What is DNA Science/Genetic Engineering?
We find it mixed in our food on the shelves in the supermarket--genetically engineered soybeans and maize. We find it growing in a plot down the lane, test field release sites with genetically engineered rape seed, sugar beet, wheat, potato, strawberries and more. There has been no warning and no consultation.
It is variously known as genetic engineering, genetic modification or genetic manipulation. All three terms mean the same thing, the reshuffling of genes usually from one species to another; existing examples include: from fish to tomato or from human to pig. Genetic engineering (GE) comes under the broad heading of biotechnology.
But how does it work? If you want to understand genetic engineering it is best to start with some basic biology.
What is a cell? A cell is the smallest living unit, the basic structural and functional unit of all living matter, whether that is a plant, an animal or a fungus.Some organisms such as amoebae, bacteria, some algae and fungi are single-celled - the entire organism is contained in just one cell. Humans are quite different and are made up of approximately 3 million cells -(3,000,000,000,000 cells). Cells can take many shapes depending on their function, but commonly they will look like a brick with rounded comers or an angular blob - a building block.Cells are stacked together to make up tissues, organs or structures (brain, liver, bones, skin, leaves, fruit etc.).
In an organism, cells depend on each other to perform various functions and tasks; some cells will produce enzymes, others will store sugars or fat; different cells again will build the skeleton or be in charge of communication like nerve cells; others are there for defence, such as white blood cells or stinging cells in jelly fish and plants. In order to be a fully functional part of the whole, most cells have got the same information and resources and the same basic equipment.
A cell belonging to higher organisms (e.g. plant or animal) is composed of: · a cell MEMBRANE enclosing the whole cell. (Plant cells have an additional cell wall for structural reinforcement.) · many ORGANELLES, which are functional components equivalent to the organs in the body of an animal e.g. for digestion, storage, excretion. · a NUCLEUS, the command centre of the cell. It contains all the vital information needed by the cell or the whole organism to function, grow and reproduce. This information is stored in the form of a genetic code on the chromosomes, which are situated inside the nucleus.
Proteins are the basic building materials of a cell, made by the cell itself. Looking at them in close-up they consist of a chain of amino-acids, small specific building blocks that easily link up. Though the basic structure of proteins is linear, they are usually folded and folded again into complex structures. Different proteins have different functions. They can be transport molecules (e.g. oxygen binding haemoglobin of the red blood cells); they can be antibodies, messengers, enzymes (e.g. digestion enzymes) or hormones (e.g. growth hormones or insulin). Another group is the structural proteins that form boundaries and provide movement, elasticity and the ability to contract. Muscle fibres, for example, are mainly made of proteins. Proteins are thus crucial in the formation of cells and in giving cells the capacity to function properly.
Chromosomes means "coloured bodies" (they can be seen under the light microscope, using a particular stain). They look like bundled up knots and loops of a long thin thread. Chromosomes are the storage place for all genetic - that is hereditary - information. This information is written along the thin thread, called DNA. "DNA" is an abbreviation for deoxyribo nucleic acid, a specific acidic material that can be found in the nucleus. The genetic information is written in the form of a code, almost like a music tape. To ensure the thread and the information are stable and safe, a twisted double stranded thread is used - the famous double helix. When a cell multiplies it will also copy all the DNA and pass it on to the daughter cell.
The totality of the genetic information of an organism is called genome. Cells of humans, for example, possess two sets of 23 different chromosomes, one set from the mother and the other from -the father. The DNA of each human cell corresponds to 2 meters of DNA if it is stretched out and it is thus crucial to organise the DNA in chromosomes, so as to avoid knots, tangles and breakages. The length of DNA contained in the human body is approximately 60,000,000,000 kilometres. This is equivalent to the distance to the moon and back 8000 times!
The information contained on the chromo-somes in the DNA is written and coded in such a way that it can be understood by almost all living species on earth. It is thus termed the universal code of life. In this coding system, cells need only four symbols (called nucleotides) to spell out all the instructions of how to make any protein. Nucleotides are the units DNA is composed of and their individual names are commonly abbreviated to the letters A, C G and T These letters are arranged in 3-letter words which in turn code for a particular amino acid - as shown in the flow diagram 1. The information for how any cell is structured or how it functions is all encoded in single and distinct genes. A Gene is a certain segment (length) of DNA with specific instructions for the production of commonly one specific protein. The coding sequence of a gene is, on average about 1000 letters long. Genes code for example for insulin, digestive enzymes, blood clotting proteins, or pigments.
