MGD Session 9 Mutagenesis Flashcards
Explain the relationship between changes in nuceleotide and amino acid sequence
A change in the genetic code (base sequence) can result in different amino acids being coded for.
What effect does a change in amino acid sequence have on its protein?
Changes it shape, so changes its function
What are two types of point mutation?
Transition or transversion
What is a transition mutation?
Purine–>Purine
or
Pyridimine –> Pyridimine
What is transversion?
Pyridimine –> Purine
or
Purine –> Pyrimidine
What are single base mutations also known as? (haircut)
Single Nucleotide Polymorphisms
Name three types of mutation other than transition?
SIlent, missense and nonsense
How can point mutations outside of coding regions be detrimental? (3)
Can occur in binding sites, promoter sequence and splice sites.,
What are insertions and deletions?
When a sequence is added or removed from dna sequence. Can be single, a few nucleotides or millions of nucleotides long
What type of mutation can insertions and deletions cause?
Frameshift mutation
What is a silent mutation?
A mutation that does not alter the amino acid specified (but sometimes can disrupt RNA splicing).
What is a missense mutation?
A mutation that replaces one amino acid with another. Some can be tolerated better than others, especially in non-critical regions of proteins.
What is a nonesense mutation?
A mutation that changes the amino acid specified to a stop codon
What is a frameshift mutation?
Addition or subtraction of nucleotides not in multiples of 3(insertions, deletions or splice site mutations. Stop codons often found
What happens to mRNAS which contain premature termination codons?
They are degraded by nonesense mediated decay, so little or no protein is produced
AUG (Met) ACG (Thr) CUA (Leu) AUG (Met) ACG (Thr) CUG (Leu)
Silent mutation
AUG (Met) ACG (Thr) UGG (Trp) AUG (Met) ACG (Thr) UGA (Stop)
Nonesense mutation
AUG (Met) ACG (Thr) GAC (Asp) AUG (Met) ACG (Thr) GAA (Glu)
Missense mutation
GGA (Gly) GCA (Ala) CCA (Pro) GGA (Gly) GGA (Gly) GCC (Ala) ACC (Thr) AGG (Arg)
Frameshift mutation
What happens if a mutation occurs at an intron splice site?
Results in skipping of exon adjacent to mutation
When an exon is skipped due to mutation an intron splice site, what happens if it is a multiple of 3 bp?
The mRNA will be shortened but still in frame
When an exon is skipped due to mutation an intron splice site, what happens if it is not a multiple of 3 bp?
mRNA will contain PTC, resulting in nonesense mediated decay
Name three ways in which spontaneous mutations occur?
Slippage during replication - One strand folds back on itself and is left out of replication
Errors during DNA repair
DNA bases have slight chemical instability
What two things does the rate of spontaneous mutation depend on?
Size and Sequence
What is spontaneous mutation not caused by?
Exposure to known mutagen
What are induced mutations caused by?
Mutagens and Carcinogens - Chemicals the cause mutations and cancer respetively
Give three examplesof mutagens and their affects
Aklylating agents - Remove a base
Acridine agents - Add or remove a base
UV radiation - Creates thymidine dimers
Define mutation
‘a change in a nucleic acid sequence, which can be the addition of one or more (or many) nucleotides [insertion], the removal of one or more (or many) nucleotides [deletions], or the rearrangement of several (or many) nucleotides’
What is a wild type?
an individual within a population displaying a wild-type trait, which is the trait that is most common in that population’
What kind of phenotype does a mutation cause?
A mutant phenotype
What does a mutation in a gene cause?
A mutant allele
What happens if mutations occur in the germline?
Germline mutations
What is mismatch repair?
This occurs when enzymes detect nucleotides that don’t base pair in newly replicated DNA.
The incorrect base paired is excised and replaced. These bases exist because of failure of proof reading
Excision Repair
DNA damaged by external agents such a chemical mutagens is removed by excision of bases and replacement by DNA polymerase.
Nucleotide excision repair
replaces up to 30 bases and is used in repair of UV Damage and some carcinogens.
Base excision repair
replaces 1-5 bases and repairs oxidative damage. Fairly long patches of new DNA synthesis on repaired strand.
Which protein is called the guardian of the genome, and what is its role?
p53 monitors repair of damaged DNA and if damage is too severe it promotes apoptosis
What can be inherited which leads to increased gene mutation?
Mutations in the genes encoding the DNA repair proteins can be inherited. Therefore, there is an overall increase in mutations, as errors/DNA damage isn’t repaired efficiently.
Base excision repair
replaces 1-5 bases and repairs oxidative damage. Fairly long patches of new DNA synthesis on repaired strand.
How are cancerous cells produced?
