Section 8: The control of gene expression Flashcards
1
Q
Mutations are changes in the sequence of nucleotides in DNA molecules. Name the four types of mutations:
A
- Insertion/Deletion
- Duplication
- Inversion
- Translocation
2
Q
Translocation is a type of mutation, describe how it occurs?
A
A group of bases become separated from the DNA sequence on one chromosome and are inserted into the DNA sequence on another chromosome.
3
Q
Describe how inversion ( a type of mutation ) occurs?
A
A group of bases become separating from the DNA sequence and then re-join at the same position but in the reverse order.
4
Q
What two type of mutations cause a frameshift
A
Insertion/deletion
Duplication
5
Q
Gene mutations can be cause by mutagenic agents that affect DNA, name 3 potential causes of gene mutations:
A
- Chemical mutagens - alcohol, benzene, tar
- Ionising radiation - UV and Xray
- Spontaneous errors in DNA replication
6
Q
Define Stem Cell
A
Undifferentiated cells which can keep dividing to give rise to other cell types
7
Q
What does pluripotent mean?
A
can form any cell type in the body, excluding placental cells. often used in replacing damaged tissues in human disorders
8
Q
What does totipotent mean?
A
A type of stem cell that can give rise to all types of specialised cells
9
Q
What does multipotent mean?
A
Can differentiate into other cells types but are more limited e.g. the cells in the bone marrow and umbilical cords
10
Q
What does unipotent mean?
A
Cells can only differentiate into one type of cell
11
Q
Name the four different types of stem cells.
A
- Totipotent
- Pluripotent
- Multipotent
- Unipotent
12
Q
Where do stem cells originate from in mammals?
A
- Embryonic stem cells
- Umbilical cord blood stem cells
- Placental stem cells
- Adult stem cells (some are in body tissues to maintain and repair tissues)
13
Q
What does iPS stand for?
A
induced pluripotent stem cells
14
Q
Describe how induced pluripotent stem (iPS) cells are generated.
(4 marks)
A
Somatic cells (e.g., skin cells) are collected from an individual.
Four specific genes (Oct4, Sox2, Klf4, and c-Myc) are introduced into these somatic cells.
These genes reprogram the somatic cells, turning them into pluripotent cells (iPS cells).
iPS cells are capable of developing into a wide variety of cell types, similar to embryonic stem cells.
15
Q
Explain two potential medical applications of iPS cells.
(4 marks)
A
Cell therapy: iPS cells can be used to generate tissues or organs for transplant, reducing the risk of rejection.
Disease modelling: iPS cells can be used to create models of diseases (e.g., Alzheimer’s, Parkinson’s) for drug testing and research.
16
Q
What are the advantages of using induced pluripotent stem (iPS) cells?
A
Lower ethical concerns: iPS cells are derived from adult somatic cells (e.g., skin cells), avoiding the destruction of embryos.
No embryo destruction: There is no involvement of embryos, avoiding debates about the moral status of embryos.
Reduced risk of rejection: Since the cells are genetically similar to the donor, there are fewer concerns about immune rejection and tumour formation.
17
Q
What are the ethical concerns associated with the use of embryonic stem cells?
A
Higher ethical concerns: Embryonic stem cells are derived from human embryos, involving the destruction of the embryo.
Moral status of embryos: The use of embryos raises debates about when human life begins and whether embryos should be considered for research.
Exploitation risks: The process may involve the exploitation of women for egg donation and potential commodification of human life.
18
Q
Describe the action of oestrogen in controlling transcription in 3 points.
A
- Oestrogen diffuses across membrane binds to receptor molecule of transcription factor
- Alters shape of DNA binding site on transcription factor
- Transcription factor therefore enters nucleus via nuclear pore and binds to DNA
19
Q
What effect foes oestrogen have on transcription?
A
Stimulates transcription of the gene that makes up DNA (does this by altering DNA binding site on transcription factor so it can bind to DNA)
20
Q
How does oestrogen enter a cell?
A
Oestrogen is lipid soluble so it can diffuse freely across the phospholipid bilayer
21
Q
What is the role of transcriptional factors in gene expression?
A
Transcriptional factors are proteins that bind to specific DNA sequences to regulate the transcription of genes.
They can either activate or repress gene expression by promoting or inhibiting the binding of RNA polymerase to the DNA.
Activators increase transcription by helping RNA polymerase bind, while repressors decrease transcription by blocking RNA polymerase or altering the chromatin structure.
22
Q
What is the role of siRNA in gene silencing?
A
siRNA binds to a complementary sequence of mRNA.
siRNA is usually single stranded and the cell therefore detects the double stranded from on mRNA and view it as abnormal.
Therefore the mRNA is broken down by enzymes preventing translation
23
Q
What are two applications of siRNA in research and medicine?
A
Gene function analysis:
siRNA is used to silence specific genes in research to study their function by knocking them down at the mRNA level.
Therapeutic use: siRNA can be designed to target and silence disease-causing genes, such as in viral infections or cancer, offering potential treatments by reducing harmful gene expression
24
Q
What is epigenetics?
