Genetic engineering: artificial manipulation and alteration of genes.
Process of Genetic Engineering:
- Isolation: process of removing DNA from cells.
Isolation involves using detergents to break open the cell membranes and nuclear membranes to release the DNA.
2. Cutting and ligation
- Cutting: removal of a gene from a piece of DNA using a restriction enzyme.
Enzymes called restriction enzymes (type of catabolic enzyme) are added to the DNA to cut out the desired gene. The same restriction enzyme is used to cut open a vector – into which the gene will be inserted (see diagram below).
It is important to use the same restriction enzyme because then the gene will attach to the vector due to the presence of complementary ends – as both pieces of DNA were cut using the same enzyme. The gene inserts into the vector (such as a plasmid or a virus) using DNA ligases (type of anabolic enzyme).
- Vector: piece of DNA (such as a bacterial plasmid) that will carry a gene of interest into a host cell.
- Ligation: joining of a gene to a vector using DNA ligase.
Once the gene has been inserted into the vector, the new piece of DNA is called ‘recombinant DNA‘ as it contains DNA from two species.
- Recombinant DNA: piece of genetically modified DNA that contains DNA from two or more different species.
3. Introduction of base sequence changes/ Transformation/ Transfection/ Transduction:
Transformation is the term used to generally describe the uptake of recombinant DNA into a host cell.
- Transformation: uptake of recombinant DNA into a bacterial cell.
- Transfection: uptake of recombinant DNA into a eukaryotic cell.
- Transduction: uptake of recombinant viral vector by any cell.
4. Selection and cloning
Not all cells take up the recombinant DNA. Geneticists want only cells that have been ‘transformed’ with the recombinant DNA. In order to do this they select cells that have taken up the DNA by killing cells that have not. Recombinant DNA usually contains an antibiotic-resistant gene that gives any cells that take it up the ability to survive in the presence of a strong antibiotic. Any cells that do not take up the recombinant DNA are killed by the antibiotic.
- Selection: process of killing any cells that did not take up the recombinant DNA.
Once transformed cells have been selected for, very few may remain. They need to be reproduced. The technique of cloning (or simply allowing the cells to reproduce themselves) is used.
- Cloning: process of producing identical copies of a cell.
Once a workable number of transformed cells have been produced, they are stimulated to produce their product.
- Expression: stimulation of a cell to produce the product of a particular gene.
Applications of genetic engineering:
Corn in Spain has been genetically modified to be resistant to herbicide and insects.
Mice have been genetically modified to glow green. A green fluorescent protein (GFP), originally discovered in jellyfish, has been successfully expressed in mice by inserting the gene responsible for producing GFP into mice. It is hoped this can be applied to human cancer cells.
E. coli bacteria have been genetically modified with human genes, such as the human insulin gene. The bacteria produce human insulin, which is then purified and used to treat type I diabetes.
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