Applied Genetics: WJEC A Level Biology Notes

Lesson 1 of 21

What were the Human Genome Project and the 100K Genome Project?

These notes cover topic 4.5, Application of reproduction and genetics, in WJEC A level Biology Unit 4: , Inheritance and Options. Each lesson follows the WJEC specification, and the exam tips come from BioWelsh analysis of WJEC past papers.

A is all the DNA in one set of an organism's chromosomes. The human is about 3 billion base pairs long. The () was an international project, begun in 1990 and completed in 2003, that worked out the order of those bases.

Identify all the in the human and find which chromosome each one is on.
Determine the sequence of the 3 billion base pairs and store it in public databases.
Improve the computer tools used to analyse the data.
Improve the understanding, diagnosis and treatment of genetic disorders.
Address the ethical, legal and social issues the knowledge raises.

The project found that humans have only about 20 000 protein-coding , far fewer than expected. Less than 2% of the codes for proteins, and much of the rest is repeated sequences. Many human are shared with other organisms, which shows how closely related all living things are.

The used (the chain termination method). The diagram below shows the idea.

Two-part diagram. On the left, eight rows each start with a grey box labelled primer followed by one to eight coloured square bases, and each row ends with a ringed base: A, C, G, T, T, A, G, C. Text beside each row reads 1 base added up to 8 bases added. On the right, a tall lane contains eight coloured bands, labelled longest at the top and shortest at the bottom, with letters beside them reading upwards A, C, G, T, T, A, G, C. An upward arrow is labelled read upwards from the shortest. A box states the sequence of the new strand as 5 prime A C G T T A G C 3 prime, and text below gives the template strand as 3 prime T G C A A T C G 5 prime. — Sanger sequencing: fragments ending in a coloured base are sorted by length. Original BioWelsh diagram.

Many copies of a short DNA fragment are copied, starting from a .
The mixture includes a few special , each carrying a coloured fluorescent label. When one is added, copying stops.
This gives fragments of every possible length, each ending in a coloured base.
sorts the fragments by length. Reading the colours from the shortest fragment to the longest gives the base sequence.

Sanger sequencing compared with next generation sequencing (NGS)
Feature	Sanger sequencing	Next generation sequencing
Fragments read at once	One at a time	Millions in parallel
Scale	Separate tubes and gels	Tiny volumes on a chip
Time for a human	Years	About a day
Cost	Very high	Much lower

The () followed in 2012. It used to sequence the whole genomes of NHS patients with rare diseases or cancer, and their families. Linking each to the patient's medical records helps doctors find the causes of disease, make faster diagnoses and choose better treatments. Its results now feed into a genomic medicine service in the NHS.

Lesson 2 of 21

What ethical issues does knowing a person's genome raise?

Knowing someone's DNA sequence is powerful. It can predict the risk of some diseases, show family relationships and reveal ancestry. That creates questions about who owns the information and who may use it.

Ethical issues raised by genome sequencing
Issue	The concern
Ownership and consent	Who owns a person's sequence? Companies should not profit from it without permission.
Insurance and employment	An insurer or employer could refuse cover or a job to someone with a higher genetic risk of disease.
Discrimination	Information about ancestry or disease risk could be used to treat people unfairly.
Relatives	A result also says something about parents, brothers, sisters and children, who may not want to know.
The right not to know	Some people would rather not learn that they are at risk of an untreatable condition, such as .
Testing children	Should parents learn a child's risk of an adult-onset disease before the child can decide?
Data security	Stored genomes could be hacked or used for a purpose other than the one agreed.

helps with these decisions. A genetic counsellor explains the chance of a person, or their children, inheriting a condition, using the family history and any test results, so that people can make informed choices.

Lesson 3 of 21

How are embryos screened for genetic disorders, and is it right?

testing people for to disease. It can be done on adults, on a foetus in the uterus, or on embryos made by ().

finds people who carry one , such as the , without having the disease.
() tests one cell from an embryo, so only embryos without the disorder are placed in the uterus.
Prenatal testing checks a foetus: at about 10 weeks, at about 15 to 20 weeks, or non-invasive testing of foetal DNA in the mother's blood.
Pre-symptomatic testing tells an adult whether they carry an for a later-onset disease such as .

The three disorders named in the specification
Disorder	How it is inherited	What goes wrong
		The CFTR chloride channel does not work, so mucus in the lungs and pancreatic duct is thick and sticky
		cells in the brain , usually from middle age; one is enough
		Too little of one haemoglobin chain is made, causing severe anaemia

Non-invasive prenatal testing uses fragments of foetal DNA found in the mother's blood. They are copied by and the amount of DNA from, for example, chromosome 21 is compared with a chromosome. A ratio of about 3:2 instead of 1:1 suggests three copies of chromosome 21 ().

Arguments for and against screening embryos
For	Against
Prevents a child being born with a severe, untreatable disorder	Embryos with the disorder are destroyed, and some believe an embryo has a right to life
Parents can make an informed choice	It could lead to selecting embryos for non-medical traits
Reduces suffering and long-term care costs	False positive or false negative results are possible
Prenatal diagnosis can prepare parents and doctors	Invasive prenatal tests carry a small risk of miscarriage, and abortion may become more common

Lesson 4 of 21

Why sequence the genomes of other organisms, such as the malaria mosquito?

