commonly used method for plant genetic engineering because of relatively high efficiency. Initially it was believed that this Agrobacterium only infects dicotyledonous plants, but it was later established that it can also be used for transformation of monocotyledonous plants such as rice.
During transformation, several components of the Ti plasmid enable effective transfer of the genes of interest into the plant cells. These include:
· T-DNA border sequences, which demarcate the DNA segment (T-DNA) to be transferred into the plant genome
· vir genes (virulence genes), which are required for transferring the T-DNA region to the plant but are not themselves transferred, and
· modified T-DNA region where the genes that cause crown gall formation are removed and replaced with the genes of interest.
The Agrobacterium-mediated transformation process involves a number of steps that has been summarized in the Figure 1.
Figure 1: Summary of Agrobacterium-mediated plant transformation
Figure 2: Agrobacterium-mediated plant transformation
Agrobacterium tumefaciens infects only dicotyledonous plants and many economically important plants such as cereals, remained accessible for genetic manipulation during long time. For these cases, many alternative direct transformation methods have been established, such as gene gun technology, protoplast fusion technique and antisense technology.