Questions on genome analysis, genetic screening and gene therapy

Question 1

Explain why the sequenced genome of a bacterium gives its proteome almost directly, while the sequenced genome of a human does not.

Accepted Answer

B1 bacterial genes are continuous, with no introns, so one gene gives one polypeptide and the coding sequence can be read straight off the DNA B1 human genes contain introns, which have to be identified and removed before the coding sequence is known B1 alternative splicing lets one human gene give several different polypeptides B1 large stretches of the human genome are regulatory rather than coding, and post-translational modification changes the polypeptide again after translation

Question 2

Compare somatic gene therapy with germ line gene therapy, referring to which cells are altered, what is inherited, and how each stands in law.

Accepted Answer

B1 somatic therapy alters body cells such as bone marrow, retina or airway epithelium, whereas germ line therapy alters a gamete, a zygote or an early embryo B1 a somatic change is not inherited, because the cells that become sperm or eggs are untouched, whereas a germ line change is present in every cell of the resulting person, including those cells, and passes to every descendant B1 somatic treatment often has to be repeated, because cells that are not stem cells are replaced within weeks, whereas a germ line change is permanent and cannot be withdrawn from the family line B1 somatic therapy is licensed for several conditions, whereas clinical use of heritable genome editing is not permitted in the UK and is banned or unauthorised in most countries that regulate it; credit any named jurisdiction stated correctly

Question 3

Trastuzumab is given only to patients whose breast tumour has been tested first, while patients starting azathioprine may have their TPMT genotype tested first. Explain what each test is for, and explain why the two tests read different genomes.

Accepted Answer

B1 trastuzumab is an antibody with a binding site complementary to the HER2 receptor, so it can only work on a tumour that over-expresses HER2 and the test decides whether the drug is given at all B1 TPMT is the enzyme that breaks azathioprine down, so a patient with two low-activity alleles would accumulate the drug and suffer bone marrow damage, and the test decides the dose rather than the drug B1 the HER2 test reads the genotype the tumour has acquired through mutation, which the rest of the patient's cells do not carry B1 the TPMT test reads alleles the patient inherited, which are present in every cell and will not change

Question 4

A health minister proposes sequencing the genome of every newborn so that all later prescribing can be personalised. Evaluate this proposal.

Accepted Answer

B1 in favour, a drug that will not work for a patient can be avoided rather than trialled, and doses can be set to prevent adverse reactions such as the bone marrow damage caused by a standard thiopurine dose in a TPMT-deficient patient B1 against, the sequencing and the interpretation cost money and staff time that the health service must fund before any saving appears B1 against, the reference databases used to judge what a variant means are drawn mostly from people of European ancestry, so the benefit would be smaller for the patients already served worst B1 a supported judgement, for example that it is worth doing for the small set of single-gene drug-handling variants with clear evidence, but not as a prediction of common disease, which is polygenic and strongly environmental

Question 5

Compare genetic screening with genetic counselling, referring to what each one does and to the meaning of non-directive.

Accepted Answer

B1 genetic screening is the laboratory test itself, carried out on an individual to look for particular alleles, whereas genetic counselling is the conversation around the test, explaining what it can and cannot show B1 counselling works out the risk to the person and to any children, and sets out the options available, which screening alone does not do B1 counselling is meant to be non-directive, meaning the counsellor does not recommend a course of action, because the decision belongs to the person or family being counselled B1 screening can happen without counselling, as in newborn heel-prick testing, but a positive predictive result for an adult-onset condition is normally accompanied by counselling before and after the test

Question 6

A woman is found to carry a pathogenic BRCA1 variant; a separate patient is found to carry the allele for Huntington's disease. Compare what each result means for the person who receives it, referring to penetrance.

Accepted Answer

B1 the BRCA1 result raises the woman's lifetime risk of breast cancer from roughly twelve to fifteen per cent in the general population to about sixty to seventy per cent, which is a large increase but not a diagnosis, since some carriers never develop cancer B1 the Huntington's disease result is different because penetrance of the allele is essentially complete, so the result says the disease will develop at some point, though not when B1 the BRCA1 result changes what the woman might reasonably choose to do, such as more frequent screening or risk-reducing surgery, whereas nothing currently changes the course of Huntington's disease B1 both results are examples of predictive genetic testing, but the certainty of the outcome each one predicts is completely different, which is why many people at risk of Huntington's disease choose not to be tested

Question 7

Describe how a DNA profile is obtained from a sample such as blood or saliva, using the terms variable number tandem repeat and PCR.

Accepted Answer

B1 DNA is extracted from the sample and a set of loci is chosen, each a variable number tandem repeat that differs in length between people B1 each locus is amplified by PCR, using primers that bind to the constant sequences flanking the repeat, so the length of the product depends only on how many repeats that person has B1 the amplified products are separated by gel electrophoresis according to their length B1 the pattern of fragment lengths obtained, one from each parent at every locus examined, is the profile, and the UK standard test examines sixteen such loci plus a marker that indicates sex

Question 8

At one locus a mother has alleles of 9 and 14 repeats, and her child has alleles of 9 and 11 repeats. Man A has alleles of 11 and 16; man B has alleles of 7 and 14. Identify which man could be the father, identify which man is excluded, and identify what a single matching locus like this does and does not show.