A
Epigenetics is the study of changes in gene expression or phenotype that do not involve changes to the DNA sequence itself.
25
How does DNA methylation affect gene expression?
DNA methylation involves adding a methyl group (-CH3) to the DNA, typically at cytosine bases.
Methylation of genes silences their expression by preventing the binding of transcription factors or RNA polymerase.
This often leads to a decrease in gene activity and can be inherited through cell divisions, affecting gene expression over generations.
26
How does DNA acetylation affect gene expression?
DNA acetylation involves the addition of an acetyl group (-COCH3) to histone proteins.
Acetylation of histones reduces the positive charge on the histones, causing them to loosen their grip on DNA.
This relaxation of DNA packaging allows transcription factors and RNA polymerase to bind more easily, promoting gene expression
27
How do histone modifications influence gene expression?
Histones are proteins that DNA wraps around, and their modification can influence gene expression.
Acetylation of histones (addition of an acetyl group) loosens DNA packaging, allowing genes to be more easily transcribed.
Methylation of histones can either activate or repress gene expression depending on the specific site of modification.
28
What is the effect of decreased DNA acetylation?
Decreased acetylation of histones increases there positive charge, so they bind to DNA more tightly. When this happens transcription factors can no longer access the DNA, so the gene is switched off.
29
Why do histones associate with DNA?
Histones are positively charged proteins that interact with the negatively charged DNA molecules.
This electrostatic attraction between the positive charge of histones and the negative charge of the phosphate groups in DNA helps DNA to coil tightly around the histones, forming nucleosomes.
30
Define cancer.
Cancer is the uncontrolled cell division caused by mutations in genes that regulate the cell cycle.
This leads to the formation of tumours and, in some cases, the spread of cancer cells to other parts of the body (metastasis)
31
What is metastasis?
Metastasis is the spread of cancer cells from the original (primary) tumor to other parts of the body, forming secondary tumors.
Cancer cells break away, travel through the blood or lymphatic system, and invade other tissues.
32
What are the differences between benign and malignant tumours?
Benign tumours consist of specialised cells and are usually encapsulated, meaning they do not invade surrounding tissues. They grow slowly and do not spread.
Malignant tumours contain unspecialised cells, are not encapsulated, and invade surrounding tissues. They can spread to other parts of the body through metastasis and often evade the body's defensive tissues
33
What are the two main types of genes that play a role in cancer?
Tumour suppressor genes and oncogenes
34
What role does proto-oncogenes play in cancer?
If a proto-oncogene mutates causing activation of an oncogene which causes:
Receptor protein on cell-surface membrane to be permeantly activated so that cell division is switched on even in the absence of growth factors.
The oncogene may code for a growth factor that is then produced in excessive amounts of
35
How are oncogenes formed?
They are formed from mutated proto-oncogenes.
36
What is the result of the formation of an oncogene? How does it do this?
Result in uncontrolled cell division.
It does this by permanently activating a cell surface receptor or coding for a growth factor
37
How do tumour suppressor genes control cell division?
cause the cell cycle to stop when damage is detected.
Also play important role in the programming of apoptosis (cell death). When switched off the cell cycle becomes unregulate.
38
What is apoptosis?
Cell death
39
How does abnormal methylation of tumour suppressor genes and oncogenes contribute to cancer?
Abnormal methylation of tumor suppressor genes (e.g., hypermethylation) can silence these genes, preventing them from inhibiting cell division and allowing cells to divide uncontrollably.
Abnormal methylation of oncogenes (e.g., hypomethylation) can activate these genes, leading to overexpression and excessive cell growth.
Both processes disrupt the normal regulation of the cell cycle and contribute to the development of cancer.
40
Why can increased oestrogen concentrations be linked to breast cancer?
Because they bind to transcription factors which activate genes promoting cell division, potentially leading to tumour formations. Elevated levels are usually found in fatty tissues call adipose tissue.
41
What is in vivo cloning?
In vivo cloning is a method of gene cloning that involves inserting a gene into a living organism (such as a bacterium) where it is replicated as the organism divides
42
What does In vitro mean?
“In glass”
DNA (containing gene) is copied many times over by Polymerase Chain Reaction using PCR machine
43
What is a vector?
A length of DNA that carries the gene of interest into the host cell.
It has to be big enough to contain the gene
44
Explain the role of a vector during in vivo gene cloning.
A vector is used to transfer the desired gene (DNA fragment) into a host cell
The vector ensures the foreign DNA is replicated when the host cell divides
It may also carry marker genes to allow identification of successfully transformed cells
45
What could be used as a vector?
Plasmids
Bacteriophages (viruses that infect bacteria)
46
What is a plasmid? Why is it commonly used as a vector?
A plasmid is a small, circular DNA molecule found in bacteria. It’s used as a vector because it can replicate independently and carry foreign genes.
47
Name the group of enzymes that cut DNA
Restriction enzymes ( also known as restriction endonucleases)
48
Name the enzyme that joint nucleotides in a DNA strand.
