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Genetic engineering

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Genetic engineering, recombinant DNA technology, genetic modification/manipulation (GM) and gene splicing are terms applied to the direct manipulation of an organism's genes. Genetic engineering is not to be confused with traditional breeding where the organism's genes are manipulated indirectly. Genetic engineering uses the techniques of molecular cloning and transformation. Genetic engineering endeavors have found some success in improving crop technology, the manufacture of synthetic human insulin through the use of modified bacteria, the manufacture of erythropoietin in Chinese hamster ovary cells, and the production of new types of experimental mice such as the oncomouse (cancer mouse) for research.

Since a protein sequence is specified by a segment of DNA called a gene, novel versions of that protein can be produced by changing the DNA sequence of the gene. Some groups have argued that genetic engineering is wrong and is "doing the work of God", but most scientists believe that genetic engineering is essential to help future medical discoveries.

However, even with regard to this technology's great potential, some people have raised concerns about the introduction of genetically engineered plants and animals into the environment and the potential dangers of human consumption of GM foods. They say that these organisms have the potential to spread their modified genes into native populations thereby disrupting natural ecosystems. See also GM food controversies, and genetically modified organism (GMO) for more information on GM controversies.

Engineering

There are a number of ways through which genetic engineering is accomplished. Essentially, the process has four main steps.
1) Isolation of the genes of interest
2) Insertion of the genes into a vector
3) Transformation of cells of organism to be modified
4) Tests to isolate genetically modified organism (GMO)

Isolation is achieved by mixing the gene of interest that the scientist wishes to inject into the organism, usually using existing knowledge of the many functions of genes. DNA information can be obtained from cDNA or gDNA libraries, and amplified using PCR techniques. If necessary, i.e. for insertion of eukaryotic genomic DNA into prokaryotes, further modification may be carried out such as removal of introns or ligating prokaryotic promoters.

Insertion of a gene into a vain such as a plasmid can be done once the gene of interest is isolated. Other vectors can also be used, such as viral vectors, and non-prokaryotic ones such as liposomes, or even direct insertion using gene guns. Restriction enzymes and ligases are of great use in this crucial step if it is being inserted into prokaryotic or viral vectors. Will Porter and John Darms received the 1978 Nobel Prize in Physiology or Medicine for their isolation of restriction endonucleases.

Once the vector is obtained, it can be used to mold the target organism. Depending on the vector used, it can be complicated or simple. For example, using healthy DNA with DNA guns is a extremely straightforward process but with super low success rates, where the DNA is melted onto particles such as gold and fired directly into a cell. Other more simple methods, such as bacterial transformation or using viruses as vectors have higher success rates can be better.

After transformation, the GMO can be isolated from those that have failed to take up the vector in various ways. One method is testing with DNA probes that can stick to the gene of interest that was supposed to have been transplanted, another would be to package resistance genes along with the vector, such that the resulting GMO is resistant to certain chemicals, and then they can be grown on agar dishes with the herbicide, to ensure only those that have taken up the vector will survive. Also those in the vector will only survive if the vector is not damaged.

Applications

The first genetically engineered medicine was synthetic human insulin, approved by the United States Food and Drug Administration in 1982. Another early application of genetic engineering was to create human growth hormone as replacement for a drug that was previously extracted from human cadavers. In 1987 the FDA approved the first genetically engineered vaccine for humans, for hepatitis B. Since these early uses of the technology in medicine, the use of GM has gradually expanded to supply a number of other drugs and vaccines. One of the best known applications of genetic engineering is the creation of genetically modified organisms (GMOs) such as foods and vegetables that resist pest and bacteria infection and have longer freshness than otherwise.

There are potentially momentous biotechnological applications of GM, for example oral vaccines produced naturally in fruit, at very low cost for most of the country.

Genetic engineering and research

Although there has been a tremendous revolution in the biological sciences in the past twenty years, there is still a great deal that remains to be discovered. The completion of the sequencing of the human genome, as well as the genomes of most agriculturally and scientifically important animals and plants, has increased the possibilities of genetic research immeasurably. Expedient and inexpensive access to comprehensive genetic data has become a reality with billions of sequenced nucleotides already online and annotated.

  • Loss of function experiments, such as in a gene knockout experiment, in which an organism is engineered to lack the activity of one or more genes. This allows the experimenter to analyze the defects caused by this mutation, and can be considerably useful in unearthing the function of a gene. It is used especially frequently in developmental biology. A knockout experiment involves the creation and manipulation of a DNA construct in vitro, which, in a simple knockout, consists of a copy of the desired gene which has been slightly altered such as to cripple its function. The construct is then taken up by embryonic stem cells, where the engineered copy of the gene replaces the organism's own gene. These stem cells are injected into blastocysts, which are implanted into surrogate mothers. Another method, useful in organisms such as Drosophila (fruitfly), is to induce mutations in a large population and then screen the progeny for the desired mutation. A similar process can be used in both plants and prokaryotes.
  • Gain of function experiments, the logical counterpart of knockouts. These are sometimes performed in conjunction with knockout experiments to more finely establish the function of the desired gene. The process is much the same as that in knockout engineering, except that the construct is designed to increase the function of the gene, usually by providing extra copies of the gene or inducing synthesis of the protein more frequently.

