Topic 8: Controlling gene expression Flashcards
1
Q
TRUE OR FALSE
ALL mutations are bad for organisms
A
FALSE
Not ALL mutations are bad. Sometimes they result in variation which allows for selection
2
Q
What is a factor that increases the mutation rate called?
A
A mutagenic agent
3
Q
What are the 6 main types of mutation?
A
- Translocation
- Inversion
- Duplication
- Deletion
- Addition
- Substitution
4
Q
What are the 3 types of substitution mutation?
A
TYPES OF SUBSTITUTION
- Nonsense mutation (codes for stop codon)
- Mis-sense mutation (codes for a different AA)
- Silent mutation (same AA is coded for)
5
Q
What is a translocation mutation?
A
When a whole section of bases within a gene is moved and inserted into a different chromosome
6
Q
What is an inversion mutation?
A
When sections of bases are put in reverse order
7
Q
What is a duplication mutation?
A
When sections of bases are copied (and thus repeated)
8
Q
What type of substitution mutation is least harmful? Why?
A
A silent mutation. The same amino acid is coded for, so the same polypeptide, and this protein is also coded for
9
Q
What type of substitution mutation is most harmful? Why?
A
Nonsense mutation. The substituted base (in the triplet) codes for a stop codon, so no more AAs are added to the polypeptide, so the tertiary structure is severely affected (reference to bonds)
10
Q
When will an addition (/insertion) or deletion mutation not cause a frame shift?
A
If a multiple of 3 bases is inserted or deleted then the other amino acids coded for my the gene will not be frame-shifted
11
Q
Why do some types of mutation not cause a change in the structure of the protein produced?
A
- The genetic code is degenerate
- Same amino acid may be coded for
- Same H/ionic/disulphide bonds form in the same place
- Protein forms same tertiary structure
(even if a different AA was coded for it MIGHT form the same H /ionic/ disulphide bonds, so still same structure)
12
Q
TRUE OR FALSE?
All the cells in the body contain the same DNA and the same genes
A
TRUE
All the cells in the body contain the same genes
(Only some of the genes are expressed)
13
Q
How do multicellular organisms have specialised cells?
A
The cells undergo differentiation to become specialised
14
Q
What it mean when a gene is expressed?
A
The coded protein is produced in transcription and translation
15
Q
What are totipotent cells?
A
Cells which have the capability to differentiate into ANY body cell
16
Q
What are stem cells?
A
- Undifferentiated, dividing cells
- Can divide to form copies of themselves (replacement)
- Only a few types in mature mammals can differentiate into other types of cell
17
Q
Where are adult stem cells found?
A
Found in all body tissues
18
Q
What are adult stem cells?
A
- Stem cells which are specific to the tissue/organ where produced
- Used to maintain + repair tissues throughout life
19
Q
What are the 4 types of stem cell?
A
- Totipotent
- Pluripotent
- Multipotent
- Unipotent
20
Q
Where are totipotent cells found?
A
In Zygote early stages
21
Q
Where are pluripotent cells found?
A
Embryo
22
Q
Where are multipotent cells found?
A
Mature mammals
23
Q
Where are unipotent cells found?
A
Mature mammals
24
Q
What can totipotent cells differentiate into?
A
All body cells
25
What can pluripotent cells differentiate into?
All body cells except the placenta + umbilical cord
26
What can multipotent cells differentiate into?
A limited number of specialised cells
27
What can unipotent cells differentiate into?
A single type of cell (tissue specific)
28
How can colonies of pluripoent cells be made specific?
A differentiation factor is added to differentiate the pluripotent cells into a specific type of cell
29
What are induced pluipotent cells (iPS)?
- Pluripotent cells produced from adult cells
- Can divide into unlimited numbers (self-renewal)
- Can be used to treat human disorders
30
How can induced pluripotent cells be produced?
- Body cells genetically altered in the lab
- Induces genes and transcription factors to express themselves
Basically turns on and off the necessary genes
31
Name 3 advantages of induced pluripotent cells.