Genomes have now been sequenced for thousands of . Organisms are chosen because they matter for medicine, farming or research. Comparing genomes also shows how closely are related, so classification can be based on DNA rather than appearance.

Some landmark genome sequences
Organism	Why it matters	Year completed
Haemophilus influenzae (a bacterium)	First free-living organism sequenced	1995
Anopheles gambiae (mosquito)	that carries	2002
Plasmodium falciparum	Parasite that causes the most dangerous	2002
Human		2003
Chimpanzee	Closest living relative of humans	2005

kills hundreds of thousands of people a year, mostly young children in Africa. It is caused by the protoctist Plasmodium and passed on when a female Anopheles mosquito bites. Both are becoming to the chemicals used against them: mosquitoes to insecticides, and Plasmodium to drugs such as chloroquine.

The two photographs below show the and the parasite.

Close-up photograph of a slender brown mosquito with long thin legs and narrow speckled wings, standing on pale skin against a bright blue background. Its long mouthparts point down towards the skin. — An *Anopheles gambiae* mosquito, the vector of malaria, on human skin. Photo: James Gathany, CDC, Public domain.

Many pale beige discs, each a red blood cell, fill the frame on a white background. Inside many of the discs are small purple rings and dark purple dots, and some discs contain two or three of them. — Light micrograph of a blood smear containing ring stages of *Plasmodium falciparum*. Photo: CDC, Dr Greene, Public domain.

The first photograph shows an Anopheles gambiae mosquito resting on human skin before feeding.
The second is a stained blood smear. Each pale disc is a red blood cell, and the small dark dots and rings inside many of them are young Plasmodium falciparum parasites.

Find that make mosquitoes to insecticides, so new insecticides can be designed or old ones used again.
Develop mosquitoes that are less or cannot carry Plasmodium.
Find proteins that Plasmodium needs, as targets for new drugs or .
Track which strains of Plasmodium are drug and where they are spreading.

Lesson 5 of 21

How does the polymerase chain reaction copy DNA?

The () makes millions of copies of a chosen section of DNA in a few hours. It is DNA replication in a tube. This matters because samples from a crime scene, a or a foetus often contain only a tiny amount of DNA.

The DNA sample to be copied (the template).
Two different : short single strands of DNA, about 6 to 25 bases long, complementary to the start of the section to be copied on each strand.
Free DNA nucleotides containing the four bases A, T, C and G.
, a DNA polymerase from the bacterium Thermus aquaticus, which lives in hot springs.
A solution to keep the pH constant.

The tube goes into a (), which changes the temperature quickly. The diagram below shows one cycle.

Three panels joined by arrows. Panel 1, denaturation at 95 degrees Celsius, shows a blue strand labelled 5 prime to 3 prime above a pink strand labelled 3 prime to 5 prime, with arrows pointing apart. Panel 2, annealing at about 55 degrees Celsius, shows the same strands with a short green primer bound under the right end of the blue strand and another above the left end of the pink strand. Panel 3, extension at about 70 degrees Celsius, shows each primer extended by a new paler strand with a yellow oval labelled Taq polymerase at its growing end, and text saying new strands grow 5 prime to 3 prime. Below, five boxes joined by arrows read 1 copy, 2 copies, 4 copies, 8 copies and 16 copies. — One cycle of the polymerase chain reaction. Original BioWelsh diagram.

at 95 °C: the hydrogen bonds between complementary bases break and the DNA separates into two single strands.
at about 55 °C: the bind to the complementary base sequences at the 3′ end of each single strand.
Extension at about 70 °C: attaches to each and adds free nucleotides by complementary base pairing, joining them with phosphodiester bonds.
Two double-stranded copies now exist, and the cycle starts again.

Two are needed because the two strands have different base sequences at their ends. The also give a double-stranded starting point, and because they match only the chosen sequence, only that section is copied. is used because it is thermostable: it does not denature at 95 °C, so it survives every cycle.

The photograph below shows a with its lid open.

A black laboratory machine on a bench. At the top left a hinged lid stands open above a square metal block with rows of round holes for small tubes. The front has a small display screen, a number keypad and buttons. — A thermal cycler with its lid open, showing the heating block for PCR tubes. Photo: Tinojasontran, Public domain.

The metal block holds the small tubes. It heats and cools rapidly to each set temperature.
The screen shows which step and cycle the programme has reached.

Lesson 6 of 21

What limits the polymerase chain reaction?

In theory doubles the DNA every cycle for ever. In practice the reaction slows down and has other weaknesses that scientists must .

The graph below compares the number of copies expected with what happens in a typical reaction. The vertical axis is a , so steady doubling appears as a straight line.

Line graph with the number of PCR cycles from 0 to 30 on the horizontal axis and log to base 10 of the number of copies of DNA from 0 to 10 on the vertical axis. A grey straight line, labelled expected if every cycle doubles the DNA, rises from 0 at cycle 0 to about 9 at cycle 30. A purple line, labelled typical real reaction, follows the grey line up to about cycle 16, then curves below it and levels off at about 7 from cycle 26 to 30. — Number of DNA copies during PCR, starting from one molecule. Original BioWelsh graph.

