Genetically modified organisms are also known as Recombinant-DNA. During the 1970s and 1980s, the practical application of microorganisms expanded almost beyond imagination with the development of new, artificial techniques for making recombinant DNA. Although natural recombination makes it possible for closely related organisms to exchange genes, the new techniques make it possible to transfer genes between completely unrelated species. So very powerful are these techniques that the term recombinant DNA is now widely understood to refer to DNA that has been artificially manipulated to combine genes from two different sources. A gene from a vertebrate animal, such as a human, can be inserted into the DNA of a bacterium, or a gene from a virus into yeast. In many cases, the recipient can then be made to express the gene, which may code for a commercially useful product. For example, bacteria with genes for human insulin are now being used to produce insulin for treating diabetes. Also, a vaccine for hepatitis B is being made by yeast carrying a gene for part of the hepatitis virus; the yeast produces a viral protein. Scientists hope that such a protein may prove to be an effective vaccine, eliminating the need to use whole viruses as in conventional vaccines. In a lighter vein, scientists working to refine recombinant DNA procedures in plants have succeeded in transferring a firefly gene to a tobacco plant and getting it to function (Barton, 2000).
The new recombinant-DNA technique can also be used to make thousands of copies of the same DNA molecule—to amplify DNA, thus generating sufficient DNA for various kinds of experimentation and analysis. This has found practical application in criminology. DNA from a few new human cells of blood or semen found at a crime scene can be amplified and then compared with the DNA of suspects. Since each person’s DNA is unique, the procedure has great potential usefulness.
Artificial gene manipulation is popularly known as genetic engineering. In fact, the term biotechnology, which had been generally defined to include all industrial applications of microorganisms, has increasingly become identified in the public mind as the industrial use of genetically engineered microorganisms (Weaver, 2004).
Genetic engineering was made possible by the discovery of bacterial enzymes that can be used to cut DNA from different sources into pieces that are easy to recombine in vitro (in vitro means “in glass,” that is, in a test tube rather than inside a living organism). Also essential was the development of methods for inserting this recombinant DNA into a cell. If a mosquito carrying the virus for yellow fever bites and infects a human, the mosquito in known as a “disease vector” because it can transmit the virus from one host to another (Weaver, 2004). The term vector has been adopted to describe a self-replicating DNA molecule that is used as a carrier to transmit a gene from one organism into another. It is also known as a cloning vector.
A. Making a Gene Product
We have just seen that one way of identifying cells carrying a particular gene is by assaying for the gene product. Such products are themselves, of course, a frequent objective of genetic engineering. Most of the earliest work in genetic engineering made use of E. coli to synthesize the gene products. E. coli has the advantage that researchers are very familiar with this easily grown organism and with its genetics. It also has several disadvantages. Like other gram-negative bacteria, it produces endotoxins as part of its outer layer. Since endotoxins cause fever and shock in animals, their accidental presence in products intended for use in humans would be in a serious problem. Another disadvantage of E. coli is that it does not usually secrete protein products. To obtain a product, cells must usually be broken open and the product purified from the resulting “soup” of cell components (Barton, 2000).
Recovering the product from such a mixture is expensive when done on an industrial scale. It is more economical to have an organism secrete the product so that it can be recovered continuously from the growth medium. One approach has been to link the product to a natural E. coli protein that the bacterium does secrete. This approach has been used to produce insulin. Certain gram-positive bacteria, such as Bacillus subtilis, are more likely to secrete their products and are often preferred industrially for that reason.
Another microbe that shows promise as a vehicle for the expression of genetically engineered genes is baker’s yeast, Saccharomyces cerevisiae. Its genome is only about four times larger than that of E. coli and is probably the best understood eukaryotic genome. Yeasts may carry plasmids; their cell walls can readily be removed to introduce plasmids carrying engineered genes. As eukaryotic cells, yeasts may be more successful in expressing foreign eukaryotic genes than bacteria. Furthermore, yeasts are likely to continuously secrete the product. Because of all these factors, yeasts have become the workhorse of eukaryotic cells. Yeasts also have a psychological advantage in the marketplace. Bacteria and viruses are, unfairly, associated in the public’s mind with diseases, whereas yeasts have a much more benign image, thanks to their association with baking, brewing, and wine-making (Barton, 2000).