1. No immune response because they are the patients' own cells
2. Stem cells can be obtained from adult cells
3. Can produce all types of cell
32
Why are induced pluripotent stem cells more ethically suitable than embryonic stem cells?
iPS are produced from adult cells, so embryonic cells are not used
33
Give two characteristics of stem cells
1. Capable of self-renewal
| 2. Can differentiate into any cell
34
TRUE OR FALSE?
| Mature plants have many totipotent cells
TRUE
| Mature plants have many totipotent cells which can develop into any cell type
35
What regulates the differentiation of totipotent cells in plants?
Growth regulators
36
Name 4 features of growth regulators in plants.
1. Have a range of effects on plant growth
2. The effects depend on the concentration of growth factors
3. The same conc affects different types of tissue in different ways
4. The effect of a growth factor can be modified by the presence of another
37
Why can't xylem differentiate into other cells?
Xylem is dead tissue, so it cannot differentiate into other cells
38
What do growth regulators enable plants to do?
If placed in a growth medium, plants cells from a single plant will develop into new plants
Called IN VITRO DEVELOPMENT
39
What is in vitro development?
When growth regulators are used to create cloned plant tissue (a tissue culture)
40
What will plants derived from tissue culture share?
The same genetic material
41
What is the term for undifferentiated plant cells?
Callus
42
How do cells differentiate?
Certain genes must be switched on (expressed), whilst others are switched off
43
How are genes expressed (or not)?
Transcription and translation are regulated
44
What is transcription (basic)?
DNA -> mRNA
45
What is a transcription factor?
- A protein/ molecule that moves from cytoplasm to DNA
- Binds to a specific gene
- Allowing / blocking protein production
46
How do transcription factors bind to DNA?
- Specific DNA binding site
- Complementary to the specific shape of a DNA base sequence
- Binds to a PROMOTER REGION
47
What do transcription factors enable?
Enables RNA polymerase to bind to DNA, allowing or preventing transcription
48
What is an example of a molecule activating transcription factors?
Oestrogen
| Has specific functional site which can bind to a receptor on a TF so the TF changes shape to become active
49
What are the proteins that Eukaryotic DNA associates with called?
Histones
50
What is the DNA associated with histones called?
Chromatin
51
What is highly-coiled DNA and histones called?
Heterochromatin
52
What is loosely-coiled DNA and histones called?
Euchromatin
53
Why can DNA be transcribed when DNA and histones are loosely associated (euchromatin)?
Transcription factors are able to bind to the promoter region of DNA
54
What is epigenetics?
A process by which environmental factors can cause heritable changes in gene function without changing the base sequence of DNA
55
What does Methylation cause? (4 points)
- DNA nucleotides pack together tightly
- Creates heterochromatin
- TFs cannot bind
- Genes are not transcribed or translated
56
What does Acetylation cause? (4 points)
- DNA nucleotides are loosely packed together
- Creates euchromatin
- TFs can bind to the DNA
- Genes are transcribed and translated
57
Does the DNA sequence change in the epigenome?
No
58
What attaches to DNA to cause methylation and acytylation?
Chemical tags causing DNA to become more/less associated with histones
59
What is it called when inactive genes are kept tightly packed in epigenetics?
Epigenetic silencing
60
What does the acetylation of histones actually do?
Masks the +ve charge of histones (makes them -ve) so less attracted to -vely charged DNA
61
What factors influence the epigenome?
The accumulation of signals in the foetus and during a lifetime
62
Where do epigenetic signals come from in the foetus?
Nutrition provided by the mother (eg smoking)
63
Where do epigenetic signals come from during life?
- Environment (diet/stress etc)
| - Signals within the body (hormones)
64
Can epigenetic tags be inherited?
Not many
- Tags are erased in gametes
- Some epigenetic tags escape the process and become inherited by the next generation
65
Considering epigenetics, why is reproductive cloning difficult?