If DNA is damaged the extent that apoptosis (programmed cell death promoted by p53 protein) doesn’t occur or the damage leads to uncontrolled growth then cancerous cells are produced.
What enzymes can potentially be rendered non-function in order for cancer to occur?
Can occur as result of mutational and resultant non-functionality in mismatch repair enzymes.
What are tumours?
- Tumours are derived from individual abnormal cells.
- They arise from the lack of normal growth control.
Which type of cell are tumours more likely to arise from?
cell types undergoing frequent cell division.
What does the behaviour of a tumour depend on?
Cell type
What are oncogenes responsible for?
Control of ceoll division
What is the function of an oncogene
To stimulate/inhibit growth
What is the role of a tumour supressor gene?
To protect the cell against one step on the path to cancer
How do viruses cause cancer?
Carry copies of oncogenes
What are the steps of PCR?
- DNA is denatured @ 950C for 1 minutes
- Anneal the primers @ 550C for 1 minute
- Copy the DNA @ 720C for 2 minutes
How is PCR used to detect sickle cell disease?
In Sickle Cell disease the restriction site for the enzyme MstII is destroyed:
CCTGAGG CCTGTGG
Therefore, with Gel Electrophoresis/Southern blotting the mutated gene will have less/one DNA fragments if digested with MstII, as it lacks the site.
Why can’t straight PCR be used to analyse Cystic Fibrosis?
3 base deletion (No frameshift)
Loss of Phenylalanine
When is southern blotting used?
when there is a need to analyse larger segments of DNA within and around a gene.
Name two conditions southern blotting is used to analyse
triplet repeat disorders such as Huntington’s disease and Fragile X syndrome.
What is array comparative genomic hybridisation used to analyse?
sub-microscopic chromosomal deletions for which the location (locus) cannot be deduced from the patient’s phenotype.
How does array CGH work?
- Array of DNA probes covering entire genome
-Patient DNA and normal control DNA labelled with different coloured fluorescent tags
Equal amounts of labbeled DNA are then hybridised to probe array
Hybridisation signals detected and compared
What happens when probes of normal DNA exceed that of the patients DNA?
the patient has a deletion of the chromosomal region from which that probe was derived.
Which protein is called the guardian of the genome, and what is its role?
p53 monitors repair of damaged DNA and if damage is too severe it promotes apoptosis
What can be inherited which leads to increased gene mutation?
Mutations in the genes encoding the DNA repair proteins can be inherited. Therefore, there is an overall increase in mutations, as errors/DNA damage isn’t repaired efficiently.
How are cancerous cells produced?
If DNA is damaged the extent that apoptosis (programmed cell death promoted by p53 protein) doesn’t occur or the damage leads to uncontrolled growth then cancerous cells are produced.
What enzymes can potentially be rendered non-function in order for cancer to occur?
Can occur as result of mutational and resultant non-functionality in mismatch repair enzymes.
What are tumours?
- Tumours are derived from individual abnormal cells.
- They arise from the lack of normal growth control.
Which type of cell are tumours more likely to arise from?
cell types undergoing frequent cell division.
What does the behaviour of a tumour depend on?
Cell type
What are oncogenes responsible for?
Control of ceoll division
What is the function of an oncogene
To stimulate/inhibit growth
What is the role of a tumour supressor gene?
To protect the cell against one step on the path to cancer
How do viruses cause cancer?
Carry copies of oncogenes
What are the steps of PCR?
- DNA is denatured @ 950C for 1 minutes
- Anneal the primers @ 550C for 1 minute
- Copy the DNA @ 720C for 2 minutes
How is PCR used to detect sickle cell disease?
In Sickle Cell disease the restriction site for the enzyme MstII is destroyed:
CCTGAGG CCTGTGG
Therefore, with Gel Electrophoresis/Southern blotting the mutated gene will have less/one DNA fragments if digested with MstII, as it lacks the site.
Why can’t straight PCR be used to analyse Cystic Fibrosis?
3 base deletion (No frameshift)
Loss of Phenylalanine
When is southern blotting used?
when there is a need to analyse larger segments of DNA within and around a gene.
Name two conditions southern blotting is used to analyse
triplet repeat disorders such as Huntington’s disease and Fragile X syndrome.
What is array comparative genomic hybridisation used to analyse?
sub-microscopic chromosomal deletions for which the location (locus) cannot be deduced from the patient’s phenotype.
How does array CGH work?
- Array of DNA probes covering entire genome
-Patient DNA and normal control DNA labelled with different coloured fluorescent tags
Equal amounts of labbeled DNA are then hybridised to probe array
Hybridisation signals detected and compared
What happens when probes of normal DNA exceed that of the patients DNA?
the patient has a deletion of the chromosomal region from which that probe was derived.