For the first 15 to 20 cycles the real reaction follows the doubling line.
After about 20 cycles it slows and then levels off at a plateau.
After 30 cycles there are about 10⁷ copies instead of the 10⁹ expected.

and free nucleotides are used up and become limiting.
slowly denatures after being heated to 95 °C many times.
At high concentrations the single strands of product pair with each other rather than with the .

Other limitations of PCR
Limitation	Why it matters
Contamination	Any stray DNA, from the air, the scientist or an earlier reaction, is copied too
Errors	cannot proofread, so wrong bases are copied into every later product
Fragment length	It works best for fragments of up to a few thousand base pairs, much shorter than many
Inhibitors	Substances in the sample, for example from soil or blood, can stop the reaction

Lesson 7 of 21

How does gel electrophoresis separate DNA fragments?

separates DNA fragments by length. It is used in , in and to check the products of .

The diagram below shows a gel seen from above during a run.

Top view of a pale purple gel in a pale blue tank. A dark bar marked with a minus sign runs across the top and a red bar marked with a plus sign across the bottom. Four white wells sit near the top. The left lane, labelled ladder, has seven bands labelled 1000, 800, 600, 400, 300, 200 and 100 base pairs from top to bottom. Sample 1 has one band level with about 500, sample 2 has bands near 700 and 200, and sample 3 has one band at 300. A downward arrow runs beside the gel. Labels name the negative electrode, the wells where samples are loaded at the negative end, the agarose gel as a mesh of pores that slows larger fragments more, and the positive electrode, to which DNA is attracted because its phosphate groups are negatively charged. — Gel electrophoresis of DNA seen from above. Original BioWelsh diagram.

The gel is made of , a polysaccharide from seaweed, which forms a mesh of tiny pores.
DNA samples are loaded into wells at the end next to the negative electrode (cathode).
A voltage is applied. DNA is negatively charged because of the phosphate groups in its backbone, so it moves towards the positive electrode (anode).
Small fragments move through the pores more easily, so they travel further in the same time. Large fragments stay near the wells.
A , a mixture of fragments of known length, is run in one lane so the length of each sample band can be estimated.

DNA is colourless, so the bands are made visible by staining the gel with a dye that glows under ultraviolet light, or by using fluorescent or radioactive labels. In the original method, the DNA was transferred to a nylon membrane () and located with labelled .

The photograph below shows samples being loaded into the wells of a gel.

Two hands in blue gloves hold a large multichannel pipette with eight tips over a clear gel in a tank of blue liquid. The tips point into a dashed dark blue line of small wells across the gel. — DNA samples being loaded into the wells of an agarose gel. Photo: U.S. Department of Agriculture, Public domain.

A multichannel pipette places a sample into each well at the same time.
The dark blue dashed line across the gel is the row of wells. The samples are dense, so they sink into the wells instead of floating away.

Lesson 8 of 21

How is a genetic fingerprint made?

About 99.9% of the DNA of any two people is identical. A () uses the small part that varies to produce a pattern of bands that is unique to each person, except identical twins. It does not show the base sequence of any .

The used are (): a short sequence of bases, often four, repeated one after another in non-coding DNA. The number of repeats at each varies between people, and each person inherits one version from each parent.

The diagram below shows two of one .

Two rows. Allele 1 has a grey flanking block, six gold blocks each labelled GATA, and a second grey flanking block. Allele 2 is the same but with nine GATA blocks. Green arrows above the left flanking block and below the right flanking block show where primers bind. Text reads that PCR copies everything between the two primers, so the product from allele 2 is 3 times 4, which is 12 bases longer. On the right, a gel lane headed one person shows two bands: the upper labelled allele 2 and the lower labelled allele 1, with the note longer fragment moves less far. — Two alleles of a short tandem repeat give PCR products of different lengths. Original BioWelsh diagram.

1 has six repeats of GATA and 2 has nine.
bind to the flanking DNA on either side of the repeats, so copies the repeats plus a little DNA each side.
The product from 2 is 12 bases longer, so on a gel it moves less far.
A person with two different shows two bands for this .

Making a

1) Extract DNA from cells, for example from blood, hair roots, saliva or a mouth swab
2) Copy several regions by , using labelled with fluorescent dyes
3) Separate the fragments by length with
4) Detect the labelled bands and compare the pattern with other samples

The first fingerprints were made a different way: DNA was cut with , separated by , blotted on to a membrane and located with radioactive . Modern profiling with needs far less DNA and is much faster.

Lesson 9 of 21

How are DNA profiles used, and what are the concerns?

Two samples come from the same person only if every band matches. Relatives share some bands because they share .

The first diagram below compares a crime scene sample with three suspects.

DNA profiles from a crime scene and three suspects. Original BioWelsh diagram.

Open full size

Suspect 2 has every band in the same position as the crime scene sample, so the DNA probably came from suspect 2.
Suspects 1 and 3 share one or two bands by chance, but most of their bands differ, so they are ruled out.

The second diagram is a paternity test.

A paternity test using DNA profiles. Original BioWelsh diagram.

Open full size

Every band in the child must come from the mother or the father.
First find the child's bands that match the mother (pink).
The remaining bands (blue) must come from the father. Man 2 has all of them and man 1 has none, so man 2 is very likely the father.