All epigenetic tags are inherited, so some genes are not able to be expressed because TFs cannot bind to promoter regions, so transcription and translation do not occur
66
How can epigenetics trigger cancers?
- Promoter regions on some tumour suppressor genes become hyper-methylated
- The promoter region becomes inactive
- No transcription of tumour surpressor occurs
- Cell division increases
(can also be reduced methylation of oncogenes, same effect)
67
1. How can drugs be used to counteract epigenetic changes which could cause cancer?
2. What is a disadvantage of this?
1. Drugs can target enzymes involved in histone acetylation and DNA methylation
2. If not carefully administered to target cancerous cells then the risk of cancer could increase in other tissues
68
How can epigenetic tags be used in medical diagnosis?
- The levels of DNA methylation and histone acytylation can allow the cancer risk to be identified
- Treatment can start sooner, so a better prognosis
69
In epigenetics, where does methylation occur?
On DNA itself
70
In epigenetics, where does acetylation occur?
On the histones in chromatin
71
Where does transcription take place?
- pre-mRNA produced in the nucleus
- pre-mRNA is spliced to remove introns
- mRNA leaves the nucleus vias nuclear pore
- mRNA ready for translation
72
Where does translation take place?
In ribosomes on he rough ER or cytoplasm
73
What is siRNA?
- Small Interfering RNA
- Inhibits the production of a gene by translation
- Breaks down mRNA before info is translated into a polypeptide
Regards interfering with gene expression
74
How does siRNA break mRNA? (4 points)
- Enzyme cuts large double-standed sections of RNA into siRNA
- One stand of the siRNA combines with an enzyme
- siRNA guides the enzyme to mRNA with comp base pairing
- Enzyme mRNA into smaller sections
75
What effect does siRNA have on protein synthesis? (3 points)
- mRNA is cut by enzyme associated with siRNA
- mRNA unable to be translated into the correct polypeptide
- The gene is not expressed
76
What is cancer?
A group of diseases caused by damage to genes that regulate mitosis and the cell cycle
Causes uncontrolled cell growth (tumour)
77
How many cells cause cancer?
Just one which subsequently divides
78
What are the two types of tumour?
Benign and malignant
79
What is a benign tumour?
A non-cancerous tumour
80
What is a malignant tumour?
A cancerous tumour
81
What are some key characteristics of malignant tumours? (4 points)
- Grow rapidly and aggressively
- The cell nucleus is distinctive
- Cause secondary tumours
- The tumours are not surrounded by a capsule, so have finger-like projections
82
What two mutations cause a cancer cell?
1. Initial mutation leads to uncontrolled mitosis in cells
| 2. Mutation in decedent cells causes subsequent cells to become abnormal
83
What two genes control cell division?
Proto-oncogens and tumour suppressor genes
84
What the normal function of proto-oncogens?
Stimulates cell division
Like an accelerator pedal in a car, it controls how fast the cell cycle goes
85
What is the normal function of tumour suppressor genes?
Slow down cell division and program cell death
86
How do proto-oncogens function normally?
- Growth factor binds to receptor protein
- Relay protein is released
- Relay proteins cause genes needed for DNA replication in the nucleus to be switched on
87
What are oncogens?
Mutations of proto-oncogens which cause oncogens to be permanently switched on
88
What are the 2 reasons for an oncogene to become permanently activated?
1. Receptor protein on the cell surface membrane can be permanently activated - no effect with growth factors
2. Growth factors may be produced in excessive amounts
89
What are the possible consequences of a mutation to a tumour suppressor gene? (4 points)
- The gene becomes inactive
- Cell division increases
- Cells become structurally/functionally different +dies
- Some cells survive and clone by mitosis = tumour
90
How many mutated alleles do you need to trigger cancer with tumour suppressors?
2
| Even if one tumour suppressor gene is mutated, faulty, the other will still work and regulate cell division
91
How many mutated alleles do you need to trigger cancer with oncogenes?
