19.2. Genetic technology applied to medicine
A subsection of Biology, 9700, through 19. Genetic technology
Listing 10 of 37 questions
In humans, the gene RPE65 encodes a protein responsible for regenerating visual pigment in rod and cone cells after they have been exposed to light. A recessive allele of this gene causes impaired vision from birth, progressing to complete blindness in early adulthood. This condition is called LCA. In 2008, trials were carried out into the possibility and safety of treating LCA using gene therapy. Suggest and explain why LCA is suitable for treatment using gene therapy. Six adults with this condition were used in the study. Genetically modified adenoviruses (a type of virus that can cause respiratory infections) were used as vectors. The vectors were injected beneath the retina of one eye of each of the participants. Suggest two ways in which the genome of the adenoviruses used as vectors would differ from that of normal adenoviruses. 1. 2. Improvements were found in the vision of all the participants, but the small number in the trials made most of these improvements not statistically significant. Suggest why these trials were designed to include such a small number of participants.
9700_w11_qp_42
THEORY
2011
Paper 4, Variant 2
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Insulin can be produced on a large scale using gene technology and prokaryotes such as Escherichia coli. Table 7.1 summarises the sequence of steps in one method of production of insulin by E. coli. Complete Table 7.1 by adding one statement in each of the empty boxes. Table 7.1 step reason for step obtain copies of gene with sticky ends the gene codes for the synthesis of insulin acts as a vector for the transfer of the gene into the host use restriction endonuclease enzyme mix vector and gene to seal the sugar-phosphate backbone to obtain transformed host E. coli cells screen for, and obtain, successfully transformed cells to obtain large amounts of insulin for extraction and purification
9700_w11_qp_42
THEORY
2011
Paper 4, Variant 2
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The steps involved in a method of production of human insulin by gene technology are listed in Table 5.1. The steps are not listed in the correct order. Table 5.1 step description A DNA coding for human insulin inserted into cut plasmid vector B genetically modified bacteria identified C mRNA for human insulin isolated in β cells D plasmid vector inserted into bacterium E genetically modified bacteria cloned F DNA for human insulin cloned G human insulin harvested H cDNA coding for human insulin synthesised Complete Table 5.2 to show the steps in the correct order. Two of the steps have been done for you. Table 5.2 correct order letter of step C D Name the enzymes responsible for the following steps: step A step H Explain two advantages of treating diabetes with human insulin produced by gene technology rather than using insulin from animals.
9700_w11_qp_43
THEORY
2011
Paper 4, Variant 3
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Human insulin can be synthesised in a laboratory strain of Escherichia coli using recombinant DNA (rDNA) technology. The starting point for the process is mRNA coding for insulin, isolated from human pancreas cells. Four enzymes are needed: • reverse transcriptase • DNA polymerase • restriction enzyme • DNA ligase. State the role of each of these enzymes in producing rDNA carrying the gene for human insulin. reverse transcriptase DNA polymerase restriction enzyme DNA ligase Outline the role of insulin in a healthy human. Describe and explain one advantage of treating diabetics with human insulin produced by rDNA technology. It is possible to use rDNA technology to produce insulin with a slightly different structure from that of human insulin. The effect of the changed structure can then be investigated. The activities of equal quantities of two insulins, both produced by E. coli, were compared in healthy, non-diabetic subjects: • human insulin • insulin X, in which the positions of two amino acids, lysine and proline, were exchanged. Lysine has a hydrophilic R group and proline has a hydrophobic R group. The results of the investigation are shown in . activity of insulin / arbitrary units time after injection / hours insulin X human insulin With reference to describe the differences in activity between human insulin and insulin X. Suggest how exchanging the position of two amino acids in the insulin molecule can result in differences in activity.
9700_w14_qp_41
THEORY
2014
Paper 4, Variant 1
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Human insulin can be synthesised in a laboratory strain of Escherichia coli using recombinant DNA (rDNA) technology. The starting point for the process is mRNA coding for insulin, isolated from human pancreas cells. Four enzymes are needed: • reverse transcriptase • DNA polymerase • restriction enzyme • DNA ligase. State the role of each of these enzymes in producing rDNA carrying the gene for human insulin. reverse transcriptase DNA polymerase restriction enzyme DNA ligase Outline the role of insulin in a healthy human. Describe and explain one advantage of treating diabetics with human insulin produced by rDNA technology. It is possible to use rDNA technology to produce insulin with a slightly different structure from that of human insulin. The effect of the changed structure can then be investigated. The activities of equal quantities of two insulins, both produced by E. coli, were compared in healthy, non-diabetic subjects: • human insulin • insulin X, in which the positions of two amino acids, lysine and proline, were exchanged. Lysine has a hydrophilic R group and proline has a hydrophobic R group. The results of the investigation are shown in . activity of insulin / arbitrary units time after injection / hours insulin X human insulin With reference to describe the differences in activity between human insulin and insulin X. Suggest how exchanging the position of two amino acids in the insulin molecule can result in differences in activity.
9700_w14_qp_42
THEORY
2014
Paper 4, Variant 2
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The diagram outlines how a gene coding for human insulin is produced by genetic engineering techniques. mRNA for human insulin gene isolated step 1 step 2 X Y step 3 step 4 single strand of complementary DNA (cDNA) produced single-stranded DNA made into double-stranded DNA additional non-coding DNA added Name the enzymes X and Y. X Y Explain why the starting point in this procedure is mRNA. Suggest why some methods of manufacturing genetically engineered insulin use eukaryotic yeast cells rather than prokaryotic bacterial cells. State three advantages of using human insulin produced by genetic engineering.
