Recombinant DNA is a DNA molecule artificially made from different sources. The different portions are combined into a single molecule. Genetic engineering with recombinant DNA is the re-designing of organisms at the gene level
Recombinant DNA is a DNA molecule artificially made from different sources. The different portions are combined into a single molecule. Genetic engineering with recombinant DNA is the re-designing of organisms at the gene level with the purpose of achieving specific goals.
The first step in genetic engineering usually involves inserting a short piece of 'foreign' DNA into the DNA of a host organism. In a procedure known as gene splicing, a piece of human DNA for example may be inserted into a bacterium. The DNA ring of the bacterial plasmid is broken open to insert the human DNA. The bacterium then acquires the ability to synthesise the protein for which the foreign DNA codes.
Gene splicing relies on a group of enzymes called restriction endonucleases.
One of these would cut a bacterial plasmid open at a specific site that is determined by the sequence of bases in that region. These bases are a DNA code, made up of adenine (A), thymine (T), guanine (G) and cytosine (C). The four bases are paired on the DNA molecule in a very specific manner, A always with T and G always with C. Connecting the base pairs are alternating sugar and phosphate units, forming a structure that resembles a ladder. The ladder is three-dimensional and takes the form of two strands twisted into a long spiral known as the 'double helix'. The same enzyme will cut foreign DNA wherever an identical base sequence occurs.
Most restriction endonucleases have the characteristic that they cut the two strands of DNA at slightly different points. The result is that each end of the foreign DNA segment has a short row of unpaired bases which match the complimentary bases at each end of the opened up plasmid. These are known as 'sticky ends'. In a suitable condition the unpaired bases of the foreign DNA and the plasmid join up, so the foreign DNA is incorporated into the plasmid. The bonding is made secure by another enzyme known as DNA ligase. Once in position, the foreign DNA replicates along with the rest of the plasmid every time the cell divides.
Another enzyme often used in genetic engineering is reverse transcriptase, this makes DNA using mRNA as a substrate. For example, cells of the human pancreas that produce large amounts of the hormone insulin, can have their mRNA extracted. Using reverse transcriptase this mRNA can be used to make single strands of DNA, sometimes known as complementary DNA (cDNA). Single stranded cDNA can then be made into the appropriate double-stranded DNA by the enzyme DNA polymerase.
Once a bacterium has taken up a piece of foreign DNA successfully, it may divide repeatedly and produce a large population of bacterial cells which contain both their original DNA and replicas of the foreign DNA, this is known as genetic cloning.
A further technique to move genes from one species to another is to simply fire them from a tiny gun! The DNA is mixed with metal particles usually made of tungsten then fired into an organism or a tissue culture of cells of the organism. The main advantage of this method is its simplicity and it is therefore widely used genetic engineering in plants, however the damage which may be caused as a result of the 'firing' means that only a small proportion of the cells take up the foreign DNA.
DNA can also be injected directly ...
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A further technique to move genes from one species to another is to simply fire them from a tiny gun! The DNA is mixed with metal particles usually made of tungsten then fired into an organism or a tissue culture of cells of the organism. The main advantage of this method is its simplicity and it is therefore widely used genetic engineering in plants, however the damage which may be caused as a result of the 'firing' means that only a small proportion of the cells take up the foreign DNA.
DNA can also be injected directly into the nucleus of an embryonic cell, this is the way that most genetic engineering in animals is conducted. This method ensures that at least some of the cells of the organism take up the foreign DNA.
Viruses can also be used to carry DNA from one species to another. The disadvantage of this however lies in the fact that the virus itself may cause problems and for this reason genetic engineers usually disable the virus in some way.
Risks and benefits of Genetic Engineering
Until the advent of genetic engineering, the insulin for insulin-dependant diabetics came from cattle (bovine insulin) and pigs (porcine insulin). Neither bovine nor porcine insulin is identical to that found in humans. Although all three hormones are proteins with fifty-one amino acids, bovine insulin differs from human insulin by three amino acids and porcine insulin by one.
Bovine and porcine insulin are obtained from the pancreases of cattle and pigs slaughtered for food. Although these kinds of insulin have allowed literally millions of people, who would otherwise have died, to lead relatively healthy lives, there are problems with their use. For instance there amino acid sequences differ from humans, which can mean that some diabetics develop reactions to them. A second problem is that if the animals from which the insulin is obtained were contaminated in some way, perhaps with a virus, the contaminant may be passed onto the diabetic. In addition to these, some people have ethical objections for not wishing to use insulin from cattle or pigs.
Synthetic human insulin has been available through genetic engineering. Sometimes bacteria are used, sometimes yeast, but in both cases, the basic idea is the same, the gene that produces human insulin is spliced into the micro-organism, which in turn multiplies and produces human insulin. This is then collected and purified then given to diabetics to inject.
A certain level of risk is involved in using human insulin in that some diabetics who have previously used bovine or porcine insulin and now use genetically modified human insulin have found its effects are slightly different. The most important one of these is it may be more difficult for a person to realise when his or her blood sugar level is becoming dangerously low. In a few cases this may have led to the death of diabetics. (Advanced Biology- M Roberts, M Reiss, G Monger published 2000)
Gene therapy means using genetic engineering to change genetic make-up for medical benefit. Germ-line therapy from germ-line cells in the ovaries of a female and the testes of a male is not permitted in any country. However somatic gene therapy from somatic cells is permitted under certain tightly regulated circumstances.