1
| A mutation to one allele will still cause the gene to be expressed and to have an effect
92
What can be done to prevent, treat and cure cancer?
- Preventing the expression of oncogenes
| - Introducing tumour suppressor genes into cancerous cells
93
What is an example of a molecule that activates transcription factors?
Oestrogen
94
What is a genome?
- The complete set of genetic info in a organism
| - Provides all the info an organism needs to function
95
What are the 3 aims of genome projects?
1. Determines the entire nucleotide sequence of organisms
2. Maps the DNA sequence of all genes on an organism
3. Maps the genes to individual chromosomes
96
How many genes do Homo sapiens have?
20,000 - 25,000
97
How many base pairs to Homo sapiens have?
3 billion
98
What % of base pairs code for proteins in humans?
1%
99
What is bioinfomatics?
The science of collecting + analysing complex biological data using algorithms + maths
100
What charge does DNA have?
Negative charge
101
In gel electrophoresis, what direction does DNA move?
From negative to positive charges because DNA is negatively charged
102
Which DNA fragments move furthest in gel electrophoresis?
The shortest fragments
103
Why do the shortest DNA fragments move the furthest in gel electrophoresis?
The fragments can move through the gel granules more easily, so move through the gel faster and thus cove more distance in a given time
104
How do you measure the size of DNA fragments in gel electrophoresis?
Using a ladder of DNA fragments of known lengths
105
What was the name of the original process used to sequence DNA?
Sanger Sequencing
106
What is a primer?
- A short piece of DNA that binds to template DNA
- Gives something for DNA polyerase to join onto to start a new chain
(used in Sanger Sequencing)
107
Why are new methods of DNA sequencing better?
- Less labour intensive
- Faster turnaround
- Longer DNA strands can be sequenced
108
What are the 3 steps involved in whole genome shotgun sequencing?
1. DNA cut into short sequences
2. Computer algorithms align overlapping segments
3. Assembles the entire genome
109
What are Single Nucleotide Polymorphisms (SNPs)?
- Single base sequences in the genome associated with some disorders
- >1.4 million SNPs in the human genome
110
What is a proteome?
All the proteins produced in a given cell/organism at a given time, under specific conditions
111
Why is determining the proteome easier in prokaryotes>
- One circular piece of DNA
- DNA unassociated with histones
- No introns
112
Why can sequencing genomes of pathogens help cure diseases?
- Identifies the DNA sequence of antigens on pathogens
| - Allows vaccines of the complementary antibody to be produced
113
TRUE OR FALSE?
| Introns are found are found between genes?
FALSE
Introns are repeated bases within genes
VNTRs are repeated base sequences found between genes
114
What is a gene?
A section of DNA that codes for a polypeptide or a functional RNA (tRNA and rRNA)
115
Why is the genetic code said to be universal?
The same triplet codes for the same amino acid in all organisms
116
What is recombinant DNA?
Combining DNA from 2 different organisms
117
What is a transgenic organism?
An organism that has had its genome modified
Also called Genetically Modified Organism (GMOs)
118
What does recombinant DNA technology do? (3 points)
- Transfers DNA from one organism/species to another
- The gene is translated in the recipient
- Due to the universal nature of DNA
119
What does IN VIVO mean?
Inside a living organism
120
What does IN VITRO mean?
In glass (not a living organism)
121
What are the 5 basic stages of recombinant DNA?
1. Isolation of gene
2. Insertion of the gene into a vecto
3. Transformation of gene + vector into a host
4. Identification of the gene that has been taken up by the host using gene markers
5. Growth/cloning of cells
122
Why can't eukaryotic genes be successfully transcribed + translated in prokaryotes?
- Eukaryotic genes contain introns
- Prokaryotes lack enzymes to remove introns
- The protein would thus contain additional AAs and would not fold correctly (BONDS + STRUCTURE)
123
How can the effect of introns in recombinant eukaryotic DNA (into Prokaryotes) be overcome?