Forensic science: linking a suspect to a crime scene, and clearing people who were wrongly accused.
Paternity and other family relationships, including cases.
Telling identical twins, who share all bands, from non-identical twins.
: identifying the that seized ivory or other wildlife products came from.
Comparing to check how closely related they are.

Arguments for and against DNA profiling
For	Against
Needs only a tiny, non-invasive sample such as a mouth swab or hair	Collecting and storing profiles may breach privacy and civil liberties
Can rule people out and has overturned wrongful convictions	Databases could be hacked or used for other purposes
Very high probability of a correct match	A match gives a probability, not certainty, and samples can be contaminated or planted

Lesson 10 of 21

How is a gene obtained for genetic engineering?

Genetic engineering moves a from one organism into another. DNA made by combining DNA from two different sources is . The classic example is putting the human insulin into the bacterium Escherichia coli, so that bacteria make human insulin for people with diabetes.

First the has to be obtained. There are three ways to do it.

1. Cut it out with . (restriction endonucleases) come from bacteria. Each cuts DNA at a specific base sequence, its . A , a labelled single strand complementary to part of the , is used to find the right fragment.

The diagram below shows the EcoRI cutting DNA.

Top half, labelled before: two rows of coloured letter squares form double-stranded DNA reading C T G A A T T C G A on the top strand, 5 prime to 3 prime, and G A C T T A A G C T on the bottom strand. A red dashed box surrounds G A A T T C and its complement, labelled recognition sequence GAATTC, which reads the same 5 prime to 3 prime on both strands. A red dashed stepped line labelled cut runs down between G and A on the top strand, across, and down between A and G on the bottom strand. Bottom half, labelled after: two separate fragments. The left fragment has C T G on top and G A C T T A A underneath, with the unpaired T T A A boxed and labelled sticky end. The right fragment has A A T T C G A on top and G C T underneath, with the unpaired A A T T boxed and labelled sticky end. — A restriction enzyme cuts at its recognition sequence and leaves sticky ends. Original BioWelsh diagram.

EcoRI recognises the sequence GAATTC. The sequence is a palindrome: it reads the same 5′ to 3′ on both strands.
It cuts each strand between G and A, so the cuts are staggered.
This leaves short single-stranded overhangs, AATT, called . They pair readily with complementary made by the same .

2. Make it from mRNA with . Pancreatic β cells contain many copies of insulin mRNA. , an from retroviruses, uses the mRNA as a template to make a single strand of (). DNA polymerase then makes the second strand.

The diagram below shows how is made.

Four numbered panels joined by arrows. Panel 1, extract mRNA, shows a pink oval cell with a purple nucleus and three wavy red strands labelled mRNA. Panel 2, reverse transcriptase, shows a wavy red mRNA strand above a straight blue strand labelled cDNA, joined by grey rungs, with a yellow oval enzyme at the end of the blue strand. Panel 3, remove the mRNA, shows a single blue strand labelled single-stranded cDNA. Panel 4, DNA polymerase, shows the blue strand paired with a green strand, labelled double-stranded DNA. Text below reads that mature mRNA has already had its introns removed, so the cDNA contains only the coding sequence. — Making a copy of a gene from mRNA with reverse transcriptase. Original BioWelsh diagram.

Mature mRNA has had its removed, so the contains only the coding sequence ().
This matters because bacteria cannot remove . A cut straight from human DNA would include them, and the protein made would not work.

3. Build it artificially. Because the base sequence is known from the , a can be synthesised base by base in a machine.

Advantages of cDNA or a synthesised gene over cutting the gene from human DNA
Problem with restriction enzymes	How the other methods avoid it
The has to be found among about 20 000 , using	No searching: start from the mRNA or the known sequence
The may cut inside the	The whole is made in one piece
Human contain that bacteria cannot remove	and synthetic contain only

Lesson 11 of 21

How is a gene put into a plasmid and then into a bacterium?

Bacteria rarely take up a loose , so it is carried in by a . The usual is a : a small circle of double-stranded DNA found in bacteria, separate from the main chromosome, that copies itself.

The diagram below shows how the is made.

Four numbered panels joined by arrows. Panel 1, cut the plasmid open, shows a blue ring with a gap at the top labelled sticky ends and a gold section labelled antibiotic resistance gene. Panel 2, cut out the gene, shows a short pink bar labelled gene with small gold squares at each end. Panel 3, mix and join, shows the pink gene above the gap in the blue ring, with a downward arrow labelled DNA ligase. Panel 4 shows a closed blue ring with a pink section at the top, labelled recombinant plasmid. Text below reads that calcium chloride and a brief heat shock at about 42 degrees Celsius make the bacterial membranes more permeable so the plasmid can enter. — Putting a gene into a plasmid to make recombinant DNA. Original BioWelsh diagram.

The is cut open with a at one .
The is cut with the same , so the of the and the are complementary.
When they are mixed, the pair by complementary base pairing (hydrogen bonds).
then joins the sugar-phosphate backbones by making phosphodiester bonds, sealing the into the .
The result is a .

Next the are mixed with bacteria. Calcium ions and a brief heat shock, from about 4 °C to 42 °C, make the membranes more permeable, and a small fraction of the bacteria take up a . A bacterium that has taken up new DNA is said to be .