DNA ligase
49
What are sticky ends?
• Sticky ends are exposed, unpaired bases at the ends of a DNA fragment.
• They are produced when DNA is cut by restriction enzymes in a staggered manner.
• Sticky ends are complementary to other sticky ends cut by the same enzyme, allowing DNA fragments to join.
50
Why is a promoter region added to the DNA fragment?
• The promoter region allows RNA polymerase to bind.
• This ensures the target gene is transcribed and expressed in the host cell.
51
Why is a terminator region added to the DNA fragment?
• The terminator region signals RNA polymerase to stop transcription.
• This ensures the correct length of mRNA is produced.
52
Give two ways a gene can be isolated from a piece of DNA. Name the enzymes involved
• Using restriction enzymes to cut the desired gene from the DNA at specific recognition sites.
• Using reverse transcriptase to produce complementary DNA (cDNA) from mRNA.
53
What does palindromic sequence mean?
• A palindromic sequence is a sequence of DNA bases that reads the same in both directions on complementary strands.
• For example:
• 5’ — GAATTC — 3’
• 3’ — CTTAAG — 5’
54
What is the name of the type of DNA formed from vector DNA and inserted DNA fragment?
Recombinant DNA
55
What is meant by the process ‘transformation’?
Transformation is the process of introducing recombinant DNA (such as a plasmid) into a host cell.
56
What is a genetic marker?
• A genetic marker is a gene that produces an observable characteristic, allowing scientists to identify cells that have successfully taken up the recombinant DNA.
• Examples include antibiotic resistance genes or fluorescent protein genes.
57
How can bacteriophages act as vectors?
Bacteriophages are viruses that infect bacteria; they can inject foreign DNA directly into bacterial cells.
58
What role do restriction enzymes play in vector preparation?
Restriction enzymes cut DNA at specific sequences, creating sticky or blunt ends that allow foreign genes to be inserted into the vector.
59
What is a recombinant plasmid?
A recombinant plasmid is a plasmid that has had a foreign gene inserted into it.
60
Describe the steps involved in inserting a gene into a plasmid vector.
The plasmid and target gene are cut with the same restriction enzyme to create complementary sticky ends. DNA ligase then joins the gene to the plasmid, forming a recombinant plasmid.
61
How are sticky ends important in forming recombinant DNA?
Sticky ends are complementary overhanging sequences that allow DNA fragments to anneal and form stable recombinant molecules.
62
How is a vector introduced into a host cell?
Through transformation, often by heat shock or electroporation.
63
What is transformation in the context of genetic engineering?
Transformation is the process of introducing a recombinant vector into a host cell.
64
Describe the process of identifying cells that have successfully taken up the vector.
Cells are grown on agar plates containing antibiotics or other selective agents. Only transformed cells with resistance genes (from the vector) will survive.
65
Why are marker genes important in vector-based genetic engineering?
Marker genes help identify cells that have successfully taken up the vector.
66
What types of marker genes are commonly used in recombinant DNA technology?
Antibiotic resistance genes or genes that produce fluorescent proteins.
67
How are antibiotic resistance genes used in vector identification?
Cells that have taken up the vector will survive on agar containing the corresponding antibiotic.
68
Explain how fluorescent markers can indicate successful gene insertion.
Fluorescent marker genes cause transformed cells to glow under UV light, indicating successful gene uptake.
69
What is the purpose of inserting a promoter sequence into a vector?
A promoter ensures the inserted gene is transcribed and expressed in the host cell.
70
Why might some cells fail to take up a recombinant plasmid during transformation?
Factors include inefficient heat shock conditions, damaged plasmids, or natural resistance of cells to transformation.
71
How can gene expression be controlled once a vector is introduced into a host?
By using inducible promoters that activate gene expression in response to specific environmental factors.
72
What are the ethical concerns surrounding the use of vectors in genetic modification?
Concerns include potential environmental impacts, gene escape, and the modification of human genes.
73
What are the 5 overview steps of gene cloning?
1. Isolation (of DNA fragments)
2. Insertion
3. Transformation
4. Selection
5. Culturing
74
Name the three methods to create fragments of DNA.
1. Reverse transcription
2. Restriction endonucleases
3. Gene machine
75
Describe the process of using reverse transcriptase to produce DNA fragments.
1. Enzyme makes DNA copies from mRNA
2. Naturally occurs in viruses, such as HIV
3. A cell that naturally produces the protein of interst is selected
4. These cells should have large amounts of mRNA for the protein
5. The reverse transcriptase enzyme joins the DNA nucleotides with complementary bases to the mRNA sequence
6. Single stranded DNA is made (cDNA)
7. To make this DNA fragment double stranded, the enzyme DNA polymerase is used
76
What is the advantage of using reverse transcription to produce DNA fragments?
The cDNA is intron free because it is based on mRNA template.
Prokaryotic cells do not have the ability to remove introns so very useful if using those
77
Describe how restriction endonucleases produce DNA fragments?