Using reverse transcriptase on mRNA (post-splicing)
124
What is REVERSE transcription?
mRNA -> complementary DNA (cDNA)
125
What is complementary DNA (cDNA)?
- DNA that does not contain introns
- Complementary to mRNA
- Made from mRNA using reverse transcriptase
126
What organisms use reverse transriptase?
RNA Viruses (used in virus replication)
127
What are restriction endonucleases?
A group of enzymes which cut DNA at specific base sequences called recognition sites
128
What are restriction endonucleases used for in recombinant DNA?
Used to cut desired genes from DNA of one organism in order to insert it into the DNA of another
129
What are the two types of 'end' cut with restriction enzymes?
Blunt- and sticky-ends
130
What are palindromic sequences?
Sequences on two single strands of DNA which are read the same backwards
Eg
ACGT on one strand
TGCA on the other
131
What is a sticky end?
- Formed when restriction endonuclease cuts at different positions on two DNA strands
- Leaves each section with a single-stranded part complementary to the other
132
What is a blunt end?
When a restriction enzyme cuts at the same position on each strand
133
Why can you join together two DNA fragments from different organisms that have both been cut by the same sticky-end restriction endonuclease?
The single strands of exposed bases are complementary because the restriction endonuclease has a specific, unique shape
134
What enzyme is used in recombinant DNA to join together two adjacent nucleotides joined by sticky ends?
DNA LIGASE (not polymerase because only one phosphodiester bond is being formed)
135
How does a gene machine artificially produce a gene? (5 steps)
1. Computer designs a series of DNA sequences
2. Small single-stranded sequences assembled by adding one nucleotide at a time
3. Small sequences combined to create a gene
4. Gene replicated using PCR
5. Gene inserted into vector (using restriction endonucleases + DNA Ligase)
6. Vector transferred to a host cell to be replicated
136
Why are gene machines useful?
- Sequence of DNA can be produced in a short amount of time
- The genes are artificial and contain no introns
- No introns = transcription + translation in prokaryotes
137
What do genes need to function effectively?
A promoter region and a terminator region
138
What are promoter regions?
The specific sequence of DNA nucleotide bases where RNA polymerase and transcription factors bind, allowing the production of mRNA by transcription
139
What are terminator regions?
The specific sequence of DNA nucleotide bases where transcription factors disengage from DNA, stopping transcription
140
What bonds does RNA polymerase form between nucleotides?
Phosphodiester bonds
141
What needs to be added to a DNA fragment before insertion into a vector?
- Promoter at the front so RNA polymerase + TFs bind and produce mRNA
- Terminator at the end of the gene to stop trnascription
142
What does a restriction endonuclease do?
Cuts DNA at specific palindromic recognition sites
143
Why are sticky ends better than blunt ends when joining DNA strands together?
- Same restriction endonuclease used (specific structure)
- The ends of the two fragments from the two species will be complementary
- H bonds will form between the complementary bases
144
What is a vector in recombinant DNA technology?
A DNA molecule that carries foreign DNA into a cell where it can replicate or be expressed
Example: Bacterial plasmid
145
What is the advantage of inserting recombinant plasmids into host cells?
Every time the bacteria containing the plasmid replicate by binary fission, the plasmid will also replicate
146
What is it called when DNA is inserted into host cells?
Transformation
147
How can transformation of bacteria and recombinant plasmids occur?
Heat shock treatment
- Bacteria mixed with calcium ions to make permeable
- Chilled on ice
- Warmed to 42 degrees
148
What is an alternative method, besides heat shock, of transformation? (recombinant DNA technology)
Electroporation
149
What is the purpose of heat shock / electroporation in transformation?
Makes the bacteria more permeable so the recombinant plasmid is taken up (and can thus be replicated or expressed in the bacteria)
150
Why do only some bacteria possess DNA fragments after transformation? (3 points)
- Only some take up the recombinant plasmid
- Some plasmids self-ligate (recreating the original plasmid)
- DNA fragments may join together to make own plasmid
151
By what process do bacterial cells divide?
