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Sources of spontaneous mutations:
- molecular decay
- errors in replication
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Types of point mutations
Transition mutations = A <-> G or C <-> T (purine to purine or pyrimidine to pyrimidine)
Transversion mutations = purine to pyrimidine or vice versa
These can arise from mispairing, insertion of base analogs, or chemical mutagens (e.g., nitrous acid, hydroxylamine, and alkylating agents)
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What is an indel type of mutation?
- insertion or deletion event
- adds or removes 1+ nucleotides
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What is a rearrangement sort of mutation?
- interchange 2+ pieces of DNA
- (inversion is also rearrangement, when 1+ fragments change direction)
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What is an inversion type of mutation?
when 1+ fragments of DNA change directions
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Which method(s) could you use to create indel mutations?
E. All of the above
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What types of point mutations occur in gene coding regions?
- Silent/sense
- missense
- nonsense
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Define sense (or silent) mutation
- a change in a DNA sequence that results in a new codon coding for the same amino acid
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Define missense mutation:
- a change in DNA that results in a different codon and in the amino acid substitution in the corresponding protein
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Define nonsense mutation:
- a change in DNA that results in a premature stop codon
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What would you need to do to regrow telomeres in a senescence cell culture?
reactivate telomerase genes. Can do by reintroducing the gene with a viral vector, and having it reintroduced in the organism under stronger promoters
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three components of telomere genes?
- scaffolding/structural component
- reverse transcriptase protein (TERT)
- large RNA component (rna template for telomerase)
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Why do different groups have different rates of mutation?
- This can be an effect of the size of genome footprint.
- The larger the size of the gene pathway, the more opportunity for mutations
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Describe the tool here:
- "Replicate plater"
- velvet fabric on a chunk of plastic that enables duplication of colonies on many plates
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Explain how you might identify auto-tropic mutants in bacteria
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Describe a mutation hotspot
a chromosomal region where mutations occur at a greater frequency than elsewhere in the genome
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How are mutational hot spots thought to arrise?
- They are thought to be caused by some inherent instability
- (like unequal crossing over or predisposition to nucleotide substitutions)
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What is an example gene with mutation hotspots within patients with colorectal tumors?
- p53 (key tumor suppressor gene)
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How might mutations arise?
- 1. Mistakes during DNA replication
- 2. Spontaneous mutations
- 3. Induced mutations
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Describe some mistakes during DNA replication
- changes in H-bonding properties of bases
- incorporation of different base analogs
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Describe how spontaneous mutations might arise:
- deanimation
- depurination/depyrimidination
- oxygen radical damage
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Describe what Induced mutations caused by environmental factors may give rise to mutations:
- Induced mutations by (chemicals, UV radiation, etc)
- or biological (viruses, transposons, etc) agentes
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Define: tautomer
molecular isomers that differ in positions of atoms in bonds
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What are the three mechanims that change the H-bonding properties of the base, so that it is incorporated incorrectly
- 1) Tautomerization of adenine
- 2) Syn vs. anti-conformation
- 3) H20 acting as a bridge
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Describe what happens during tautomerization of adenine:
- The H-boning properties of the base are changed
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Describe what happens when hte glycosidic bond is flipped (syn vs anti conformation)
- The H-boning properties of the base are changed
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Describe what happens when H2O acts as a brige
- The H-boning properties of the base are changed
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What is the prominent conformation of the glycosidic bond (syn vs anti)
- anti
- (can make an A pair with a G)
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How can a syn vs anti base conformation occur in nucleotides?
- The base can rotate around the glycosidic bond in respect to the sugar
- In naturally occurring nucleotide, the anti conformation predominates
- (can make an A pair with a G)
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Describe how mutations can be caused by base analogs during DNA replication
- base analogs that are similar enough to nitrogen bases to get accidentally incorporated
- ex: 5-BU is a thymine analog tha gets incorporated during replication opposite of A, but frequently pairs with G
- T-A --> C-G
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Describe a scenario where base analog incorporation during DNA replication can cause mutation
- 5-BU is a thymine analog that gets incorporated during replication opposite of A, but frequently pairs with G
- T-A --> C-G
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Example of Oxidative deanimation
- conversion of methylcytosine to thyminine
- if the methylcytosine is deanimated to T, there is no repair system that can recognize it to remove it. This is a transition mutation
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Why is oxidative deamination of methycytosine to thymine dangerous?
- Methylcytosine naturally occurs in the human genome.
- if the meC is deanimated to T, there is no repair system to recognize the change
- meC-G--> T-A
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Describe how spontaneous base loss may occur
- Within a mammalian cell, several thousand purines/pyrimidines are lost per cell per day
- The loss creates an apurinic/apyrimidinic (AP) site
- also known as abasic site
- During replication, a random base can be inserted across from abasic site
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What is oxygen radical damage?
Reactive oxygen species that can cause oxidative damage to DNA, as well as to precursors of DNA, resulting in mutations
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What ultimately happens when an oxygen radical induces damage?
- Some base modifications result to mispairing
- some fail to pair and are removed by DNA repair machinery
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What can oxidative damage to DNA lead to?
mutations and cancer
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Three ways that mutations arise:
- 1. Mistakes during DNA replication
- 2. Spontaneous mutations
- 3. Oxygen radical damage
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Examples of mutations arising during DNA replication:
- due to changes in H-bonding properties of bases
- due to incorporation of base analogs
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Examples of mutations arising from spontaneous mutations:
- deanimation
- depurination/depyrimidination
- oxygen radical exchange
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Examples of how mutations arise via the enviroment:
- chemicals
- UV radiation
- viruses
- transposons
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