In the late 1990's trials to investigate the possibility of treating cystic fibrosis by means of somatic gene therapy began in the UK and USA. Cystic fibrosis results from an individual being homozygous recessive for the faulty form of the cystic fibrosis gene. It is caused by inborn errors in single genes. The gene therapy for this will hopefully open the possibility of giving those affected by the condition, nasal sprays which would contain the healthy form of the cystic fibrosis gene packaged in a harmless virus, a hollow sphere of lipid molecules or some other vector. The idea is that healthy genes enter the lung epithelial cells, splice with their nucleic DNA and therefore make the CTFR protein, which the faulty cystic fibrosis gene cannot, thus allowing the mucus secreted by these cells to have its normal runny consistency. (www.newscientist.com)
During the early 1990's research began in genetic engineering in tomatoes. The average tomato is picked before it is ripe when it is still green. It is sprayed with a liquid known as ethylene, which makes it turn red. Because the ripening of the tomato isn't natural, their taste and texture isn't right. One company developed a genetically modified tomato known as the "Flav'r Sav'r". The engineering takes the form of preventing the plant from making polygalacturonase.
Polygalacturonase synthesis takes place in tomatoes as they ripen from green to red. It makes the tomatoes soften by breaking down some of the parts in the cell walls between tomato cells. This process is done by getting the cells to make a substance that binds to its mRNA, which then cannot make the protein coded for by that gene. The result is a red firm tomato.
However during 1999 the tomatoes were withdrawn from sale 'on commercial grounds'. They were found to be lower than normal in Vitamin C. Also marker genes that are antibiotic resistant which were used to tell genetic engineers if the crops have worked were contained still in the tomatoes and that there was a possibility that such genes may move to pathogenic bacteria and make them also immune to antibiotics. As such it also seemed likely that the antibiotic immunity could be passed onto humans, which in medicine today is becoming an increasing problem. (www.msn.com/magazines/gmfoods)
Embryonic stem cell research has been classed as the way ahead with human genetic engineering, labelled the 'super-cell' it has the potential to create one of the human body's two hundred or so tissues.
Embryonic stem cells appear a few days after an egg is fertilised and known as a blastocyst. The cell walls of this will later become the placenta, but it is the small mass of cells within, which will later grow to form the embryo. They only spend a relatively short space of time as stem cells and formulate into specialised cells of the body at a rapid rate.
Much of the early research began with tumours, some of which can grow their own muscle tissue, nerves, teeth, tufts of hair and even deformed arms and legs.
A discovery was made by Angelo Vescovi's team at the National Neurological institute, that when testing on neural stem cells in mice, injecting the cells into the bone marrow of mice which had had most of their own white blood cells destroyed, they turned into new white blood cells. This is known as therapeutic cloning. In humans for example this could provide treatments for diseases such as Parkinson's disease by growing healthy cells to replace and then reproduce the dysfunctional cells that cause it.
Presently more cells die or ignore signals to convert into their new cell format, even if a few of the desired cells can be activated, they would have to be grown for many generations to make enough for a transplant. This prematurely ages the tissue.
There are many factors that have affected advances in stem cell engineering such as law. The British government commissioned a report on stem cells research, which was completed in August this year. Liam Donaldson (the Chief Medical Officer) stated at the press launch of the report, that "My own view, and that of the committee, is that stem cell research opens up a new medical frontier. It's got major, major potential". Problems were highlighted in the report regarding premature ageing, potentially hazardous mutations of tissues derived from stem cells. Also the ethical issue of using embryonic stem cells was raised and other ways of sourcing the stem cells were discussed, but using embryonic stem cells was not ruled out. The expert group concluded that the great potential to relieve suffering and treat disease meant that research was warranted across the whole range of possible sources of stem cells in the first instance, including embryo's. In summation of this essay, although the Donaldson Report stated embryonic stem cell engineering was a major benefit to mankind the expert group also put forward the following recommendations, which I think are relevant to highlight as well as monitor the possible problems of genetic engineering; -
Recommendation 1
Research using embryos (whether created by in-vitro fertilisation or cell nuclear replacement) to increase understanding about human disease and disorders and their cell based treatments should be permitted subject to the controls in the Human Fertilisation and Embryology Act 1990 (HFE Act 1990).
Recommendation 2
In licensing any research using embryos created by cell nuclear replacement, the Human Fertilisation and Embryology Authority (HFEA) should satisfy itself that there are no other means of meeting the objectives of the research.
Recommendation 3
Individuals whose eggs or sperm are used to create the embryos to be used in research should give specific consent indicating whether the resulting embryos could be used in a research project to derive stem cells.
Recommendation 4
Research to increase understanding of, and develop treatments for, mitochondrial diseases using the cell nuclear replacement technique in human eggs, which are subsequently fertilised by human sperm, should be permitted subjected to the controls in the HFE Act 1990.
Recommendation 5
The progress of research involving stem cells, which have been derived from embryonic sources, should be monitored by an appropriate body to establish whether the research is delivering the anticipated benefits and to identify any concerns, which may arise.
Recommendation 6
The mixing of human adult somatic cells with the live eggs of any animal species should not be permitted.
Recommendation 7
The transfer of an embryo created by cell nuclear replacement into the uterus of a woman (reproductive cloning) should remain a criminal offence.
Recommendation 8
The need for legislation to permit the use of embryo derived cells in treatments developed from this new research should be kept under review.
Recommendation 9
The Research Councils should be encouraged to establish a programme for stem cell research and to consider the feasibility of establishing collections of stem cells for research use.