Binary fission
152
How can transformed bacteria be identified? (recombinant DNA)
Using genetic markers to identify which bacteria contain the plasmid with the desire gene
153
What are 3 types of genetic markers?
1. Enzymes
2. Antibiotic resistance genes
3. Florescence genes
154
How were marker genes previously used?
- Desired gene inserted into one marker gene (antibiotic resistance gene) using comp restriction enzymes, sticky ends and ligase
- The antibiotic is added and the bacteria with the desire genes die (as gene is not intact)
155
How are marker genes now used?
- The antibiotic resistance marker gene is connected directly to the gene of interest
- Only the bacteria which survive the antibiotic have the desired gene
156
When growing bacteria in a culture to obtain desired genes, what should be done? Why?
- The bacteria should be diluted
- Allows the bacterial solution to spread out on the agar jelly
- Allows room for each individual bacteria to produce a clear colony
157
What will happen if a desired gene is inserted directly into a fluorescence gene? (Recombinant DNA tech)
- When UV light is shined onto the bacterial culture the bacteria which are not glowing will contain the desired gene
- The fluorescence gene will be split so no longer functions (as cannot be transcribed + translated)
158
Why is using fluorescent gene markers better than using antibiotic gene markers when identifying recombinant bacteria?
- Unlike with antibiotics, the bacterial cells with the desired gene are not killed
- Thus no need for REPLICA PLATING
- So faster
159
What is a method of in vivo gene cloning?
Using bacterial plasmids and transforming bacteria to replicate the desired gene
160
What is a method of in vitro gene cloning?
The Polymerase Chain Reaction (PCR)
161
What 5 components are needed to replicate DNA in vitro (PCR)?
1. DNA fragment to be copied
2. Pair of primers
3. DNA nucleotides in excess
4. Thermostable DNA polymerase
5 Thermocycler
162
What are DNA primers?
Short lengths of DNA that are complementary to the 3' end of each DNA strand
163
Why are DNA primers needed for PCR?
- Polymerase can only add nucleotides to an existing strand
- Gives polymerase something to join new nucleotides to
- Prevents the separated strands from rejoining
164
Why is a heat stable DNA polymerase used in PCR?
Ordinary DNA polymerase (enzyme) will denature in the high temps used in PCR
165
What is a thermocycler?
Where is it used?
A computer-controlled machine which varies temperatures precisely over a period of time
Used in the polymerase chain reaction (PCR)
166
What are the 3 steps of the polymerase chain reaction?
1. Separation of DNA strands
2. Annealing of primers
3. Synthesis of DNA
167
At what temperatures do the 3 steps of the polymerase chain reaction take place?
1. Separation of DNA strands
95C
2. Annealing of primers
50-60C
3. Synthesis of DNA
72C
168
What takes place during the first step (separation of DNA strands) in PCR?
- H bonds break in high temp (95C)
- Strands separate
- Each strand acts as a template
169
What takes place during the second step (annealing of primers) in PCR?
- Primers join to complementary exposed bases at the end of the DNA fragment
- Provide starting sequence of DNA polymerase
- Prevents original strands rejoining
170
What takes place during the third step (synthesis of DNA) in PCR?
- Complementary DNA nucleotides make H bonds with exposed bases on each template strand
- Beginning at the primer, DNA polymerase joins nucleotides by phosphodiester bonds along one of the template strands, forming 2 copies
171
How are hydrogen bonds between complementary bases broken in the polymerase chain reaction?
H bonds broken by heat
172
How are hydrogen bonds between complementary bases broken in DNA replication?
H bonds broken by DNA helicase
173
Are primers (supposedly) needed in DNA replication?
No (although technically they do)
174
Why is PCR useful in forensic science?
