Mechanisms and Effects of Mutations

Question 1

How do mutations arise?

Answer 1

• Random errors in DNA replication/repair
• Exposure to mutagenic agents

Question 2

What are polymorphisms and what types are there?

Answer 2

• Non-harmful mutations
• Sequence variant in non-coding DNA
• Sequence variant in gene but doesn’t change amino acid
• Sequence variant changes amino acid but not protein function

Question 3

What is a single nucleotide polymorphism?

Answer 3

One base in the sequence is changed (eg. C changed to T)

Question 4

How can the bases of the genome be examined?

Answer 4

• Sequencing
• Microarray analysis

Question 5

How can large blocks of DNA be examined?

Answer 5

• Microarray analysis
• Fluorescence in situ hybridisation (FISH)

Question 6

How can chromosomes be examined?

Answer 6

Light microscopy

Question 7

Method of DNA sequencing

Answer 7

Sanger sequencing:
- Amplify very small amounts of target DNA fragments (PCR)
- Fragments added to 4 mixtures, all containing all 4 deoxynucleotides (normal nucleotides), and each containing 1 of the dideoxynucleotides (OH group on 3’ carbon > H)
- No nucleotides can be added after a dideoxynucleotide as they can’t form phosphodiester bonds (no OH)
- This generates a set of fragments increasing by 1 base each time
- Fragments separated by size in their separate mixtures, so comparing them shows which the next biggest (and therefore which base next) in the sequence
- Base sequence put together and then inverted to show the original template fragment being tested

Question 8

Why is DNA sequenced?

Answer 8

• Determine exact position of mutation
• Determine type of mutation

Question 9

Endogenous mechanisms causing damage to DNA

Answer 9

• Depurination
• Deamination
• Reactive oxygen attacking purine/pyrimidine rings
• Methylation of cytosines

Question 10

What is depurination?

Answer 10

• Spontaneous break of bond between purine base and the sugar
• Leads to deletion of base or incorrect nucleotide in new strand

Question 11

What is deamination?

Answer 11

• Cytosine deaminates to uracil
• Causes substitution of A into new strand

Question 12

What is methylation of cytosines and what can it cause?

Answer 12

• Cytosine at CpG dinucleotide is methylated
• Spontaneous deamination of 5-methyl-cytosine into thymine

Question 13

Extracellular agents causing damage to DNA

Answer 13

• UV light
• Environmental chemicals
• Ionizing radiation

Question 14

What does UV light do to DNA?

Answer 14

Cross-links adjacent thymines to form a dimer

Question 15

What do environmental chemicals do to DNA?

Answer 15

Cause DNA breaks or chromosome aneuploidy

Question 16

What does ionizing radiation do to DNA?

Answer 16

Cause breaks in DNA

Question 17

2 types of mutation relating to location (and if they’re heritable or not)

Answer 17

• Germline mutation (heritable)
• Somatic mutation (non-heritable)

Question 18

What are the stages of checking in order to remove mutations from replication?

Answer 18

• DNA polymerase adds a base, checks it and moves on
• The DNA mismatch repair system fixes most mutations missed by DNA polymerase

Question 19

How does the mismatch repair system work?

Answer 19

• Mismatch is identified
• Newly synthesised strand is excised by a mismatch repair protein
• Gap left behind is refilled by DNA polymerase and ligase

Question 20

How are DNA double stranded breaks fixed?

Answer 20

• Broken at same point on both strands = DNA on homologous chromosome used as template
• Broken at different points on strands = extra bits removed then ends joined

Question 21

What is a splice-site mutation?

Answer 21

A single base substitution at the end of an exon where splicing was meant to take place

Question 22

What is the normal method of splicing and how does a splice-site mutation affect this?

Answer 22

• Normally each exon has a splice donor site and splice acceptor site with the DNA in between being removed
• Due to mutation, one of these sites is removed
• Either an exon is removed or an intron is incorporated

Question 23

How do copy number variants lead to large deletions or insertions?

Answer 23

• Areas of repeats at different loci on different chromosomes line up and crossing over happens here
• One chromosome then has more DNA and one has less than they usually have