Site author Richard Steane
The BioTopics website gives access to interactive resource material, developed to support the learning and teaching of Biology at a variety of levels.
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A gene is a unit of hereditary information (i.e. it normally passes
on characteristics from one generation to another), and is composed of DNA.
Gene manipulation may be advantageous because it makes the resulting genetically modified ortransgenic
organism easier to grow or manage, or to transfer a characteristic to a
different crop, etc.
It differs from selective breeding which only involves
members of the same species, in that usually only single genes are
moved, often in addition to that organism's normal complement of genes
("genome").
Because selective breeding involves the normal methods of sexual
reproduction (gamete transfer, fertilisation and development, etc.), it
only results in large combinations of genes being transferred (the haploid number
of chromosomes contained in a gamete is in effect half a genome), and
the effect of these genes may be masked or diluted due to dominance by
other genes.
Restriction enzymes break DNA at specific parts of the
molecule (nucleotide base sequences) - usually leaving so called
"sticky ends". This can be done to both DNA from which genes are being
taken, and to DNA in which genes are being inserted.
DNA ligase enzymes may be used to rejoin such sections into
the other DNA.
The DNA containing the selected gene for the desired characteristic
may then be inserted into cells of the target organism by means of vectors
(here used in the service of Man, not disease organisms).
There are 2 main types of vectors:
plasmids and viruses (see previous notes on
micro-organisms).
Agrobacterium tumefaciens
is a bacterium which in nature causes plant disease - "crown gall
disease" - but only in some dicots ("broad-leaved plants"), not in
monocots (grasses and cereals). A gall is a mass of undifferentiated plant tissue - similar
to a cancerous tumour - produced in response to such an infection.
Crown galls are usually produced on the stem just above the surface of
the ground.
The bacterium contains a section of DNA called a plasmid in
addition to its usual component of DNA. This Ti (tumour inducing)
plasmid normally incorporates its DNA into the cells of the plant host
("integrating with their genome").
The ability of this organism can be utilised in genetic
engineering to insert other genes into crop plants.
However, it is said that, due to commercial pressures, the main use
of gene transfer to date has been to confer resistance to pests or
diseases, rather than more direct impact on yield or other desirable
characteristics. A gene thought to be useful may be obtained from a
variety of sources, e.g.:-
The gene for resistance to herbicide (weedkiller) may
be obtained from (occasional) weeds which survive treatment with this
chemical. This could perhaps usefully be incorporated into a crop which
would then benefit from reduced competition from weeds, less hoeing
etc, when sprayed with the appropriate herbicide. Commercially it would
also mean that the seed and herbicide would be part of the same supply
deal.
This procedure has actually been applied to crops of commercial
significance, e.g. soya (beans),
sugar beet, tobacco, and oilseed rape.
The Bt gene for production of insecticidal toxin fromBacillus
thuringensis has been incorporated into several crops in order to
protect them against insect pests.
Protection of crops from insect damage has also been tested using
the gene for venom from scorpions!
Similarly, the effects of incorporating pest resistance genes from
snowdrops into potatoes has been investigated.
Other novel ideas include the transfer of genes coding for important
animal proteins such as the hormone insulin into plants, such as
potatoes, which are easily grown and processed, and the transfer of
genes into easily managed animals such as cows, sheep, etc, which may
produce milk containing valuable proteins such as human antibodies and
anti-cancer products.
Other alternative approaches
involve isolation and modification of genes so that normal
developmental changes do not occur. For example, there are several
enzyme-controlled stages in the ripening and subsequent
deterioration (spoilage) of fruit. Modification
(inhibition) of the genes producing these enzymes can slow down the
changes which occur after fruit is ripe. As a result, the keeping
quality or shelf-life of the fruit is increased, and possibly the
quality of products derived from these fruits is improved, as well as
reducing the processing costs.
This has been achieved and licensed in the case of tomatoes and
products such as puree.
It is also said that attempts are being made to produce strains of
soya beans which will flourish in temperate climates, and which are
tall enough to facilitate mechanical harvesting.
Interestingly, several biotechnology companies working in the field have attracted the attention of investors excited by the prospect of profits to be made. However, much venture capital has been used in the process, and there is considerable commercial rivalry and secrecy as to the exact details of the processes. Similarly, there is much public distrust as to the true intentions of workers in the field, and campaigners on each side have raised the profile of these activities in relation to regulatory authorities.
Recently there have been a variety of developments:
- Organisations representing consumer interests wish to ban GMOs (genetically modified
organisms) from entering the food supply chain, or to have
it kept separate from other food supplies, and have its origin
specifically stated in the product labelling
- Supermarket chains
have in some cases responded either by sourcing supplies of non
genetically modified foods, or by identifying such ingredients in the
labelling of the food, even if only a minor constituent. This is an
ongoing development! Iceland was one of the first to do this, and on
the 28th April 1999 Tesco also announced it was stopping using GMOs.
- Growers or importers have
mixed genetically modified foods with non genetically modified foods,
either on the grounds that to do otherwise would increase costs, or in
order to confuse the issue, in the hope of speedy acceptance of the
product. This has been the case with soya beans, which are a
major export from the USA.