Allows lots of DNA to be amplified from a small amount extracted at a crime scene
175
What happens to the amount of DNA in a PCR machine each time it completes one cycle?
The amount doubles
176
What are the 4 advantages of in vitro gene cloning (PCR)?
1. Makes lots of DNA from a small amount
2. Faster than conventional methods
3. Allows for forensic analysis
4. Cells do not need to be cultured
177
What are the 4 advantages of in vivo gene cloning (plasmids in bacteria)?
1. Desired gene can be easily transferred once in a vector to another organism
2. Only the gene with complementary sticky ends is cloned, so less risk of contamination
3. Accurate replication in organisms
4. Can clone specific genes
178
What are the 2 disadvantages of in vitro gene cloning (PCR)?
1. Risk of contamination (errors) - the contaminated DNA is also amplified
2. Less accurate than in vivo gene cloning
179
What are somatic cells?
Any cell other than reproductive cells
180
What are germ cells?
Reproductive cells/gametes
181
Why are genes isolated and cloned using recombinant DNA technology?
Allows genetic disorders to be treated
182
What is somatic cell therapy?
- Targets specific tissues (eg lungs)
- The altered gene is not present in other cells
- Cells are constantly replaced by mitosis, so treatment needs to be repeated
183
What is germ cell therapy?
- Occurs in fertilised eggs
- Results in all cells of organisms possessing the gene
- Faulty gene not passed onto future generations
CURRENTLY PROHIBITED
184
Why is the risk of contamination very low in vivo gene cloning?
- The gene and the plasmid are both cut by the same restriction endonuclease which is specific to the structure of complementary bases
- Only the desired gene is taken up
185
Why is in vivo gene cloning better for transforming genes into living organisms?
- Uses vectors
- Transformed plasmids can be used to deliver the gene to another organism
- The basics of gene therapy
186
What type of mutation causes cystic fibrosis?
A deletion mutation of 3 adenine bases
187
Why are viruses suitable for gene therapy?
- Specifically target certain cells with specific receptors
- Adapted to insert DNA into host cells
- Host cells produce viral proteins by transcription + translation
188
How can a virus be used to treat genetic respiratory disorders (CF)?
- The desired gene is inserted into harmful gene
- Harmful gene becomes inactive
- The desired gene is inserted into the affected cell
189
Why are viruses used in gene therapy grown in a human cell culture?
The viruses need a cellular host to replicate as the host possesses the enzymes + nucleotides required for replication
190
Why is using a virus as a method of gene therapy risky? (2 reasons)
- Viruses could still cause infections by recovering the ability to cause disease
- The patient may develop immunity to the virus (as it is a foreign body), so antibodies may be produced for a secondary response to future treatment
191
What is an alternative method of gene therapy to the use of viruses?
Liposomes
192
Why are liposomes used in respiratory gene therapy?
They are made from lipids so fuse with the phospholipid bi layer and offload the desired gene
193
What are 3 disadvantages of gene therapy?
1. If viruses are used they could become harmful
2. The genes inserted are not always expressed
3. Only useful for diseases caused by a single faulty gene (i.e. recessive alleles)
194
Why does somatic gene therapy have to be repeated?
- The inserted gene is not incorporated into chromosomes
| - When the cell replicates the gene is not copied
195
Why does gene therapy become less effective with successive treatments?
- Virus/liposomes trigger an immune response
- Subsequent treatments also trigger an immune response which is of greater magnitude + speed
- Antibodies coat virus/liposomes, preventing the desired infetion of target cells
196
Why is it difficult for genetic therapy to be used to treat genetic diseases caused by multiple genes?
- All the corrected genes need to be expressed in the same cell
- Low probability of this
197
What is a DNA probe?
A short single-stranded length of DNA that has a label attached to make it identifiable
198
Name two examples of DNA probes used
1. Radioactively-labelled probes
| 2. Fluorescently labelled probes
199
What is a DNA probe made of?