It copies itself inside the cell.
It is small and easy to take up.
It is not broken down by the 's .
It carries so that cells containing it can be identified.

Lesson 12 of 21

How are the bacteria that took up the gene identified?

Only a small fraction of bacteria take up a , and some close up without the inside. on the show which bacteria have what.

The diagram below shows one common method: a with two , and .

On the left, a blue ring plasmid with a green section labelled tetracycline resistance gene and a gold section labelled ampicillin resistance gene, with a short pink section in the middle of the gold labelled inserted gene. Text beneath says the inserted gene splits the ampicillin resistance gene, so it no longer works. In the centre, a circular agar plate labelled plate A, agar with tetracycline, has eight white colonies. An arrow through a box labelled velvet pad points to plate B, agar with ampicillin, which has the same pattern but three colonies are replaced by red dashed circles, explained as colonies that did not grow. Three numbered notes explain that plate A colonies took up a plasmid, the missing colonies carry the recombinant plasmid, and those colonies are picked from plate A. — Finding transformed bacteria with two resistance genes and replica plating. Original BioWelsh diagram.

The carries for to tetracycline and to ampicillin. The new is inserted into the middle of the ampicillin , which stops that working.
The bacteria are grown on with tetracycline (plate A). Only bacteria that took up a , with or without the new , can grow.
A sterile velvet pad presses the from plate A on to plate B, which contains ampicillin, in exactly the same pattern.
that grow on A but are missing on B are to tetracycline but not ampicillin: they carry the .
These are picked from plate A and grown on.

Other work the same way. A for a fluorescent protein can make glow under ultraviolet light. In blue-white , the new is inserted into the lacZ , which would otherwise turn a chemical called X-gal blue: with the stay white.

Finally, scientists check that the chosen cells really make the product, because a that has been inserted is not always expressed.

Lesson 13 of 21

How do bacteria make human insulin, and what are the pros and cons?

Producing human insulin with bacteria

1) Obtain the insulin ( from β cell mRNA, or a synthesised )
2) Insert it into with a and
3) Transform E. coli with the
4) Select bacteria using
5) Grow them in large fermenters; each cell's are copied every time it divides
6) Bacteria transcribe and translate the ; the insulin is extracted and purified

Large numbers are easy to reach. If each bacterium makes 3 × 10⁶ molecules of a protein and a fermenter contains 1 × 10⁸ bacteria, the total is 3 × 10⁶ × 1 × 10⁸ = 3 × 10¹⁴ molecules.

Advantages and concerns of genetically modified bacteria
Advantages	Concerns
Human insulin is identical to natural insulin, so it causes fewer immune reactions than animal insulin	pass easily between bacteria, so could spread to
Large amounts are made cheaply and reliably, without using animals	Human DNA used could contain sequences that switch on
Other products: clotting factors for and human growth hormone, which are safer than old extracts from donors	Modified bacteria released by accident could harm the environment
Modified bacteria can also clean up pollution and detect such as arsenic	Bacteria cannot make every human protein correctly

Lesson 14 of 21

How are genetically modified crops made?

A () crop has had its DNA changed by technology. If the new comes from another , the plant is . The aim is usually to protect the crop from pests or disease, or to give it a useful feature.

The most common method uses a soil bacterium, Agrobacterium tumefaciens. The diagram below shows the steps.

Five numbered panels joined by arrows. Panel 1 shows a rounded green bacterium containing a wavy chromosome and a small blue ring labelled Ti plasmid. Panel 2 shows a blue ring with a gap, labelled opened plasmid. Panel 3 shows a closed blue ring with a pink section labelled chosen gene. Panel 4 shows a green square plant cell with a purple nucleus and a small bacterium containing a ring. Panel 5 shows a seedling with two leaves growing from a cream block, labelled transgenic plantlet. Captions describe removing the plasmid, cutting it and removing the tumour-forming genes, inserting the chosen gene with DNA ligase, returning the plasmid to the bacterium to infect plant cells, and growing the cells in tissue culture into plants. — Making a transgenic plant with the Ti plasmid of *Agrobacterium tumefaciens*. Original BioWelsh diagram.

A. tumefaciens carries a Ti (-inducing) . Normally part of this enters plant cells and causes a called a .
The is cut with a and the -forming are removed.
The chosen is inserted with .
The bacteria infect plant cells, and the section of carrying the joins a plant chromosome.
The cells grow into whole plants in tissue culture. Every cell, including the seeds, carries the .

The photograph below shows on a plant infected by A. tumefaciens with its normal , which is why the are removed first.

A potted plant with finely divided green leaves. Its stem, from the soil up to the lowest leaves, is swollen into a stack of large, rough, pale brown knobbly lumps. A black plastic pot with a handwritten label is at the bottom. — Crown galls on a plant infected by *Agrobacterium tumefaciens*. Photo: Cazadores de Microbios, CC0.

The swollen, knobbly lumps on the stem near the soil are the galls.
Their cells divide out of because of from the bacterium's .

A gun fires tiny gold or tungsten particles coated with DNA into plant cells.
Electroporation uses an electric field to make membranes more permeable to DNA.

