Biotechnology and Genetic Modification

Microorganisms as living factories

Biotechnology is the use of living organisms, or the parts of them, to make useful products or carry out useful processes. Most of the examples in this topic use microorganisms: bacteria, yeast and fungi.

Bacteria are especially useful for two reasons. They have a rapid reproduction rate, so under good conditions a population can double every 20 minutes or so; a small starter culture soon becomes an enormous number of cells, all making the product you want. They can also make complex molecules, including proteins such as human insulin, that are difficult or impossible to make by chemistry alone.

Two further reasons are tested at Extended level. There are few ethical concerns over manipulating and growing bacteria, so engineering a bacterium raises far fewer objections than experimenting on animals. And bacteria contain plasmids: small circular loops of DNA separate from the main chromosome, which can be removed, cut open and used to carry a gene from another organism into the bacterium.

• Biotechnology = using living organisms, or their parts, to make useful products or carry out useful processes.
• Bacteria reproduce rapidly, so a large population and a large amount of product build up quickly.
• Bacteria can make complex molecules such as human proteins.
• Bacteria are easy and cheap to grow, and engineering them raises few ethical concerns.
• Plasmids are small circular loops of DNA that can act as vectors, carrying a gene into a bacterium.

Fermentation: yeast, bread, beer and biofuels

Yeast is a single-celled fungus. When oxygen is present it respires aerobically, but in the absence of oxygen it respires anaerobically (ferments), converting glucose into ethanol and carbon dioxide. The word equation is: glucose -> ethanol + carbon dioxide.

In bread-making the useful product is the carbon dioxide. Yeast is mixed with flour, water and sugar to make dough. As the yeast ferments the sugar, bubbles of carbon dioxide are trapped in the dough, so it rises and becomes light and spongy. The small amount of ethanol produced is baked off in the oven, so it does not affect the taste.

In brewing the useful product is the ethanol. Yeast ferments the sugars from malted barley or grapes in a vessel kept closed so that oxygen cannot get in, and the carbon dioxide produced gives the drink its fizz. In the production of biofuels the same reaction is used deliberately to make ethanol from sugar cane or maize, which is distilled and burned as a fuel for vehicles.

Enzymes in industry: washing powders, fruit juice and lactose-free milk

Enzymes are used in industry because they work at low temperatures and pressures, which saves energy, and because each enzyme is specific, so it acts on one substrate without producing unwanted side products.

Pectinase in fruit juice production. Pectin is a substance in plant cell walls that makes fruit pulp thick and cloudy. Pectinase breaks pectin down, so the pulp releases more juice and the juice is clearer. It also makes the juice easier to filter and increases the yield.

Biological washing powders. These contain proteases and lipases that digest protein and fat stains such as blood, egg and grease. Because the enzymes work best at around 40 °C, the wash can be done at a lower temperature, which saves energy. Washing at too high a temperature would denature the enzymes and they would stop working.

Lactase and lactose-free milk. Lactase is an enzyme that breaks down lactose, the sugar in milk, into glucose and galactose. Some people cannot digest lactose because they do not make enough lactase of their own, so drinking milk gives them diarrhoea and wind. Treating milk with lactase removes the lactose, so these people can drink it and still get the calcium and protein they need.

• Enzymes work at low temperatures, so using them saves energy compared with high-temperature chemical processes.
• Enzymes are specific, so they produce the product wanted with few unwanted side products.
• Pectinase breaks down pectin, increasing the yield of clear juice from fruit pulp.
• Proteases and lipases in biological washing powders digest protein and fat stains.
• Lactase breaks lactose into glucose and galactose, making milk that lactose-intolerant people can drink.

The fermenter: growing microorganisms on an industrial scale

A fermenter is a large vessel used to grow microorganisms on an industrial scale so that the product they make can be harvested in bulk. Bacteria and fungi are grown in fermenters to make insulin, penicillin and mycoprotein.

The vessel is a large tank, usually stainless steel, that holds the culture. The stirrer or paddles keep the microorganisms, nutrients and oxygen evenly mixed and stop them settling, so every cell has access to what it needs. The water jacket around the vessel carries water at a controlled temperature, removing the heat released by respiration so the culture does not overheat. The air supply pumps sterile air in for aerobic organisms, and the outlets let waste gases out, allow samples to be taken, and let the product be harvested at the end.