• Restriction endonucleases are enzymes that cut DNA at specific sequences called recognition sites.
• These recognition sites are palindromic sequences, meaning they read the same in opposite directions on complementary strands.
• The enzyme cuts the sugar-phosphate backbone of the DNA at these sites.
• This cut can produce either:
• Sticky ends — staggered cuts that leave exposed, unpaired bases at each end. These are useful for joining DNA fragments with complementary sticky ends.
• Blunt ends — straight cuts that leave no exposed bases.
78
What are sticky ends?
staggered cuts that leave exposed, unpaired bases at each end. These are useful for joining DNA fragments with complementary sticky ends.
79
80
What are blunt ends?
straight cuts that leave no exposed bases.
81
What is the gene machine?
The gene machine is a method of synthesising DNA fragments without the need for a pre-existing DNA template.
82
What is the starting point for synthesising a gene using the gene machine?
The desired gene sequence is designed on a computer.
83
How does the gene machine assemble the DNA fragment?
The computer designs small, overlapping sections of DNA called oligonucleotides, which are chemically assembled and joined together to form the complete DNA sequence.
84
What happens after the oligonucleotides are assembled?
The assembled DNA sequence is checked for errors and replicated using the polymerase chain reaction (PCR).
85
What is added to the synthetic DNA to allow it to be inserted into a vector?
Sticky ends are often added to enable the DNA fragment to be inserted into a plasmid or other vector.
86
Name an advantage and disadvantage of using the gene machine to produce DNA fragments.
Advantage: Can desing exact DNA fragment you want, with sticky ends, labels and preferential codons
Disadvantage: Need to know the sequence of amino-acids or bases
87
Why is the removal of introns important in gene machine synthesis?
Prokaryotic cells cannot splice out introns, so producing intron-free DNA ensures successful gene expression.
88
What are some applications of the gene machine?
• Producing genes for medical research.
• Creating genes for genetically modified organisms (GMOs).
• Developing synthetic biology solutions.
89
Name an advantage and disadvantage of using reverse transcriptase to produce DNA fragments.
Advantage: mRNA present in cell is from actively transcribed genes, so lots of the mRNA of interest available to make cDNA
Disadvantage: More steps so more time consuming and technically more difficult
90
Name an advantage and disadvantage of using Restriction endonuclease to produce DNA fragments.
Advantage: Sticky ends on DNA fragment make it easier to insert to make recombinant DNA
Disadvantage: Still contains introns
91
Give two epigenetic mechanisms used to prevent transcription
Increased methylation of DNA
Decreased acetylation of histones
92
What name is used for the non-coding sections of a gene?
Introns
93
Name one type of gene that can prevent tumour growth
Tumour suppressor gene
94
During which part of the cell cycle are gene mutations most likely to occur? Suggest why.
Interphase and S phase
Interphase is when DNA replication happens most chance of spontaneous mutation
95
96
A translocation mutation is, in effect, a combination of two other different types of gene mutation. Deduce which two types of mutation these are and explain your answer.
Deletion and addition because the bases are deleted from one chromosome and added to a different one.
97
Explain why the effects of a single adfdtional base in a sequence of DNA bases may have:
a) a considerable effect on the polypeptide produced
b) little effect on the polypeptide produced
a) It will cause a frame shift causing triplets (codons) in a sequence to be read differently because each has been shifted to the right by one base. If the additional base is inserted early in the sequence most codons will be changes, so will the amino acids they code for. The resultant polypeptide will be very different from normal.
b) If the additional base is inserted at the end of the sequence few, if any codons will be changed. Few, if any, amino acids they code for will differ and the resultant polypeptide will be normal or near normal.
98
A mutation causes three bases in the DNA of a gene to become duplicated. Explain how the effects of this mutation might differ if the duplicated bases are consecutive rather than in three separate locations on the DNA molecule.
Where the duplicated bases are consecutive, the frame shift is three bases long and so the subsequent codons are not affected. The polypeptide will have an additional amino acid but otherwise be unchanged. If the bases are separate, the frame shift will initially be one base long, becoming two bases long after the second duplicate base is added. Codons after both the duplications will be changed and the polypeptide will have many different amino acids (but not necessarily all - degenerate code). After the third duplicate base the codons will be unchanged.
99
Suggest two reasons why the addition of a single base into a DNA sequence may not alter the amino acid sequence in the resultant polypeptide.
Some codons will be changed to ones that code for the same amino acids (degenerate code).
The frame shift might not alter some codons because the replacement bases are the same as the originals.
100
What is a steroid hormone? Name an example.
A steroid hormone is a lipid-soluble hormone made from cholesterol that passes through cell membranes and affects gene expression by binding to receptors inside the cell.
Oestrogen, Testosterone, ect.
Don’t need to know it directly just that these work differently to protein hormones like insulin
101
Once a transcription factor has bound to the promoter region, what does this allow to happen next?
The enzyme RNA polymerase to bind to the DNA and transcribe the gene to produce mRNA.