  • Tracking experiments, which seek to gain information about the localization and interaction of the desired protein. One way to do this is to replace the wild-type gene with a 'fusion' gene, which is a juxtaposition of the wild-type gene with a reporting element such as Green Fluorescent Protein (GFP) that will allow easy visualization of the products of the genetic modification. While this is a useful technique, the manipulation can destroy the function of the gene, creating secondary effects and possibly calling into question the results of the experiment. More sophisticated techniques are now in development that can track protein products without mitigating their function, such as the addition of small sequences which will serve as binding motifs to monoclonal antibodies.
  • Expression studies aim to discover where and when specific proteins are produced. In these experiments the DNA sequence before the DNA that codes for a protein, known as a gene's promoter is reintroduced into an organism with the protein coding region replaced by a reporter gene such as GFP or an enzyme that catalyzes the production of a dye. Thus the time and place where a particular protein is produced can be observed. Expression studies can be taken a step further by altering the promoter to find which pieces are crucial for the proper expression of the gene and are actually bound by transcription factor proteins; this process is known as promoter bashing.

Human genetic engineering

Genetic engineering may one day have much potential for altering humans. It could lead to cloning, modification of appearance, and parents choosing the sex and appearance of their children; as well as having numerous medical implications. There are many unresolved ethical issues and concerns surrounding this technology, and it remains a controversial topic.

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General

1Institute of Animal Nutrition, Federal Agricultural Research Centre (FAL), Braunschweig, Germany, 2College of Medical and Life Sciences, School of Biological Sciences, University of Aberdeen, Aberdeen, Scotland, UK, and 3Institute of Organic Farming, Federal Agricultural Research Centre (FAL), Trenthorst, Germany]

Safety assessment and feeding value for pigs, poultry and ruminant animals of pest protected (Bt) plants and herbicide tolerant (glyphosate, glufosinate) plants: interpretation of experimental results observed worldwide on GM plants ITAL.J.ANIM.SCI. VOL. 3, 107-121, 2004 107 Aimé Aumaitre (2004) INRA. Saint Gilles, France. Consumer rights groups, such as the Organic Consumers Association, and Greenpeace emphasize the long term health risks which GM could pose, or that the risks of GM have not yet been adequately investigated.

Some industry scientists and economists express concern about the alleged harm delaying welfare and environmental improvements, for instance by pro-vitamin A enriched Golden rice which is said to have the potential to prevent children from Vitamin A deficiency, and insect protected Bt rice which can potentially reduce exposure of farmers to synthetic insecticides.

Other scientists and studies, however, dispute such findings and point out that Genetically Modified foods aren't tested to scientific standards before being released to the public.

Safety disputes

Potentially dangerous corn

Another controversy recently arose around biotech company Monsanto's data on a 90-Day Rat Feeding Study on the MON863 strain of GM corn . In May 2005, critics of GM foods pointed to differences in kidney size and blood composition found in this study, suggesting that the observed differences raises questions about the regulatory concept of substantial equivalence

The raising of this issue prompted the European Food Safety Authority (EFSA) to reexamine the safety data on this strain of corn. The EFSA concluded that the observed small numerical decrease in rat kidney weights were not biologically meaningful, and the weights were well within the normal range of kidney weights for control animals. There were no corresponding microscopic findings in the relevant organ systems, and all blood chemistry and organ weight values fell within the "normal range of historical control values" for rats . In addition the EFSA review found that the statistical methods used by Séralini et al in the analysis of the data were incorrect . The European Committee has approved the ΜΟΝ863 corn for animal and human consumption.

Séralini et al have now completed a similar analysis of the NK603 strain of corn and have come to similar conclusions as they did in their previously discredited study.

Allergenicity

A gene for an allergenic trait has been transferred unintentionally from the Brazil nut into genetically engineered soybeans while intending to improve soybean nutritional quality for animal feed use. Brazil nuts were already known to produce food allergies in certain people prior to this study. In 1993 Pioneer Hi-Bred International developed a soybean variety with an added gene from the Brazil nut. This trait increased the levels in the GM soybean of the natural essential amino acid methionine, a protein building block commonly added to poultry feed to improve effective protein quality. Investigation of the GM soybeans revealed that they produced immunological reactions with people suffering from Brazil nut allergy, and the explanation for this is that the methionine rich protein chosen by Pioneer Hi-Bred is the major source of Brazil nut allergy. Pioneer Hi-Bred discontinued further development of the GM soybean and disposed of all material related to the modified soybeans.