Animal viruses have also been used in making engineered gene products, primarily in the field of vaccine production. For example, scientists have been able to insert genes for the surface proteins of pathogenic microbes into the generally harmless vaccinia virus. The result is a sort of “sheep in wolf’s clothing,” a virus that has the external proteins of a pathogen but dies not cause disease. When an animal host is infected with the engineered virus, the host’s immune system recognizes these proteins as foreign and, in response, develops an immunity that can protect it against the actual pathogen. Because the vaccinia virus is unusually large and has room for several extra genes, a genetically engineered vaccinia virus might theoretically be used as a vaccine for several diseases simultaneously (Weaver, 2004).
Mammalian cells in culture, even human cells, can be used much like bacteria to produce genetically engineered products. Scientists have developed effective methods for growing mammalian cells in culture as hosts for growing viruses. In genetic engineering, mammalian cells are often the best suited to make protein products for medical use; these products include hormones, lymphokines (which regulate cells of the immune system), and interferon (a natural antiviral substance that is also used to treat some cancers) (Anderson & Diacumakos, 2001). While plant cells can also be grown in culture, altered by recombinant-DNA techniques, and then used to generate genetically engineered plants. Such plants may prove useful as sources of valuable plant products, such as alkaloids (the painkiller codeine, fro example) and the isoprenoids that are the basis of synthetic rubber.
A. Evidence of Pro and Con
A strong opponent of spontaneous generation, the Italian physician Francesco Redi, set out in 1668 (even before van Leeuwenhoek’s discovery of microscopic life) to demonstrate that maggots do not arise spontaneously from decaying meat, as was commonly believed. Redi filled three jars with decaying meat and sealed them tightly. Then he arranged three other jars similarly but left them open. Maggots appeared in the open vessels after flies entered the jars and laid their eggs. But the sealed containers showed no signs pf maggot larvae. Still, Redi’s antagonists were not convinced; they claimed that fresh air was needed for spontaneous generation. So Redi set up a second experiment, in which three jars were covered with a fine net instead of being sealed. No larvae appeared in the gauze-covered jars, even though air was present (Weaver, 2004). Maggots appeared only if flies were allowed to leave their eggs on the meat.
Redi’s results were a serious blow to the long-held belief that large forms of life could rise from nonlife. However, many scientists still believed that small organisms, such as van Leeuwenhoek’s “animalcules,” were simple enough to be generated from nonliving materials.
B. The Role of Microorganisms in Food Production/Foodservice Industry
In the latter part of the nineteenth century, microbes used in food production were grown in pure culture for the first time. This development quickly led to improved understanding of the relationship between specific microbes and their products and activities. This period can be considered the beginning of industrial food microbiology. For example, once it was understood that a certain yeast grown under certain bacteria could spoil the beer; brewers were better able to control the quality of their products. Specific industries became active in microbiological research and selected certain microbes for their special qualities. For example, the brewing industry extensively investigated the isolation and identification of yeast and selected those that were able to produce more alcohol (Whitman, 2000).
Closely related organisms can exchange genes in natural recombination. Genes can be transferred between unrelated species via laboratory manipulations called genetic engineering. Recombinant DNA refers to DNA that has been artificially manipulated to combine genes from two different sources. Biotechnology includes all industrial applications of microorganisms as well as industrial uses of genetically engineered cells. A DNA molecule used to carry a desired gene from one organism to another is called a vector.
A restriction enzyme recognizes and cuts only one particular nucleotide sequence in DNA. Some restriction enzymes produce sticky ends, short stretches of single-stranded DNA at the ends of the DNA fragments. Fragments from DNA produced by the same restriction enzyme will spontaneously join by hydrogen-bonding. DNA ligase can covalently link the DNA backbones.
Anderson, W.F. and E.G. Diacumakos. “Genetic engineering in mammalian cells.” Scientific American 245 (1): 10121, July 2001. Describes methods of inserting genes into mammalian cells with the goal of curing inherited diseases.
Barton, J.H. “patenting life.” Scientific American 246 (3):40-46, March 2000. This article the legal issues of ownership of genetically engineered cells.
Weaver, R.F. “Changing life’s genetic blueprint.” National Geographic Magazine 166 (6): 818-847, December 2004. Well illustrated with diagrams and photographs explaining the basics of genetic engineering.
Whitman, D. Genetically Modified Foods: Harmful or Helpful? ProQuest. http://www.csa.com/discoveryguides/gmfood/overview.php