Examples of GM crops
Crop	Gene added	Effect
tomato, maize and cotton	for an insecticidal protein from the bacterium thuringiensis	Insects that eat the leaves are killed, so less insecticide spraying is needed
Herbicide- soya	for an that is not affected by the herbicide glyphosate	Fields can be sprayed to kill weeds without harming the crop
Golden Rice	to make beta-carotene, which the body turns into vitamin A	Helps prevent vitamin A deficiency and blindness

Lesson 15 of 21

What are the arguments for and against GM crops?

crops could help feed a growing world , but they raise scientific, environmental and economic concerns. A good answer weighs both sides.

Arguments about GM crops
For	Against
Higher yields, because fewer plants are lost to pests, disease, drought or salty soil	could carry the new to wild relatives, for example making herbicide- weeds
Less insecticide is sprayed, protecting insects that are not pests	Insect pests may evolve to the , because the crop applies a strong
Food can be made more nutritious, as in Golden Rice	Unknown long-term effects of eating new proteins, although no harm has been shown
Crops can make medicines and (pharming)	Beneficial insects and food chains may be affected, reducing biodiversity
Crops that fix their own nitrogen could reduce use	may contaminate organic crops, and farmers may depend on a few seed companies

in pests is . A few insects carry an that lets them survive the . They breed and pass it on, so the becomes more common. Growing some non- plants nearby, or crops with two different , slows this down.

Lesson 16 of 21

What is gene therapy, and what are its advantages and disadvantages?

treats a genetic disorder by putting a working (functional) into a patient's cells, or by correcting the faulty one. It is most useful for disorders caused by a single .

Somatic and germ line gene therapy
Feature	Somatic cell therapy	Germ line therapy
Cells changed	Body cells in the affected tissue	Gametes, a or an early embryo
Passed to children?	No	Yes, to all later generations
Status	Used in treatment and trials	Not allowed in humans in the UK

Viruses: altered so they cannot cause disease, they carry the into cells.
: tiny spheres of phospholipid that fuse with cell membranes.
, or naked DNA injected directly.
editing, such as , which cuts the DNA at the faulty so it can be repaired.

In , carrying the normal CFTR can be breathed in as an aerosol. They fuse with the epithelial cells lining the airways, which then make working CFTR channels. The effect lasts only as long as those cells, so treatment has to be repeated.

For sickle cell disease, can be taken from the patient's own bone marrow, corrected in the laboratory (for example with ), and returned. The corrected keep dividing and make red blood cells with normal haemoglobin, so one treatment can last.

Advantages and disadvantages of gene therapy
Advantages	Disadvantages
Treats the cause, not just the symptoms	Only a small proportion of the introduced are expressed
Using the patient's own cells less chance of rejection	The may trigger an immune response
Changes to can last for years	A virus could insert the in the wrong place and activate an
Treating cells affects only the patient	treatment must often be repeated; germ line changes have unknown effects on future generations

Lesson 17 of 21

How can exon skipping treat Duchenne muscular dystrophy?

() is caused by a of the on the X chromosome, so it mostly affects boys. holds the membranes of steady. Without it, muscle cells are damaged and die, and muscles waste away. (If inheritance is unfamiliar, read the Inheritance lesson on sex linkage.)

The is huge, with 79 . Most cases of are deletions of one or more . If the number of bases deleted is not a multiple of three, the shifts: every codon after the deletion is read wrongly, a stop codon appears early, and no is made.

The diagram below shows how works, using one real example.

Four rows of labelled blocks. Healthy gene: exons 48, 49, 50, 51 and 52, labelled in frame, full-length dystrophin. DMD, exon 50 deleted: exon 50 is a dashed empty box labelled 50 lost, and the row is labelled reading frame shifted, early stop codon, no dystrophin. Exon skipping drug: the same row with a green bar above exon 51 labelled antisense oligonucleotide, labelled frame restored, shorter, partly working dystrophin. An arrow leads down to mRNA after splicing: exons 48, 49 and 52 only. Text beside it says the patch makes exon 51 be spliced out too, and exon 49 now joins exon 52 in the correct reading frame. — Exon skipping restores the reading frame of the dystrophin mRNA. Original BioWelsh diagram.

In a healthy , the join in the correct and full-length is made.
When 50 is deleted, 49 joins 51 and the shifts, so no is made.
The drug is an , a short nucleic acid complementary to part of 51. It binds to the pre-mRNA like a .
The patched is skipped during splicing, so 49 joins 52. The is restored.
A shorter is made. It is missing a few amino acids in the middle but still partly works, so the disease is milder.

Each drug skips one particular , so it helps only patients with matching deletions.
The mRNA is changed, not the DNA, so injections must be repeated for life.
The drug has to reach muscles all over the body, including the heart.

Lesson 18 of 21

How could genomics change healthcare?

is the study of the structure and function of whole genomes. It combines DNA sequencing, computing (bioinformatics) and medical records. As sequencing becomes faster and cheaper, doctors can use a patient's to guide care. This is called personalised or precision medicine.