102
If the protein or polypeptide is not required by the cell, what does the transcription factor do?
Detaches from the DNA which causes transcription to stop.
103
Not all point mutations in DNA result in a change of one amino acid in a polypeptide. Give two reasons why?
Degeneracy of code / explained re. mutation may code for same amino acid ;
Mutation may be in non-coding DNA / in an intron ;
Mutation may give stop signal ( truncated polypeptide) ;
Mutation may cause a frame shift / described ;
104
During which part of the cell cycle are gene mutations most likely to occur? Suggest an explanation for your answer. 2 marks
(i) Interphase / S / synthesis (phase);
(ii) DNA / gene replication / synthesis occurs / longest stage;
Allow ‘genetic information’ = DNA.
Allow ‘copied’ or ‘formed’ = replication / synthesis
105
Suggest how a mutation can lead to the production of a protein that has one amino acid missing. 2 marks
Loss of 3 bases / triplet = 2 marks;;
‘Stop codon / code formed’ = 1 mark max unless
related to the last amino acid
Loss of base(s) = 1 mark;
eg triplet for last amino acid is changed to a stop
codon / code = 2 marks
3 bases / triplet forms an intron = 2 marks
Accept: descriptions for ‘intron’ eg non-coding DNA
‘Loss of codon’ = 2 mark
106
Suggest how the production of a protein with one amino acid missing may lead to a genetic disorder.
Change in tertiary structure / active site;
Neutral: change in 3D shape / structure
(So) faulty / non-functional protein / enzyme;
Accept: reference to examples of loss of function eg
fewer E-S complexes formed
107
Give two characteristic features of stem cells.
Will replace themselves / keep dividing / replicate;
Undifferentiated / can differentiate / develop into other cells / totipotent / multipotent / pluripotent;
Accept tissues
108
Suggest one way that putting Embryonic pluripotent stem cells (ESCs) into a persons heart might lead to more harm to the person.
Might divide out of control, leading to tumour/cancer
May differentiate into the wrong type of cells
109
Suggest how the growth of new blood vessels into damaged heart tissues could increase the rate of repair of tissues. 3 marks
Greater blood supply (to damaged areas);
Bringing more oxygen / glucose for respiration;
Brings more amino acids for protein synthesis;
For cell repair / mitosis / division;
110
The scientist used an optical microscope to measure the number of capillaries in thin sections cut from samples of heart muscle.
Describe the method they would have used to fine the mean number of capillaries per mmsquared. 4 marks
Measure diameter of field of view and calculate area;
Using micrometer slide and eyepiece graticule;
Accept descriptions
Count number of capillaries in large number of fields of view and calculate mean;
Select fields of view randomly
111
Papaya plants reproduce sexually by means of seeds. Papaya plants grown from seeds are very variable in their yeild. Explain why. 2 marks
Meiosis / independent assortment / crossing over;
(Fusion of) genetically different gametes / random fertilisation;
112
Explain the advantage of growing a plant that reproduces sexually from tissue culture rather than seeds. 1 mark
Will be clones / produced by mitosis / will be genetically identical / less variation / all plants will have desired characteristics;
If the reference is to identical must be genetically identical,
but allow less variation without the reference to genetical.
113
A male has a sex-linked mutation, where would it be?
Non-homologous section of an X chromosome
114
Define heterochromatin
Heterochromatin is a tightly packed form of DNA that is not usually transcribed because the genes are inaccessible to transcription machinery.
115
What is Euchromatin?
Euchromatin is a loosely packed form of DNA where genes are accessible and can be actively transcribed.
116
When the gene is active and transcriptional factors have access to the DNA, what is the chromatin type?
Ehchromatin
117
What does decreased acetylation of the histones and increased methylation of the DNA do?
DNA-histone complex because more condensed (tightly packed)
Chromatin type is heterochromatin
Transcriptional factors have no access
The gene is inactive
118
Give two epigenetic mechanisms used to prevent transcription.
Increased methylation of DNA
Decreased acetylation of histones
119
What is histone acetylation?
Adding an acetyl group to histone proteins, changing heterochromatin to euchromatin
120
What is DNA methylation?
When a methyl group is added to the cytosines bases of DNA
This prevents transcriptional factors from binding to the DNA and triggers histone de acetylation. which attracts proteins that condense the DNA-histone complex making the DNA inaccessible to transcription factors.
Transcription cant take place
121
What is the epigenome?
The epigenome is the collection of chemical tags on DNA and histones that control gene expression without changing the DNA sequence.
122
Explain how an activated oestrogen receptors affects the target cell. 2 marks
(Receptor / transcription factor) binds to promoter which
stimulates RNA polymerase / enzyme X;
Transcribes gene / increase transcription;
123
Oestrogen only affects target cells. explain
Other cells do not have the / oestrogen / ERα receptors;
124
Transcriptional factors are important in the synthesis of particular proteins. Describe how.
