Lecture 38. Mutagenesis and Mutations

Question 1

How can mutants be created?

Answer 1

Mutants can be spontaneous or a result of being exposed to mutagens

Question 2

What can bacteria produce to protect from radiation damage?

Answer 2

Melanin and other molecules

Question 3

How often does E coli DNA make a substitution?

Answer 3

~1 in every 10⁷ bases

Question 4

How long is the E. coli genome?

Answer 4

5.4 x10⁶ base pairs

Question 5

Spontaneous mutation examples

Answer 5

Replication errors - Wrong base inserted by DNA polymerase
Wrong base inserted by DNA polymerase - Tautomers forms have different H-bonding pattern
Base pair slipping - Repeat nucleotides can lead to frameshift mutations

Question 6

What are mutagens?

Answer 6

Chemical or physical agents causing damage to DNA

Question 7

Examples of mutagens

Answer 7

Nitrous acid, Reactive oxygen species, Alkylating agents, Intercalating agents and UV light

Question 8

Intercalating agents

Answer 8

Usually have flat multiple ring structures
Binds between base pairs
Distorts the helix
Can also lead to frameshift mutations

Question 9

Intercalating agent example

Answer 9

Ethidium bromide

Question 10

What is a point mutation?

Answer 10

A change to one base pair

Question 11

What is a transition substitution?

Answer 11

A change from purine to purine or pyrimidine to pyrimidine

Question 12

What is a transversion substitution?

Answer 12

A change from a purine to a pyrimidine and vice versa

Question 13

What could result in segregation of mismatched base pairs?

Answer 13

Error in replication, tautamerisation or damage such as deamination

Question 14

Consequences of point mutations in non-coding DNA

Answer 14

May have no consequence at all, but non-coding DNA can have other functions e.g. promoters and other regulatory sequences

Question 15

Consequences of point mutations in promoters

Answer 15

Can affect transcription (up or down) but may have no consequence at all

Question 16

Consequences of point mutations in genes under control of promoter

Answer 16

Protein coding part: Depends
Can affect sequence or regulation of translation

Question 17

What are the three kinds of mutations caused by substitutions?

Answer 17

Silent, Missense and Nonsense

Question 18

Silent mutation

Answer 18

Nothing happens. Genotype changed but no change in phenotype

Question 19

Missense mutation

Answer 19

Often nothing happens but can be detrimental. Genotype changed and phenotype may be changed

Question 20

What mutations occur due to insertions and deletions?

Answer 20

Frameshift mutations

Question 21

Nonsense and frameshift mutations

Answer 21

Usually detrimental but can be tolerated close to C-terminus. Genotype and phenotype changed

Question 22

Deletions

Answer 22

Can remove kilobases and lose several genes or be just a single base pair
Can result in frameshift mutations one or a few bases are lost within a gene affecting coding or regulation

Question 23

Inversions

Answer 23

Can flip kilobases and several genes or be much shorter
Can disrupt genes or just invert them

Question 24

Tandem repeats

Answer 24

Part of genome is duplicated
Can lead to overproduction of proteins encoded in the duplicated region
Also leads to evolution of proteins

Question 25

Transposons

Answer 25

Transposons are nucleotide sequences that are able to move themselves around
Can disrupt genes

Question 26

Reversions of mutations

Answer 26

Reversion is a point mutation resulting in restoration of the original sequence

Question 27

Supressor mutations

Answer 27

A second mutation happens that results in the original phenotype being restored

Question 28

Intragenic suppression

Answer 28

A second mutation occurs in a different gene to the one in which the first mutation occurs.
The effect is to suppress the phenotype of the first mutation

Question 29

Nonsense suppression

Answer 29

Strains with nonsense suppressors are normally sick because cell tends to translate past normal stop codons and produce longer proteins than normal.
These may not function correctly because they may fold incorrectly.

Question 30

Making histadine auxotroph

Answer 30

1. Expose to mutagen
2. Grow in complex medium to allow expression of phenotype
3. Penicillin enrichment: Change medium to minimal medium, add penicillin, penicillin will kill any growing bacteria, auxotrophs will not grow
4. Plate on minimal medium + histidine and grow

Question 31

What is phenotype lag?

Answer 31

Phenotype is not seen for several generations

Question 32

What is cross feeding?

Answer 32

Metabolites immediately before block will accumulate If metabolite can diffuse from cell another bacteria can take it up. Both mutants can now grow but dependent on each other

Question 33

What is the Ames test used for?

Answer 33

Used to identify chemicals that are mutagenic and therefore carcinogenic. Assumes that if chemical is mutagenic to bacteria it is also mutagenic to humans

Question 34

What bacteria does the Ames test use?

Answer 34

A His auxotroph of Salmonella typhimurium

Question 35

What do the results of an Ames test show?

Answer 35

If chemical is mutagenic then lots of reversions
If not mutagenic only a few reversions