Ways genomics is changing healthcare
Use	Example
Faster, more accurate diagnosis	Sequencing a child with an undiagnosed rare disease can find the in weeks instead of years
Choosing the right dose ()	Variants of liver change how fast drugs such as warfarin or codeine are processed, so the dose can be adjusted
Targeted cancer treatment	Tumours with extra copies of the HER2 respond to a drug that targets the HER2 protein
Predicting risk	People with that increase the risk of some cancers can be screened more often
Tracking infections	Sequencing a virus or bacterium shows how an outbreak spreads and whether it is drug

Many diseases depend on many and the environment, so risks are often only probabilities.
The data must be stored securely and used only with consent.
Tests and treatments are expensive, so not everyone may have equal access.

Lesson 19 of 21

What are stem cells, and how do the different types compare?

A is an undifferentiated cell that can divide repeatedly by mitosis and can into specialised cells. Each daughter cell can either stay a or become specialised. differ in how many cell types they can become.

The diagram below shows the main types.

A row of four stages joined by arrows. A pink circle with a small nucleus is labelled zygote, totipotent. A large pale blue circle ringed with small pink cells and a cluster of purple cells inside at the upper left is labelled blastocyst, about 5 days, inner cell mass, embryonic stem cells, pluripotent. A cream box labelled adult tissue, for example bone marrow, contains one cell and is labelled adult stem cells, multipotent. Three circles are labelled red blood cell, white blood cell and platelets, with the note a few related cell types. A box below explains that induced pluripotent stem cells are adult cells such as skin fibroblasts reprogrammed by adding a few genes, so they become pluripotent again. — Totipotent, pluripotent and multipotent stem cells. Original BioWelsh diagram.

cells, such as the , can become any cell, including the cells of the .
cells, such as embryonic from the of a , can become any cell type in the body.
cells, the adult found in tissues such as bone marrow, can become only a few related cell types, for example the different blood cells.
Induced stem () cells are adult cells, such as skin cells, reprogrammed by adding a few so that they become again.

The micrograph below shows a of human embryonic growing in culture.

Grey microscope image. A large, dense, roughly rounded patch of small, tightly packed cells fills the centre. All around it, long thin spindle-shaped cells lie side by side in flowing streams. — Phase contrast micrograph of a colony of human embryonic stem cells on a layer of feeder cells. Photo: Id711 (Wikimedia Commons), Public domain.

The dense, rounded patch in the middle is the of small, tightly packed .
The long, thin cells around it are a feeder layer of mouse fibroblasts that support the .

Embryonic and adult stem cells compared
Feature	Embryonic stem cells	Adult stem cells
Potency	: any body cell	: a few related cell types
Source	of an embryo a few days old	Tissues such as bone marrow
Growth in culture	Divide readily in large numbers	Fewer and harder to grow
Rejection	From another person, so may be rejected	Can come from the patient, so less likely to be rejected
Ethics	The embryo is destroyed	Few ethical objections

Lesson 20 of 21

How can stem cells replace damaged tissues, and what are the issues?

The need for tissues and organs for transplant is far greater than the supply of donors. offer a renewable source of replacement cells. Examples include bone marrow transplants for leukaemia, which have been used for decades, and trials for type 1 diabetes, damaged heart muscle, spinal cord injury and sight loss.

grows new tissue outside the body. The diagram below shows the steps.

Four numbered panels joined by arrows. Panel 1, take cells, shows five pink cells with purple nuclei, captioned from the patient so they are less likely to be rejected. Panel 2, multiply them, shows a row of pink cells in a cream culture dish, captioned in culture with nutrients, oxygen and growth factors. Panel 3, seed a scaffold, shows pink cells scattered on a grey mesh, captioned a porous, biodegradable 3D framework the cells attach to. Panel 4, implant the tissue, shows a pink block with wavy lines labelled new tissue, captioned the scaffold breaks down as the new tissue forms. — Tissue engineering on a biodegradable scaffold. Original BioWelsh diagram.

Cells are taken, ideally from the patient ( cells), so the new tissue is less likely to be rejected.
They are multiplied in culture with nutrients, oxygen and , at the right temperature and pH.
They are seeded on to a : a porous, biodegradable 3D framework that the cells can attach to and that lets nutrients diffuse in.
The tissue is implanted. The slowly breaks down as the cells make their own tissue.

is a way to make embryonic that match a patient. The of a body cell is placed into an egg cell whose has been removed, the cell grows into an early embryo, and are taken from it. In the UK is allowed under licence, and reproductive cloning of humans is illegal.

Issues with using embryonic stem cells
Arguments against	Arguments for
The embryo is destroyed, and some believe life begins at fertilisation	Embryos are used only up to 14 days old, before any nervous system forms
Embryos might be created only to be used	Most are spare embryos from that would be discarded anyway
Knowledge could be misused for reproductive cloning	The research could treat many serious diseases

Adult and avoid destroying embryos. They can come from the patient, so there is less chance of rejection. However, adult can become only a few cell types, and may carry a risk of forming tumours. All treatments are expensive, and their long-term effects are still being studied.

Lesson 21 of 21

Putting the topic together

How the ideas in applied genetics connect

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Application of reproduction and genetics
- Reading genomes
  - Human and 100K projects; ethics and
  - Other genomes: Anopheles and Plasmodium
  - in healthcare
- Tools for working with DNA
  - copies DNA
  - separates by size; genetic fingerprints
- Changing DNA
  - : insulin in bacteria
  - crops
  - and
  - and

At 95 °C only the hydrogen bonds between the strands break.
must withstand 95 °C, not just 70 °C.
DNA moves to the positive electrode because of its phosphate groups; small fragments move furthest.
Use the same on the and the ; joins the backbones.
makes DNA from mRNA, which has no .
is not inherited; say less chance of rejection, not no rejection.
gives a shorter, partly working .