Bind to DNA / gene;
Generally attaching to DNA
At specific region / base sequence / promoter sequence;
At specific place
Stimulate transcription / prevents transcription / turn on gene /
turn off gene;
Accept description of transcription.
Do not accept protein synthesis.
125
What do RNAi and siRNA stand for?
RNA interference
small interfering RNA
126
How does the mechanism involving siRNA inhibit translation of mRNA?
1. Enzyme cuts large double-stranded molecules of RNA into smaller sections called siRNA
2. One of the two strands combines with an enzyme
3. The siRNA strand pairs with complementary bases on a mRNA strand
4. The enzyme cuts the mRNA into smaller sections
5. This means the mRNA is no longer capable of enough translated and the gene has no been expressed/ has been blocked
127
Where does siRNA come from?
When an enzyme cuts large double-stranded molecules of RNA into smaller sections. This happens in the cytoplasm.
128
Compare benign and malignant tumours.
Benign grow very slowly Malignant grow rapidly
Benign are often specialised cells Malignant become unspecialised
Benign adhere together so don’t from secondary tumours Malignant spread in a process called metastasis forming secondary tumours
Benign tumours are surrounded by a capsule Malignant are not surrounded by a capsule
129
Name one type of gene that can prevent tumour growth.
Tumour suppressor gene
130
Describe a process by which oestrogen might cause breast cancer in post-menopausal women.
After menopause, fat cells in breast tissue produce more oestrogen.
Oestrogen can act as a transcription factor by binding to specific receptors in the cytoplasm of breast cells.
This oestrogen–receptor complex enters the nucleus and binds to specific genes, including proto-oncogenes.
This can increase the transcription of proto-oncogenes, converting them into oncogenes.
Oncogenes produce proteins that stimulate cell division.
As a result, the rate of mitosis increases, which can lead to uncontrolled cell division and the formation of a tumour, potentially causing breast cancer.
131
Explain why the activation of a proto-oncogenes can cause the development of a tumour while it requires deactivation of a tumour suppressor gene to do so.
Proto-oncogenes increase the rate of cell division and so their activation produces a mass of cells but tumour suppressor gens descrease the rate of cell diction and so their deactivation produces a tumour.
132
Suggest two reasons why the surgical removal of a benign tumour is usually sufficient treatment to prevent the tumour growing again.
Produce adhesion molecules that make them stick together and are surrounded by a capsule of dense tissue.
The surgical removal is likely to remove all tumour cells.
133
Suggest why the surgical removal of a malignant tumour requires follow-up treatment.
Malignant tumours spread to other regions of the body and so even though sure grey can remove the obvious larger ones, tiny ones will require other therapies to prevent these regrowing into new tumours.
134
What is a proto-oncogene?
A proto-oncogene is a gene that normally codes for proteins involved in the stimulation of cell division, growth, and differentiation.
135
What role does proto-oncogenes have in cancer?
If a proto-oncogene mutates causing activation of an oncogene which causes:
Receptor protein on cell-surface membrane to be permeantly activated so that cell division is switched on even in the absence of growth factors.
The oncogene may code for a growth factor that is then produced in excessive amounts
136
What is aptosis and what gene initiates it?
Aptosis is programmed cell death
Stimulated by tumour suppressor genes
137
How can hyper methylation lead to cancer?
Hypermethylation occurs in a promoter region of tumour suppressor genes
This leads the the tumour suppressor gene being inactivated
As a result transcription of the promoter regions of tumour suppressor genes inhibited
The tumour suppressor gene is therefore silenced
Tumour suppressor gene normally slows the rate of cell division its inactivation leads to increased cell division and the formation of a tumour.
138
How can hypomethylation lead to cancer?
Reduced methylation found to occur in oncogenes where it leads their activation and hence formation of tumours.
139
140
Why is determining the proteome of prokaryotic organisms like bacteria relatively easy?
One circular piece of DNA not assosiated with histones
No none-coding portions of DNA
141
Explain how knowledge of the proteome of a pathogen might be able to help control the disease it causes.
It allows identification of those proteins that act as antigens on the surfaces of the pathogens. These act as antigens on the surfaces of the pathogens. These antigens can then be used to produce effective vaccines against the disease.
142
What is a stem cell?
Undifferentiated cells that can continually divide and become specialised.
143
What does iPS cells stand for?
Induced pluripotent stem cells
144
Define recombinant DNA.
DNA that has been altered by incorporating fragments of DNA from a different source
145
What are the three methods of producing DNA fragments.
Conversion of mRNA to cDNA using reverse transcriptase
Using restriction endonucleases to cut fragments containing the desired genes from DNA
Creating the gene in a gene machine (usually based on a known protein structure)
146
What does the enzyme reverse transcriptase do?
catalyses tthe production of DNA from RNA
147
Describe how reverse transcriptase is used to isolate a gene.
1. mRNA Extraction: Identify and extract mRNA from cells expressing the target gene.
2. cDNA Synthesis: Use reverse transcriptase to synthesize a complementary DNA strand from the mRNA template.