- A short sequence of DNA
- Complementary nucleotide base sequence of gene of interest
- Label added
200
How is DNA to be screened using a probe broken into single strands?
Using an alkali to break the H bonds between opposite complementary bases
201
How can short, known sequences of DNA be made (eg for use as a probe)?
Using a gene machine
202
How can multiple copies of a DNA probe be made?
Using PCR
203
How does a DNA probe bind to a screened DNA fragment?
- If the gene being screened for is present
- The probe will bind to this gene
- By complementary base pairing
204
Why is screening of tumour suppressor genes important?
- Mutations to both tumour suppressor alleles could make the tumour suppressor gene inactive
- Could cause a tumour
- Screening allows for hetrozygous individuals to be identified
205
What is personalised medicine?
A method adopted by doctors which allows advice and healthcare to be prescribed based on a patients genotype
206
What is the main benefit of personalised medicine?
Allows the most effective drug to be given to a patient so drug selection and dosage is tailored.
This saves money and sometimes prevents harm
207
What is genetic counselling?
Advice given to patients or relatives at risk from an inherited disorder
208
What sort of advice do genetic counsellors give?
- Info in consequences + nature of the disorder
- Probability of developing/inheriting it
- Possible options for family planning
209
What is DNA gel electrophoresis?
A technique used to separate DNA strands according to their length
210
How can the lengths of DNA be calculated from gel electrophoresis?
Run a DNA ladder on the gel which indicates fragments of known sizes
211
What is genetic fingerprinting?
A diagnosis tool used by forensic and medical scientists
212
Is the DNA of EVERY individual unique?
No EVERY individual. Every individual except identical twins
213
What are Variable Number Tandem Repeats (VNTRs)?
Non-coding parts of DNA found BETWEEN GENES
214
What is the difference between introns and VNTRs?
Introns are non-coding parts of DNA found WITHIN genes
VNTRs are non-coding parts of DNA found BETWEEN genes
215
How many VNTRs between the same gene do different people have?
Different numbers, unique to individuals
216
Why are VNTRs useful?
Each individual has a different number of VNTRs between the same genes, so a unique "bar code" exists
This is a genetic fingerprint
217
How can DNA VNTRs be isolated for genetic fingerprinting?
With the use of restriction endonucleases
218
Which VNTR moves the furthest in gel electrophoresis?
The shortest
219
Which specific part of DNA is negatively charged?
The phosphate group
220
Why are DNA fragments (VNTRs) transferred onto a nylon membrane in gel electrophoresis?
The gel is fragile
221
What are the 7 steps of gel electrophoresis?
1. DNA extracted from a sample and cut into fragments using restriction endonucleases
2. Fragments added to electrophoreisis gel and separates according to size (shortest furthest etc)
3. DNA fragments transferred onto nylon membrane
4. DNA made single stranded (heat/alkali)
5. Radioactively-labelled DNA probes are added
6. Membrane put onto x-ray film
7. X-ray film develops with distinctive dark bands
222
Which type of cell contains no nucleus?
Red blood cells
223
Why can't red blood cells be used for genetic fingerprinting?
No nucleus, so no genetic material (DNA) is stored to be used
224
What is the difference between a gene machine and PCR?
Gene machine makes short sequences of DNA
| PCR makes multiple copies of genes (long sequences)
225
In gel electrophoresis, why is the nylon membrane washed after adding probes and before exposing to x-ray film?
It washes away any unbound DNA probes so that they do not contaminate the x-ray film so no false positive results
226
Give 5 uses of genetic fingerprinting
1. Forensics
2. Paternity tests
3. Archaeology
4. Animal population studies
5. Organ donor selection
227
What are ethics?
- Narrower than social morals
| - A set of standards set up by a society to regulate behaviour
228
What are social issues?
- Relates to human society and organisation
| - Concerns the mutual relationship of human beings, their independence and cooperation to benefit all
229
What is required with recombinant DNA technology?
Evaluations
Must revise this briefly (page 542)