9700_w14_qp_43
THEORY
2014
Paper 4, Variant 3
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Haemophilia A and haemophilia B are common hereditary disorders of blood clotting. Haemophilia A is a sex-linked genetic disorder that affects approximately 1 in 20 000 males worldwide. It is caused by a recessive allele of a gene coding for a clotting factor and results in excessive bleeding. There is currently no cure, but symptoms of haemophilia can be treated with a transfusion of a clotting factor to slow down the bleeding. State how genetic screening could reduce the number of cases of haemophilia. Some genetic disorders can be treated with gene therapy. Outline the aims of gene therapy. Suggest why haemophilia A is a suitable disorder for treatment with gene therapy. Haemophilia A and haemophilia B are caused by mutations in different blood clotting genes, F8 and F9 respectively. Both disorders have been treated with gene therapy involving the use of a vector. Table 4.1 shows the lengths, in kilobases , of the F8 and F9 genes. Table 4.1 haemophilia gene gene length / kb A F8 >8 B F9 1.4 With reference to Table 4.1, suggest why gene therapy using the F9 gene has been more successful than using the F8 gene. Two frequently used vectors in gene therapy are compared in Table 4.2. Table 4.2 feature vector adenovirus retrovirus genetic material of virus double-stranded DNA single-stranded RNA expression of inserted gene high gene expression gene expression in dividing cells only host immune response to virus high low With reference to Table 4.2, explain the advantages and disadvantages of using adenovirus rather than retrovirus as a vector.
9700_w15_qp_43
THEORY
2015
Paper 4, Variant 3
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Huntington’s disease is caused by a dominant allele of the gene that codes for the production of the huntingtin protein. This protein affects the development of many different tissues, including brain tissue. • The Huntington allele contains several repeats of the base sequence CAG, which codes for glutamine. • This results in a polyglutamine section in the synthesised protein. • A gene with more than 39 CAG repeats produces a protein that does not fold properly and does not function. • Symptoms of Huntington’s disease usually first appear between the ages of 30 and 45 years. • There is no treatment for the disease, which is progressive and always fatal. • Some people with between 27 and 35 CAG repeats do not develop the disease, but may still pass on the Huntington allele to their children, who may develop the disease as the number of repeats tends to increase when gametes are produced. The amino acid sequences on either side of the polyglutamine section of the huntingtin protein are not changed by the presence of the CAG repeats in the Huntington allele. Explain why this is so. With reference to the information given, explain why Huntington’s disease cannot be treated with gene therapy. Young people who have a parent with Huntington’s disease can choose to be screened for the presence of the Huntington allele. State the probability that a young person who has one parent with Huntington’s disease will inherit the Huntington allele. Suggest one advantage and one disadvantage of screening for Huntington’s disease before any symptoms occur. advantage disadvantage A couple, in which one partner has the Huntington allele, may choose to use IVF (in vitro fertilisation) to have a child. Any embryos obtained from the IVF procedure can be screened in the following way: • carry out an embryo biopsy • use PCR • test for the presence of the Huntington allele • only implant embryos that do not contain the Huntington allele. State what is meant by the term embryo biopsy. Explain why PCR is used in this procedure. Outline two social or ethical implications of screening embryos in this way.
9700_w17_qp_42
THEORY
2017
Paper 4, Variant 2
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Leber’s congenital amaurosis (LCA) is an autosomal recessive eye disease. LCA results in eye disorders, including severe loss of vision, at birth. LCA has been successfully treated by gene therapy, using a virus instead of a plasmid as the vector. Adeno‑associated virus (AAvectors containing the therapeutic allele were injected directly into the retina, the layer at the back of the eye containing the photoreceptor cells. People who had been blind from a young age were able to see again. There is a risk associated with the injection method used to deliver the vectors, as it might cause the retina to detach, damaging vision. This method of delivery was first used for LCA before being trialled on other retinal diseases that gradually reduce the vision of people as they get older. Suggest the main steps involved in creating recombinant DNA for this example of gene therapy. Explain why the fact that LCA is an autosomal recessive genetic disease makes it suitable for treatment with gene therapy. Suggest why the retinal injection method of gene therapy was used for LCA before it was trialled on other retinal diseases that gradually reduce the vision of people as they get older. Scientists tried to create an improved virus vector for gene therapy. step 1 – The scientists used a special form of the polymerase chain reaction (PCR). This form of PCR causes mutations in the DNA sequence of AAV by base substitution. step 2 – The viruses containing different base substitutions were tested. This was done by using the different viruses to deliver a new gene, the gene for green fluorescent protein (GFP), into the photoreceptor cells of mice, using the retinal injection method. step 3 – The best virus, known as 7m8, caused the photoreceptor cells in the retina of the mouse to fluoresce brightly, even when the recombinant virus was injected into the fluid inside the eye instead of into the retina itself. step 4 – The 7m8 virus was used to cure a mouse with LCA by injecting this virus containing the therapeutic allele into the fluid inside the eye of the mouse. Suggest how errors occurring during PCR can cause base substitution mutations in the DNA sequence of AAV. Explain why the photoreceptor cells of the mouse fluoresced in step 3. Predict the impact of the 7m8 AAV on treatment for age‑related retinal diseases.
9700_w18_qp_41
THEORY
2018
Paper 4, Variant 1
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Questions Discovered
37