3. Double-Stranded DNA Formation: Utilize DNA polymerase to form the complementary strand, resulting in double-stranded cDNA.
4. Gene Isolation: The cDNA now represents the isolated gene, free from introns, suitable for cloning or further analysis.
148
What is cDNA?
cDNA stands for complementary DNA. It’s a form of DNA that is synthesized from messenger RNA (mRNA) using the enzyme reverse transcriptase.
149
How is cDNA formed?
1. mRNA is extracted from a cell.
2. Reverse transcriptase builds a single-stranded cDNA from the mRNA template.
3. DNA polymerase makes it double-stranded.
150
Some bacteria defend themselves by producing enzymes that cut up viral DNA. What are these enzymes called?
Restriction endonulceases
151
What is a palindrome?
A palindromic DNA sequence is a sequence of base pairs that reads the same on both strands.
152
What are the advantages of using the gene machine to obtain a gene opposed to other methods like using restriction endonuclease or reverse transcriptase?
• No need for mRNA or a donor organism.
• Introns and non-coding DNA are not included (ideal for. prokaryotic expression).
• Genes can be optimized for expression in different organisms.
• Fast and accurate.
153
Describe the steps in gene machine synthesis.
1. Desired nucleotide fed into a computer
2. Synthesis of oligonucleotides
3. Assembly of gene (olgionucleotides are overlapped then joined together and made double stranded using the PCR)
4. Gene cloning (gene inserted into a bacterial plasmid)
5. Genes are sequenced and those with errors are rejected
6. Gene is usually delivered incorporated into plasmid
154
Sticky ends are produced when DNA is cut by…
Restriction endonuclease
155
What are the sequences of DNA that are cut by restriction endonuclease called?
Recognition sites
156
What enzyme binds complementary bases of two sticky ends?
DNA ligase
157
What is the most commonly used vector?
plasmids
158
Describe the steps of In Vivo Cloning.
1. Isolate the gene (methods previously learnt ie reverse transcriptase, restriction enzymes or gene machine)
2. Prepare the vector (use the same restriction enzyme to cut a plasmid creating complementary sticky ends)
3. Insert the gene ( Mix gene + plasmid + DNA ligase enzyme forming recombinant DNA )
4. Introduce vector into host (transformation)
5. Identify successful uptake (use marker genes)
6. Clone the gene
159
Whate are the advantages of In Vivo cloning.
• Useful when introducing genes into organisms.
• Allows for expression of the gene → protein production.
• No risk of mutation during synthesis (unlike in vitro PCR).
• Enables use of control elements (promoters, terminators).
160
What three marker genes can you use?
Antibiotic-resistance marker genes
Fluorescent markers
Enzyme markers
161
What is in vitro cloning?
It’s the process of cloning DNA outside of a living organism, in a test tube, using the Polymerase Chain Reaction (PCR) to rapidly make millions of copies of a DNA fragment.
162
Describe the polymerase chain reaction.
1. Reaction mixture set up (DNA fragment, primers, DNA polymerase, free nucleotides, buffer solution)
2. Heat to 95degrees - Denaturation (h bonds break DNA strands separate)
3. Cool to 55 degrees - Annealing (primers bind to complementary strands)
4. Synthesis of DNA (Heat to 72 degrees DNA polymerase adds nucleotides forming new strands starting from the primers)
163
Advantages of In Vitro cloning (PCR)
• Rapid (faster than in vivo).
• Only need a small amount of DNA to start.
• Doesn’t require living cells.
164
Disadvantages of In vitro cloning (PCR)
• High risk of contamination.
• DNA polymerase can introduce errors (mutations).
• Can’t produce proteins, only copies DNA.
• Needs prior knowledge of DNA sequence (to design primers).
165
What is a DNA probe?
Short single-stranded length of DNA that has some sort of label attached that makes it easily identifiable
166
During In viva cloning the DNA fragments mus the modified to ensure transcription of these genes can occur. What is this modification?
A promoter region must be added - added at hte start of the DNa fragment. This is a sequence of DNA which is the binding site for RNA polymerase to enable transcription to occur.
A terminator region must be added - This is added at the end of the gene. It causes RNA polymerase to detach and stop transcription, so only one gene at a time is copied into mRNA.
167
How do you insert DNA into a vector?
The plasmid is cut open using the same restriction endonuclease
This created the same sticky ends
Therefore, the DNA fragments sticky ends (exposed nucleotides) are complementary to the sticky needs on the plasmid
168
What does PCR stand for?
Polymerase chain reaction
169
Where are taq polymerase found and why are they used over DNA polymerase?
taq polymerase found in bacteria that
170
All cells possess the same genes and yet some cells cannot produce the one protein but they can produce another. Explain why.
If the gene that codes for a certain protein is not expressed (the site on the transcriptional factor that binds to DNA is not active/gene is switched off) than the genetic code for the protein will not be translated. Thus the cell cannot produce that protein.
171
Suggest a reason why skin cells retain an ability to divide by being unipotent when the cells of some other organs do not.