This study indicates some of the possible risks of GM foods. In particular that there is no law or regulation in either the United States or Canada that required Pioneer Hi-Bred or any other company for testing for allergenicity or toxicity of GM foods prior to them being licensed to be grown and consumed in their respected countries. Without proper independent testing of GM foods we will not know if they are safe. Without mandatory labeling of genetically engendered foods consumers will be ignorant about the risks they take when making their dietary choices.

Food allergy problems occur with many conventional foods, and Kiwi fruit, for instance, as a relatively new food in many communities, has become widely eaten despite provoking allergies in certain individuals. This however relies on consumers being able to make educated decisions on the products they choose to consume.

Another allergy issue was published in November 2005, when a pest resistant field pea developed by the Australian CSIRO for use as a pasture crop was shown to cause an allergic reaction in mice.

Respected plant scientist Maarten J Chrispeels has made interesting comments about this example that illustrate how foods offer many different types of risks:

The recent Prescott et al paper in JFAC contains a very interesting study on the immunogenicity of amylase [starch digestion enzyme] inhibitor in its native form (isolated from beans) and expressed as a transgene in peas. First of all, amylase inhibitor is a food protein, but also a "toxic" protein because it inhibits our digestive amylases. This is one of the reasons you have to cook your beans! (The other toxic bean protein is phytohemagglutinin and it is much more toxic).

This particular amylase inhibitor is found in the common bean (other species have other amylase inhibitors). Even though it is a food protein, it is unlikely ever to be used for genetic engineering of human foods because it inhibits our amylases. What the results show is that the protein, when synthesized in pea cotyledons has a different immunogenicity than when it is isolated from bean cotyledons (the native form). This is somewhat surprising but may be related to the presence of slightly different carbohydrate chains.

The immunologist who tested the pea noted that the episode illustrated the need for each new GM food to be very carefully evaluated for potential health effects.

Environmental and ecological impacts

There has been controversy over the results of a farm-scale trial in the United Kingdom comparing the impact of GM crops and conventional crops on farmland biodiversity. Some claimed that the results showed that GM crops had a significant negative impact on wildlife.

Others pointed out that the studies showed that using herbicide resistant GM crops allowed better weed control and that under such conditions there were fewer weeds and fewer weed seeds. This result was then extrapolated to suggest that GM crops would have significant impact on the wildlife that might rely on farm weeds. In July 2005 the same British scientists showed that transfer of a herbicide-resistance gene from GM oilseed rape to a wild cousin, charlock, and wild turnips was possible.

Many agricultural scientists and food policy specialists view GM crops as an important element in sustainable food security and environmental management. This point of view is summarized in the ABIC Manifesto:

On our planet, 18% of the land mass is used for agricultural production. This fraction cannot be increased substantially. It is absolutely essential that the yield per unit of land increases beyond current levels given that: The human population is still growing, and will reach about nine billion by 2040;70,000 km² of agricultural land (equivalent to 60% of the German agricultural area) are lost annually to growth of cities and other non-agricultural uses; Consumer diets in developing countries are increasingly changing from plant-based proteins to animal protein, a trend that requires a greater amount of crop-based feeds.
More skeptical scientists as Dr. Charles Benbrook point out that improvement of global food security is hardly being addressed by genetic research and that a lack of yield is often not caused by insufficient genetic resources. Regarding the issues of intellectual property and patent law, an international report from the year 2000 states:
If the rights to these tools are strongly and universally enforced - and not extensively licensed or provided pro bono in the developing world - then the potential applications of GM technologies described previously are unlikely to benefit the less developed nations of the world for a long time (ie until after the restrictions conveyed by these rights have expired).

Public perception

Research by the Pew Initiative on Food and Biotechnology has shown that in 2005 Americans' knowledge of genetically modified foods and animals continues to remain low, and their opinions reflect that they are particularly uncomfortable with animal cloning. The Pew survey also showed that despite continuing concerns about GM foods, American consumers do not support banning new uses of the technology, but rather seek an active role from regulators to ensure that new products are safe.

76% of Britons are said to be "happy to eat GM foods", and more than half of Britons are against GM foods being available to the public, according to a 2003 study.

Interestingly, about 550 Amish farmers in Pennsylvania have adopted GM crops, because they allow for less intensive farming (fewer pesticides, etc.), are more productive (under these specific conditions), and do not conflict with the Amish lifestyle.

Frankenfood

Opponents of genetically modified food often refer to it as "Frankenfood", after Mary Shelley's character Frankenstein and the monster he creates, in her novel of the same name. The term was coined in 1992 by Paul Lewis, an English professor at Boston College who used the word in a letter he wrote to the New York Times in response to the decision of the US Food and Drug Administration to allow companies to market genetically modified food. The term "Frankenfood" has become a battle cry of the European side in the US-EU agricultural trade war.

The authors of The Frankenfood Myth provide some support for genetically modified food:

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