Genetic Engineering

Genetic engineering, also called genetic modification, is the direct human manipulation of an organism's genome using modern DNA technology. It involves the introduction of foreign DNA or synthetic genes into the organism of interest. The introduction of new DNA does not require the use of classical genetic methods, however traditional breedingmethods are typically used for the propagation of recombinant organisms.


                         The most common form of genetic engineering involves the insertion of new genetic material at an unspecified location in the host genome. This is done by isolating and copying the genetic material of interest using molecular cloning methods to generate a DNA sequence containing the required genetic elements for expression, and then inserting this construct into the host organism. Other forms of genetic engineering include gene targeting and knocking out specific genes via engineered nucleases such aszinc finger nucleases or engineered homing endonucleases.The first field trials of genetically engineered plants occurred in France and the USA in 1986, tobacco plants were engineered to be resistant to herbicides

Procedure :

Isolating gene-

First, the gene to be inserted into the genetically modified organism must be chosen and isolated.Once chosen the genes must be isolated. This typically involves multiplying the gene using polymerase chain reaction (PCR). If the chosen gene or the donor organism's genome has been well studied it may be present in a genetic library.  Once isolated, the gene is inserted into a bacterial plasmid.

Construct:

The gene to be inserted into the genetically modified organism must be combined with other genetic elements in order for it to work properly. The gene can also be modified at this stage for better expression or effectiveness. As well as the gene to be inserted most constructs contain a promoter and terminator region as well as a selectable marker gene. The constructs are made using recombinant DNA techniques, such molecular cloning.

Targeting Gene:

The most common form of genetic engineering involves inserting new genetic material randomly within the host genome. Other techniques allow new genetic material to be inserted at a specific location in the host genome or generate mutations at desired genomic loci capable of knocking out endogenous genes. The technique of gene targeting uses homologous recombination to target desired changes to a specific endogenous gene.

Transformation:

The genes to be inserted are cloned into a binary vector, which contains T-DNA and can be grown in both E. Coli and Agrobacterium. Once the binary vector is constructed the plasmid is transformed into Agrobacterium containing no plasmids and plant cells are infected. The Agrobacterium will then naturally insert the genetic material into the plant cells.About 1% of bacteria are naturally able to take up foreign DNA but it can also be induced in other bacteria. Stressing the bacteria for example, with a heat shock or an electric shock, can make the cell membrane permeable to DNA that may then incorporate into their genome or exist as extrachromosomal DNA. 

Selection:

Not all the organism's cells will be transformed with the new genetic material; in most cases a selectable marker is used to differentiate transformed from untransformed cells. If a cell has been successfully transformed with the DNA it will also contain the marker gene. 

Regeneration:

As often only a single cell is transformed with genetic material the organism must be regrown from that single cell. As bacteria consist of a single cell and reproduce clonally regeneration is not necessary. In plants this is accomplished through the use of tissue culture. Each plant species has different requirements for successful regeneration through tissue culture. If successful an adult plant is produced that contains the transgene in every cell. In animals it is necessary to ensure that the inserted DNA is present in the embryonic stem cells.

Confirmation:

The finding that a recombinant organism contains the inserted genes is not usually sufficient to ensure that the genes will be expressed in an appropriate manner in the intended tissues of the recombinant organism. To examine the presence of the gene, further analysis frequently uses PCR, Southern hybridization, and DNA sequencing, which serve to determine the chromosomal location and copy number of the inserted gene.


APPLICATIONS:

• Medicine
• Agriculture
• Research
• Industrial