Skin cells, being on the outside of the body, are subjected to much wear and tear so need replacing frequently. Many other organs are less prone to damage and need little cell replacement.
172
How is translation of a polypeptide inhibited? Four steps
An enzyme can cut the mRNA into siRNA.
One strand of the siRNA then combines with another enzyme.
This siRNA-enzyme complex will bind via complementary base pairing to another mRNA molecule.
Once bound the enzyme will cut up the mRNA so it cannot be translated.
173
What are DNA probes?
Short single-stranded pieces of DNA complementary in base sequence to the VNTRs. The probes are labelled radioactively or fluorescently so they can be identified.
This is used to locate specific alleles of genes and to screen patients for heritable conditions, drug responses or health risks.
174
What are the main stages of genetic fingerprinting?
Collection
Extraction
Digestion
Separation
(southern blotting)
Hybridisation
Development
Analysis
175
What method is used to create DNA fragments for genetic fingerprinting and why?
Restriction endonuclease which cuts DNA at a specific sequence. (sticky or staggered ends)
176
How are the DNA fragments separated before DNA hybridisation in genetic fingerprinting?
Seperated based on size using a process called gel electrophoresis
Electric current is applied along the gel the negatively charged DNA is attracted to the positive end while the shorter DNA fragments move faster than the longer fragments
177
What are variable number tandem repeats (VNTRs)?
Found in the introns of DNA (non-coding regions of DNA)
178
What sections of DNA are analysed in genetic fingerprinting and why?
VNTR’s (variable number tandem repeats)
The probability of two individuals having the same VNTRs is very low, however the more closely related you are the more similar you VNTRs are.
179
Describe the digestion step in the genetic fingerprinting process?
Restriction endonucleases are added to cut the DNA into smaller fragments.
Enzymes which cut close to the VNTR’s are added.
180
What is southern blotting?
An alkaline is added to separate the double strands of DNA
A nylons membrane is placed over electrophoresis gel
Absorbent paper which draws up liquid containing DNA
Fixed using UV light
181
Describe the Hybridisation step in genetic fingerprinting.
Radioactive gene probes are put on to the nylon membrane
The probes are incubated (heated then temperature reduces) so that they’ll attach to any complimentary bits of DNA
182
What is the development and Analysis stage of Genetic fingerprinting?
The nylon sheet can then be exposed to x-rays to visualise the position of radioactive gene probes, or UV light if fluorescence probes were used.
The pattern of fragment distribution is then analysed
183
What are the uses of genetic fingerprinting?
Forensic science to please suspects at crime scenes
For medical diagnosis
To ensure animal and plants are not closely related before being bred
184
Describe DNA hybridisation.
The patients DNA samples are loaded is heated to make it single-stranded (hydrogen bonds break)
The patients single stranded DNA sample is mixed with the DNA probe, and cooled to and any complementary sequences can align and form hydrogen bonds (anneal)
Some of the patients DNA samples will anneal back together and some will with the DNA probe.
185
How do you locate specific alleles of genes?
Create the DNA (based on the known DNA base sequence on a specific allele)
The fragment of DNA can be produced using a gene machine
The fragment can be amplified using PCR
The label is then added (either radioactive nucleotide isotope P32 or a fluorescent label with emits light under UV light
After hybridisation the DNA is washed so that any unbound DNA probes are washed away
The presences of radioactive fluorescence label therefore indicated that the allele of interest is present in the patients DNA
186
Name two techniques the scientists may have used when analysing viral DNA to determine that the viruses were closely related.
For one mark, accept any two of the following:
• The polymerase chain reaction
• Genetic/DNA fingerprinting
• (Gel) electrophoresis
• DNA/genome sequencing;
Accept PCR for polymerase chain reaction
Accept autoradiography
Accept DNA hybridisation
Accept compare DNA/base sequence for ‘DNA
sequencing’
Ignore compare mRNA base sequence
Ignore compare amino acid sequence
Ignore DNA probes
187
Determining the genome of the viruses could allow scientists to develop a vaccine.
(The scientists) could identify proteins (that derive from the genetic code)
OR
(The scientists) could identify the proteome;
(They) could (then) identify potential antigens (to use in the vaccine);
Reject if answer suggests vaccine contains antibodies
188
Steroid hormones are hydrophobic. Explain why steroid hormones can rapidly enter a cell by passing through its cell-surface membrane. 2 marks
Lipid soluble
Diffuses through phospholipid bi layer
(if something is hydrophobic it is generally considered to be lipid soluble)
189
Suggest and explained how a protein could stimulate gene expression. 2 marks
Protein is transcriptional factor
Bind to DNA/promoter
Stimulates RNA polymerase
190
Define what is meant by epigenetics. 2 marks
Heritable changes in gene function
Without changes to the base sequence of DNA
191
What does acetyl bind to?
Protein histones
192
Explain how increased methylation could lead to cancer. 3 marks
Methyl groups added to a tumour suppressor gene
THe transcription of tumour suppressor genes is inhibited
Leading to uncontrolled cell division