How does a cell know when to produce which protein and how much of it? In front of each gene there is a stretch of DNA that contains the regulatory elements for that specific gene, most of which is known as the promoter. It functions like a "control tower," constantly holding a "flag" up for the gene it controls. Take insulin production (which we produce to enable the burning of the blood sugar). When a message arrives in the form of a molecule that says, ’more insulin", the insulin control tower will signal the location of the insulin gene and say "over here". The message molecule will "dock in" and thus activate a "switch" to start the whole process of gene expression.
How does the information contained in the DNA get turned into a protein at the right time? As shown in picture 2, each gene consists of 3 main components: a "control tower" (promoter), an information block and a polyA signal element. If there is not enough of a specific protein present in the cell, a message will be sent into the nucleus to find the relevant gene. If the control tower recognises the message as valid it will open the "gate" to the information block. Immediately the information is copied - or transcribed - into a threadlike molecule, called RNA. RNA is very similar to DNA, except it is single stranded. After the copy is made, a string of up to 200 "A"-type nucleotides - a polyA tail - is added to its end (picture 2). This process is called poly-adenylation and is initiated by a polyA signal located towards the end of the gene. A polyA tail is thought to stabilise the RNA message against degradation for a limited time. Now the RNA copies of the gene leave the nucleus and get distributed within the cell to little work units that translate the information into proteins.
No cell will ever make use of all the information coded in its DNA. Cells divide the work up amongst one other - they specialise. Brain cells will not produce insulin, liver cells will not produce saliva, nor will skin cells start producing bone. If they did, our bodies could be chaos!
The same is true for plants: root cells will not produce the green chlorophyll, nor will the leaves produce pollen or nectar. Furthermore, expression is age dependent: young shoots will not express any genes to do with fruit ripening, while old people will not usually start developing another set of teeth (exceptions have been known).
All in all, gene regulation is very specific to the environment in which the cell finds itself and is also linked to the developmental stages of an organism. So f I want the leaves of poppy plants to produce the red colour of the flower petals I will not be able to do so by traditional breeding methods, despite the fact that leaf ells will have all the genetic information necessary. There is a block that prevents he leaves from going red. This block may be caused by two things: · The "red" gene has been permanently shut down and bundled up thoroughly in all leaf cells. Thus the information cannot be accessed any more. · The leaf cells do not need the colour red and thus do not request RNA copies of this information. Therefore no message molecule is docking at the "red" control tower to activate the gene.
Of course - you might have guessed - there is a trick to fool the plant and make it turn red against its own will. We can bring the red gene in like a Trojan horse, hidden behind the control tower of a different gene. But for this we need to cut the genes up and glue them together in a different form. This is where breeding ends and genetic engineering begins.
BREEDING is the natural process of sexual reproduction within the same species. The hereditary information of both parents is combined and passed on to the offspring. In this process the same sections of DNA can be exchanged between the same chromosomes, but genes will always remain at their very own and precise position and order on the chromosomes. A gene will thus always be surrounded by the same DNA unless mutations or accidents occur. Species that are closely related might be able to interbreed, like a donkey and a horse, but their offspring will usually be infertile (e.g. mule). This is a natural safety devise, preventing the mixing of genes that might not be compatible and to secure the survival of the species.
Genetic engineering (GE) is used to take genes and segments of DNA from one species, e.g. fish, and put them into another species, e.g. tomato. To do so, GE provides a set of techniques to cut DNA either randomly or at a number of specific sites. Once isolated one can study the different segments of DNA, multiply them up and splice them (stick them) next to any other DNA of another cell or organism. GE makes it possible to break through the species barrier and to shuffle information between completely unrelated species; for example, to splice the anti-freeze gene from flounder into tomatoes or strawberries, an insect-killing toxin gene from bacteria into maize, cotton or rape seed, or genes from humans into pig.
Yet there is a problem - a fish gene will not work in tomato unless I give it a promoter with a "flag" the tomato cells will recognise. Such a control sequence should either be a tomato sequence or something similar. Most companies and scientists do a shortcut here and don’t even bother to look for an appropriate tomato promoter as it would take years to understand how the cell’s internal communication and regulation works. In order to avoid long testing and adjusting, most genetic engineering of plants is done with viral promoters. Viruses - as you will be aware - are very active. Nothing, or almost nothing, will stop them once they have found a new victim or rather host. They integrate their genetic information into the DNA of a host cell (such as one of your own), multiply, infect the next cells and multiply. This is possible because viruses have evolved very powerful promoters which command the host cell to constantly read the viral genes and produce viral proteins. Simply by taking a control element (promoter) from a plant virus and sticking it in front of the information block of the fish gene, you can get this combined virus/fish gene (known as a "construct’) to work wherever and whenever you want in a plant.
This might sound great, the drawback though is that it can’t be stopped either, it can’t be switched off. The plant no longer has a say in the expression of the new gene, even when the constant involuntary production of the "new" product is weakening the plant’s defences or growth.