Accepted Answer

B1 the child's allele of 9 repeats could have come from the mother, since she carries a 9 B1 the child's other allele, 11 repeats, is not carried by the mother, so it must have come from the biological father, who must therefore carry an 11 B1 man B, with alleles 7 and 14, carries no 11 at this locus and is excluded here, which is strong evidence against paternity (confirmed at further loci, since repeat numbers occasionally mutate), whereas man A, with alleles 11 and 16, could be the father, since he is not excluded B1 a single matching locus only shows that man A is not excluded, since an 11-repeat allele may be carried by a substantial fraction of the population; it does not by itself demonstrate paternity, which needs the same result repeated across fifteen or more loci

Question 9

Explain how a fully sequenced and annotated bacterial genome can be used to identify a candidate antigen for a new vaccine without culturing the pathogen itself, and explain why the same approach is much harder for a eukaryotic parasite.

Accepted Answer

B1 annotation of a bacterial genome is relatively direct because bacterial genes are continuous with no introns, so the sequence can be translated straight into a predicted list of the proteins the bacterium can make B1 a surface protein that every strain carries and that the human host does not make is a candidate vaccine antigen among those predicted proteins, since it is one the immune system would meet first B1 screening the predicted protein list on a computer is faster and safer than growing the pathogen to find its surface proteins directly, which matters most for organisms dangerous to culture B1 the same route is much harder in a eukaryotic parasite because introns must be identified and removed, alternative splicing can give one gene several polypeptides, and much of the genome is regulatory rather than coding

Question 10

Explain how a labelled DNA probe is used to find out whether a person carries a particular allele.

Accepted Answer

B1 the probe is a short single strand of DNA whose base sequence is complementary to the sequence being looked for, carrying a radioactive or a fluorescent label B1 the sample DNA is made single-stranded by heating and the probe is added, and hybridisation occurs wherever the probe meets a complementary sequence, held by hydrogen bonds between complementary bases B1 unbound probe is washed away, and the label is then detected, on photographic film or under ultraviolet light, only where the probe has bound

Question 11

A profile from a crime scene matches a suspect, and the court is told that the probability of such a match in an unrelated person chosen at random is one in a billion. A lawyer says there is therefore a one in a billion chance that the suspect is innocent. Explain why that statement is wrong.

Accepted Answer

B1 the quoted figure is the probability of obtaining a matching profile given that the person is not the source of the sample B1 the lawyer has reversed it into the probability that the person is not the source given that the profile matches, which is a different quantity and is not what the figure measures; reversing the two is the prosecutor's fallacy B1 the chance that the suspect is not the source depends on how many other people could have left the sample and on the rest of the evidence, which the match probability does not include; accept that the figure assumes an unrelated person, and relatives share far more alleles

Question 12

Explain the difference between what a DNA microarray shows when labelled DNA from a sample is washed over it, and what it shows when labelled cDNA made from a tissue's mRNA is washed over it.

Accepted Answer

B1 washing labelled DNA from a sample over a microarray identifies which alleles at each probed position that person carries, which is a measurement of genotype B1 washing labelled cDNA made from a tissue's mRNA over a microarray identifies which of those genes the tissue is transcribing at the time the sample was taken, which is a measurement of expression rather than genotype B1 the microarray itself works the same way in both cases, by hybridisation to a complementary sequence, so the difference lies entirely in what is used as the starting material

Question 13

Explain why genotyping every patient's CYP2C9 and VKORC1 variants before starting warfarin has not been shown to give a clear benefit over the existing practice of adjusting the dose by monitoring, even though the mechanism linking genotype to dose is well established.

Accepted Answer

B1 common variants of CYP2C9 and VKORC1 do shift the dose of warfarin a patient needs by several times, which is a genuine and well-established mechanism B1 existing practice starts every patient on a low dose and adjusts it using blood clotting tests, and this monitoring identifies the correct dose for a given patient within a week or two in any case B1 because monitoring reaches the same dose about as quickly as genotyping would predict it, trials comparing the two approaches have not shown a clear improvement in outcome from genotyping first

Question 14

Describe the two kinds of vector used to deliver a gene into a patient's cells in somatic gene therapy, and describe one limitation of using a viral vector.

Accepted Answer

B1 a modified virus, stripped of the genes that would let it cause disease and carrying the therapeutic gene instead, can be used to carry the gene into cells B1 alternatively a liposome, a small sphere of phospholipid that fuses with the cell-surface membrane, can be used to deliver the gene without any virus at all B1 a limitation of a viral vector is that the patient's immune system may attack it, or that a vector integrating at a random position in the genome may land inside another gene, such as a proto-oncogene, causing serious harm

Question 15

State what a genome project produces, and state what annotation adds to that product afterwards.

Accepted Answer

B1 a sequence of bases for the organism, coding and non-coding alike B1 annotation locates the genes within that sequence and works out which non-coding regions do something, and it is a separate and slower job done afterwards

Person	Alleles at this locus (repeats)
Mother	9, 14
Child	9, 11
Man A	11, 16
Man B	7, 14