- Test plots of varieties of plants being assessed for future
use are covered by a variety of regulations designed to reduce the
likelihood of any transfer of genes to surrounding crops.
In some cases corners have been cut and tempers have run high. Some
pressure groups have advocated a moratorium on these trials, i.e.
postponing them for several years.
- Farmers and growers must sign undertakings not to save seed from
the crop for use to start another crop next year, because agrochemical
companies have patent and
other rights to the varieties used, and expect an exclusive agreement
to use a combination of seed and control chemicals from the same
supplier.
- The impartiality of some of the more important committees overseeing
trials carried out by large companies has been called into question.
Many of the decisions used to be made by employees of companies with
interests in genetic modification, and a company owned by Lord
Sainsbury, a government minister, holds patent rights to, and therefore
profits from, important techniques in genetic manipulation.
DNA from the donor organism is broken up into short lengths with
"sticky ends" using a restriction enzyme.
One or some of these fragments should include the gene for the
desirable characteristic, but often there is an element of chance, so
the procedure is frequently repeated many times.
The tumour inducing plasmid which
consists of DNA from Agrobacterium tumefaciens is similarly
treated with the same restriction enzyme, opening out the
circle of DNA leaving 2 sticky ends.
The presumed gene DNA is then mixed with the plasmid DNA, and
conditions provided for the DNA ligase enzyme
to work. In a number of cases, this will result in the plasmid
re-joining, but with the gene incorporated into it.
The plasmid is reintroduced into the bacterium, which can then be
grown up in large numbers by standard microbiological methods.
When plants are infected with these bacteria, they will form galls of
undifferentiated tissue, some cells of which will contain the required
gene.
Sections of the gall may be encouraged to grow by special plant
tissue culture techniques, possibly bulked up in the lab
before conditions in the medium are changed to encourage growth of
roots and shoots.
The resulting small plants may eventually be potted up and
finally transferred to the field!
This also involves the activities of a species of bacterium (Rhizobium
leguminosarum) which enters a plant organ (root of a legume)
resulting in a change in the plant cell growth to form a root
nodule, in which bacteria grow and perform chemical transformations.
It is hoped that genes for nitrogen fixation (nif cluster)
may be transferred to non-leguminous plants. However there is more
genetic information in these (12/20-30genes than can be easily
transferred using plasmids, so more ambitious methods are being tried.
Gene expression (turning them on) is a problem, especially as bacteria
(prokaryotes - lacking nuclei/chromosomes) differ greatly from higher
plants (eukaryotes - chromosomes protect DNA inside nuclei).
Certain viruses can infect cells
of animals and plants "without completing a destructive cycle"
so they may also be used as gene vectors. They can usually carry larger
portions of DNA than plasmids can . An example is 
(lambda) phage - a bacteriophage which
can modify bacteria. DNA from a so-called temperate phage
becomes incorporated into the DNA of its host: the bacterium Escherichia
coli (E. coli), and can remain there indefinitely without having any
harmful effect.
The phage DNA can be opened using restriction enzymes and foreign
DNA may be inserted, so that the viral DNA can integrate with the host
cells's "chromosome" (it is then called a prophage), and
replicates with it at cell division.
Similarly, plant viruses may be used to transform plant cells
genetically.
Ballistic techniques
Minute tungsten particles are coated with the DNA to be inserted,
then shot into the target cells with an explosive charge. Apparently,
this does not, however, cause significant structural damage to the
cells.
Electroporation
In this technique, a brief pulse of electric current is passed
through the cell, temporarily increasing surface permeability so
that DNA is taken up from the surrounding liquid. This has been
especially useful with pollen tubes and has resulted in the genetic
transformation of seeds. Certain chemicals may have the same effect on
the permeability of the cell wall.
The same techniques used in the production of insulin and antibiotics may be applied to the use of genetically engineered bacteria in food production. Examples include yeasts with high alcohol tolerance, microbes with enhanced ability to digest waste straw, peat, coal, oil, etc., and improvements in capacity to produce valuable substances e.g. enzymes, flavourings, colourings. To some extent, industry has favoured the application of genetic modification processes to organisms which have achieved public acceptance, such as yeasts and lactic acid bacteria (Lactobacilli), which are responsible for cheese production as well as yoghurt and soy sauce.
However, potential human medical applications have been seen
to offer great opportunities. Production of blood clotting factor
(needed by sufferers of the genetic condition haemophilia)
can be induced in the milk of sheep. So-called "designer milk"
containig low cholesterol could probably find a profitable market.
More controversially, it has been said that transgenic organisms
such as pigs could be used as sources of organs for transplants
into humans, if human genes were transferred into these organisms at the
embryo stage. This could reduce problems of rejection due to the immune
system of the donor. However, the risk of transfer of potentially very
serious virus diseases from one species to another has become more
obvious in the light of scrapie/BSE/CJD which is said to have "jumped
the species barrier".
These possibilities pose many ethical dilemmas.
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