And furthermore, the theory doesn’t hold up with reality. Often, for no apparent reason, the new gene only works for a limited amount of time and then "falls silent". But there is no way to know in advance if this will happen.
Though often hailed as a precise method, the final stage of placing the new gene into a receiving higher organism is rather crude, seriously lacking both precision and predictability. The "new" gene can end up anywhere, next to any
gene or even within another gene, disturbing its function or regulation. If the "new" gene gets into the "quiet" non-expressed areas of the cell’s DNA, it is likely to interfere with the regulation of gene expression of the whole region. It could potentially cause genes in the "quiet" DNA to become active.
Often genetic engineering will not only use the information of one gene and put it behind the promoter of another gene, but will also take bits and pieces from other genes and other species. Although this is aimed to benefit the expression and function of the "new" gene it also causes more interference and enhances the risks of unpredictable effects.
How to get the gene into the other cell.?
There are different ways to get a gene from A to B or to transform a plant with a "new" gene. A VECTOR is something that can carry the gene into the host, or rather into the nucleus of a host cell. Vectors are commonly bacterial plasmids (see below and next page) or viruses (a). Another method is the "SHOTGUN TECHNIQUE" also known as "bio-ballistics," which blindly shoots masses of tiny gold particles coated with the gene into a plate of plant cells, hoping to land a hit somewhere in the cell’s DNA (b).
What is a plasmid?
PLASMIDS can be found in many bacteria and are small rings of DNA with a limited number of genes. Plasmids are not essential for the survival of bacteria but can make life a lot easier for them. Whilst all bacteria - no matter which species - will have their bacterial chromosome with all the crucial hereditary information of how to survive and multiply, they invented a tool to exchange information rapidly. If one likens the chromosome to a bookshelf with manuals and handbooks, and a single gene to a recipe or a specific building instruction, a plasmid,could be seen as a pamphlet. Plasmids self-replicate and are thus easily reproduced and passed around. Plasmids often contain genes for antibiotic resistance. This type of information which can easily be passed on, can be crucial to bacterial strains which are under attack by drugs and is indeed a major reason for the quick spread of antibiotic resistance.
Working with plasmids?
Plasmids are relatively small, replicate very quickly and are thus easy to study and to manipulate. It is easy to determine the sequence of its DNA, that is, to find out the sequence of the letters (A, C, G and 1) and number them. Certain letter combinations -such as CAATTG - are easy to cut with the help of specific enzymes (see proteins). Ilese cutting enzymes, called restriction enzymes, are part of the Genetic Engineering "tool-kit" of biochemists. So if I want to splice a gene from fish into a plasmid, I have to take the following steps: I place a large number of a known plasmid in a little test tube and add a specific enzyme that will cut the plasmid at only one site. After an hour I stop the digest, purify the cut plasmid DNA and mix it with copies of the fish gene; after some time the fish gene places itself into the cut ring of the plasmid. I quickly add some "glue" from my "tool-kit" - an enzyme called ligase - and place the mended plasmids back into bacteria, leaving them to grow and multiply. But how do I know which bacteria will have my precious plasmid? For this reason I need MARKER GENES, such as antibiotic resistance genes. So I make sure my plasmid has a marker gene before I splice my fish gene into it. If thA I plasmid is marked with a gene antibiotic resistance I can now add specific antibiotic to the food supply of the bacteria. All those which do not have the plasmid will die, and all those that do have the plasmid will multiply.
What’s wrong with Genetic Engineering ?
Genetic Engineering is a test tube science and is prematurely applied in food production. A gene studied in a test tube can only tell what this gene does and how it behaves in that particular test tube. It cannot tell us what its role and behaviour are in the organism it came from or what it might do if we place it into a completely different species. Genes for the colour red placed into petunia flowers not only changed the colour of the petals but also decreased fertility and altered the growth of the roots and leaves. Salmon genetically engineered with a growth hormone gene not only grew too big too fast but also turned green. These are unpredictable side effects, scientifically termed pleiotropic effects.
We also know very little about what a gene (or for that matter any of its DNA sequence) might trigger or interrupt depending on where it got inserted into the new host (plant or animal). These are open questions around positional effects. And what about gene silencing and gene instability? How do we know that a genetically engineered food plant will not produce new toxins and allergenic substances or increase the level of dormant toxins and allergens? How about the nutritional value? And what are the effects on the environment and on wild life? All these questions are important questions yet they remain unanswered. Until we have an answer to all of these, genetic engineering should be kept to the test tubes. Biotechnology married to corporations tends to ignore the precautionary principle but it also igpores some basic scientific principles.
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