The conditions must be controlled carefully. The temperature is kept constant, because too low slows growth and too high denatures the enzymes and kills the microorganisms. The pH is held near the optimum for the organism. A supply of nutrients such as glucose and mineral ions is provided for growth, and for aerobic organisms there is a continuous supply of oxygen. Waste products are removed, because they would otherwise build up and become toxic and slow the growth of the culture.

Finally, the culture must be kept sterile. The vessel, the nutrients and the air are all sterilised and the fermenter is sealed, so unwanted microorganisms cannot grow. A contaminant would compete for nutrients, reduce the yield and could spoil the product.

Genetic modification: the five steps

Genetic modification (genetic engineering) is changing the genetic material of an organism by removing, changing or inserting individual genes. Because the genetic code is the same in almost every organism, a gene taken from a human can be read and used by a bacterium. That is what makes genetic modification possible.

The gene is carried into the host cell by a vector, most often a plasmid taken from a bacterium. Learn the five steps in order, with the enzymes named, because examiners mark the sequence as well as the detail.

1. Cut out the gene. A restriction enzyme recognises a specific base sequence and cuts the DNA there, isolating the gene. The cut leaves short single-stranded lengths of DNA called sticky ends. 2. Cut open the vector. The same restriction enzyme cuts open the bacterial plasmid DNA, so it has complementary sticky ends that can pair with the sticky ends of the gene. 3. Join gene to vector. DNA ligase joins the gene to the plasmid DNA, forming a recombinant plasmid. 4. Transfer into the host. The recombinant plasmids are inserted into bacteria, and the bacteria are grown and multiplied, so the plasmid is copied every time a bacterium divides. 5. Harvest the product. The gene is expressed by the bacteria: they read it and make the human protein, which is then extracted and purified.

Human insulin from genetically modified bacteria

Diabetes is a condition in which the body cannot control its blood glucose concentration. Many people with diabetes need regular insulin injections. Insulin used to be extracted from the pancreases of pigs and cattle, but it is now made by genetically modified bacteria.

The human insulin gene is cut out with a restriction enzyme and joined to a plasmid with DNA ligase, making a recombinant plasmid. The plasmid is inserted into a bacterium, which multiplies rapidly in a fermenter, and every cell reads the human gene and makes human insulin. The insulin is then harvested and purified.

The advantages are clear. The supply is unlimited and does not depend on the number of animals slaughtered, so it can be produced on a large scale. The product is identical to human insulin, so it works more reliably and causes fewer allergic reactions than animal insulin. And because bacteria multiply rapidly in a fermenter, the insulin can be made quickly and at low cost.

GM crops: benefits, concerns and selective breeding

Genes can be inserted into crop plants to give them a useful characteristic. Cambridge asks for resistance to herbicides, resistance to insect pests, and improved nutritional qualities. A crop given resistance to a herbicide can be sprayed so that weeds are killed but the crop survives; a crop given resistance to pests is damaged less and needs fewer insecticides; a crop given improved nutritional qualities is more nutritious. The named examples are soya, maize and rice.

Golden rice is the standard example of improved nutritional content: rice has been given genes that let it make beta-carotene, which the body converts into vitamin A. In countries where rice is the main food and vitamin A deficiency causes blindness, this could improve health. Crops have also been modified to have a longer shelf life, so less food is wasted in transport and in shops.

Concerns. Modified crops may cross-breed with wild plants and transfer their genes to wild populations, which could produce weeds resistant to herbicides. Growing a single modified variety over huge areas reduces biodiversity, because the whole crop is genetically similar. There are also ethical concerns (should humans change the genetic material of other organisms at all?) and economic concerns (farmers may have to buy new seed and herbicide every year).

Genetic modification is not the same as selective breeding. Selective breeding chooses organisms that already show a useful characteristic and breeds them together; the genes come from the same species, and it takes many generations. Genetic modification takes a gene from one organism and inserts it into another, so it can move a gene between different species and produce a new variety in a single generation.