What were the Human Genome Project and the 100K Genome Project?

What ethical issues does knowing a person's genome raise?

How are embryos screened for genetic disorders, and is it right?

Why sequence the genomes of other organisms, such as the malaria mosquito?

How does the polymerase chain reaction copy DNA?

What limits the polymerase chain reaction?

How does gel electrophoresis separate DNA fragments?

How is a genetic fingerprint made?

How are DNA profiles used, and what are the concerns?

How is a gene obtained for genetic engineering?

How is a gene put into a plasmid and then into a bacterium?

How are the bacteria that took up the gene identified?

How do bacteria make human insulin, and what are the pros and cons?

How are genetically modified crops made?

What are the arguments for and against GM crops?

What is gene therapy, and what are its advantages and disadvantages?

How can exon skipping treat Duchenne muscular dystrophy?

How could genomics change healthcare?

What are stem cells, and how do the different types compare?

How can stem cells replace damaged tissues, and what are the issues?

Putting the topic together

Worksheets

Worksheet 1: 4.5 Applied genetics (154 marks)

Check your understanding.

Which of the following was an aim of the Human Genome Project?

Application of reproduction and genetics

What were the Human Genome Project and the 100K Genome Project?BWRBioWelsh recommendedWe recommend revising this because it has appeared regularly in past papers. Asked in 3 of 7 WJEC exam series (2017 to 2025), 12 marks in total. This is a revision priority, not a prediction of your exam.

What ethical issues does knowing a person's genome raise?

How are embryos screened for genetic disorders, and is it right?

Why sequence the genomes of other organisms, such as the malaria mosquito?

How does the polymerase chain reaction copy DNA?BWRBioWelsh recommendedWe recommend revising this because it has appeared regularly in past papers. Asked in 3 of 7 WJEC exam series (2017 to 2025), 16 marks in total. This is a revision priority, not a prediction of your exam.

What limits the polymerase chain reaction?BWRBioWelsh recommendedWe recommend revising this because it has appeared regularly in past papers. Asked in 3 of 7 WJEC exam series (2017 to 2025), 16 marks in total. This is a revision priority, not a prediction of your exam.

How does gel electrophoresis separate DNA fragments?

How is a genetic fingerprint made?

How are DNA profiles used, and what are the concerns?

How is a gene obtained for genetic engineering?BWRBioWelsh recommendedWe recommend revising this because it has appeared regularly in past papers. Asked in 2 of 7 WJEC exam series (2017 to 2025), 16 marks in total. This is a revision priority, not a prediction of your exam.

How is a gene put into a plasmid and then into a bacterium?BWRBioWelsh recommendedWe recommend revising this because it has appeared regularly in past papers. Asked in 2 of 7 WJEC exam series (2017 to 2025), 16 marks in total. This is a revision priority, not a prediction of your exam.

How are the bacteria that took up the gene identified?BWRBioWelsh recommendedWe recommend revising this because it has appeared regularly in past papers. Asked in 2 of 7 WJEC exam series (2017 to 2025), 14 marks in total. This is a revision priority, not a prediction of your exam.

How do bacteria make human insulin, and what are the pros and cons?

How are genetically modified crops made?

What are the arguments for and against GM crops?

What is gene therapy, and what are its advantages and disadvantages?BWRBioWelsh recommendedWe recommend revising this because it has appeared regularly in past papers. Asked in 3 of 7 WJEC exam series (2017 to 2025), 14 marks in total. This is a revision priority, not a prediction of your exam.

How can exon skipping treat Duchenne muscular dystrophy?BWRBioWelsh recommendedWe recommend revising this because it has appeared regularly in past papers. Asked in 2 of 7 WJEC exam series (2017 to 2025), 12 marks in total. This is a revision priority, not a prediction of your exam.

How could genomics change healthcare?

What are stem cells, and how do the different types compare?BWRBioWelsh recommendedWe recommend revising this because it has appeared regularly in past papers. Asked in 3 of 7 WJEC exam series (2017 to 2025), 7 marks in total. This is a revision priority, not a prediction of your exam.

How can stem cells replace damaged tissues, and what are the issues?BWRBioWelsh recommendedWe recommend revising this because it has appeared regularly in past papers. Asked in 3 of 7 WJEC exam series (2017 to 2025), 7 marks in total. This is a revision priority, not a prediction of your exam.

Putting the topic together

Worksheets

Worksheet 1: 4.5 Applied genetics (154 marks)

Check your understanding.

What were the Human Genome Project and the 100K Genome Project?

How does the polymerase chain reaction copy DNA?

What limits the polymerase chain reaction?

How is a gene obtained for genetic engineering?

How is a gene put into a plasmid and then into a bacterium?

How are the bacteria that took up the gene identified?

What is gene therapy, and what are its advantages and disadvantages?

How can exon skipping treat Duchenne muscular dystrophy?

What are stem cells, and how do the different types compare?

How can stem cells replace damaged tissues, and what are the issues?