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| Genetic Engineering A Comprehensive Review Learn the Fundamentals of Biotechnology 1. What is biotechnology? Biotechnology is the application of biological knowledge to obtain new techniques, materials and compounds of pharmaceutical, medical, agrarian, industrial and scientific use, i.e., of practical use. The pioneer fields of biotechnology were agriculture and the food industry but nowadays many other practical fields use its techniques. 2. What is genetic engineering? Genetic engineering is the use of genetic knowledge to artificially manipulate genes: It is one of the fields of biotechnology. 3. At the present level of the biotechnology what are the main techniques of genetic engineering? The main techniques of genetic engineering today are: the recombinant DNA technology (also called genetic engineering itself) in which pieces of genes from an organism are inserted into the genetic material of another organism producing recombinant beings; the nucleus transplantation technology, popularly known as “cloning”, in which a nucleus of a cell is grafted into a enucleated egg cell of the same species to create a genetic copy of the donor (of the nucleus) individual; the technology of DNA amplification, or PCR (polymerase chain reaction), that allows millions replications of chosen fragments of a DNA molecule. The recombinant DNA technology is used to create transgenic organisms, like mutant insulin-producing bacteria. The nucleus transplantation technology is in its initial development but it is the basis, for example, of the creation of “Dolly” the sheep. PCR has numerous practical uses, as in medical tests to detect microorganisms present in blood and tissues, DNA fingerprint and obtainment of DNA samples for research. 4. What are restriction enzymes? How do these enzymes participate in the recombinant DNA technology? Restriction enzymes, or restriction endonucleases, are enzymes specialized in the cutting of DNA fragments each acting upon specific sites of the DNA molecule. Restriction enzymes are used in the recombinant DNA technology to obtain with precision pieces of DNA molecules to be later inserted into other DNA molecules cut by the same enzymes. Image Diversity: recombinant DNA technology restriction enzymes 5. What are DNA ligases? How do these enzymes participate in the recombinant DNA technology? DNA ligases are enzymes specialized in tying the complementary DNA chains that form the DNA double helix. These enzymes are used in the recombinant DNA technology to insert pieces of DNA cut by restriction enzymes into other DNA molecules submitted to the action of the same endonucleases. Image Diversity: DNA ligases 6. What are plasmids? Plasmids are circular DNA molecules present in the genetic material of some bacteria. They may contain genes responsible for bacterial resistance to some antibiotics and for proteins that cause virulence (pathogenic hostility). Image Diversity: plasmid 7. How is genetic engineering used to create bacteria capable of producing human insulin? In the production of human insulin by bacteria the human insulin gene is incorporated into the genetic material of these microorganisms. The mutant bacteria multiply forming lineages of insulin-producing bacteria. In bacteria there are circular strands of DNA called plasmids, minichromosomes which act as an accessory to the main DNA. To create a mutant bacteria capable of producing insulin a plasmid is submitted to the action of restriction enzymes (restriction endonucleases) specialized in cutting DNA fragments. The once circular plasmid is open by the restriction enzyme. The same enzyme is used to cut a human DNA molecule containing the insulin gene. The piece of human DNA containing the insulin gene then has its extremities bound to the plasmid with the help of DNA ligases. The recombinant plasmid containing the human insulin gene is then inserted into the bacteria. Another human hormone already produced by recombinant bacteria is GH (somatotropin, or growth hormone). The insertion of DNA molecules into cells of an individual is also the method of the gene therapy, a promising treatment for genetic diseases. In gene therapy cells from an organism deficient in the production of a given protein receive (by means of vectors, e.g., virus) pieces of DNA containing the protein gene and they then begin to synthesize the protein. 8. What is cloning? Cloning is the making of an organism genetically identical to another by means of genetic engineering. The basis of cloning is the nucleus transplantation technology. A nucleus from a cell is extracted, generally from an embryonic (not differentiated) cell and this nucleus is inserted into a previously enucleated reproductive cell (in general an egg cell); the egg is then implanted in the organ where the embryonic development will take place. If embryonic development occurs the new organism will have identical genetic patrimony to the organism owner of the cell whose nucleus was used in the transplantation. Image Diversity: genetic cloning Dolly the sheep 9. What is PCR? How does PCR works? PCR, polymerase chain reaction, is a method to synthesize many copies of specific regions of a DNA molecule known as target-regions. Its inventor, Kary Mullis, won the Nobel prize for Chemistry in 1993. First, the DNA to be tested is heated to cause the double helix to rupture and the polynucleotide chains to be exposed. Then small synthetic sequences of DNA known as primers and containing nucleotide sequences similar to the sequences of the extremities of the region to be studied (for example, a region containing a known gene exclusive of a given organism) are added. The primers paired with the original DNA in the extremities of the gene to be amplified. Enzymes known as polymerases, that catalyze DNA replication, and nucleotide supply are added. The primers then are completed and the chosen region is replicated. In the presence of more primers and more nucleotides millions of copies of that specific region are generated. (PCR is very sensitive even using a minimal amount of DNA). Image Diversity: polymerase chain reaction 10. What is the fact of Molecular Biology on which DNA fingerprint is based? DNA fingerprint, the method of individual identification using DNA, is based on the fact that the DNA of every individual (with exception of identical twins and individual clones) contains nucleotide sequences exclusive to each individual. Although normal individuals of the same species have the same genes in their chromosomes, each individual has different alleles and even in the inactive portions of the chromosomes (heterochromatin) there are differences in nucleotide sequences among individuals. Image Diversity: DNA fingerprint 11. Why are the recombinant DNA technology and the nucleus transplantation technology still dangerous? The recombinant DNA technology and the nucleus transplantation technology (cloning) are extremely dangerous since they are able to modify, in a very short time, the ecological balance that evolution has taken millions of years to create on the planet. During the evolutionary process, under the slow and gradual action of mutations, genetic recombinations and of natural selection species emerged and were modified and genetic patrimonies were formed. With genetic engineering however humans can mix and modify genes, making changes of unpredictable long term consequences, risking creating new plant or animal diseases, new types of cancers and new disease outbreaks. It is a field as potentially dangerous as the manipulation of nuclear energy. 12. What is the main moral problem about the cloning of human individuals? Besides biological perils, a very serious moral problem involves the nucleus transplantation technology concerning humans: an individual right of a human being is offended when a man or woman is made as a copy of another. Since it is impossible to first ask if the person to be generated wants or not to be a genetic copy of another person, certainly the most important human right is being offended, one's individual freedom, when a human being is obliged to be a genetic copy of another. It is indeed a danger to democracy, whose most basic principle must be nonviolation of individual freedom. |