Mutation and Genetic Variation
For every gene, there are many different alleles
- alleles are versions of the same gene that differ in their
DNA base sequence
- some alleles differ in the protein product of the gene;
others do not (more on this to follow)
- your combination of alleles is unique, and makes you you
Alleles are generated by mutations, which are changes in the
DNA base sequence
Evolution and Alleles
Define evolution: genetic change in a population over time
But what does “genetic change” mean, exactly?
1. Mutation can create new alleles that were not previously
present in that population
2. The frequency of alleles may change
Today we will consider mutation in some detail; we will then
then discuss in detail the forces that can change allele
frequencies in natural populations
Mutation and Genetic Variation
Changes in the DNA
sequence create new
alleles in every
generation
The first mutation that was characterized at the molecular
level (that is, at the DNA sequence level) was the sickle-cell
anemia allele of the hemoglobin gene
In people afflicted with the disease, a mutation was found at
amino acid #6 of the 146-amino acid hemoglobin protein
The is an example of a point mutation
=
1 of 2 sources:
(a) random error during DNA replication
(b) error in the repair of damaged DNA
-- chemical mutagens, free radicals, radiation
Errors during DNA replication
DNA polymerase occasionally (but rarely) adds the wrong
nucleotide during DNA replication
The mutation is
a transition if..
-
Purines
-
Pyrimidines
Errors during DNA replication
DNA polymerase occasionally (but rarely) adds the wrong
nucleotide during DNA replication
If one type gets
swapped for the
other type, it’s a
transversion
Transversions
only show up in
DNA sequences
half as often as
transitions
Purines
Pyrimidines
Why are transition mutations more common?
inserted
guanine
Transversions stick two
non-complimentary bases
against each other
- steric clash disrupts the
shape of the DNA helix,
making a bulge
- the bulge in the DNA is
more often noticed and
fixed by repair enzymes
2nd source of mutations: error in the repair of damaged DNA
(1) chemical mutagens
- can intercalate (stick into) the DNA helix, distorting
the shape and causing excision (cutting out) of bases
(2) free radicals
- uncharged oxygen atoms or OH molecules
that have single electrons
- extremely oxidizing (electron-hungry)
- damage DNA bases
(3) radiation
- UV from sunlight can cause thymine dimers, which
need to be excised (= cut out)
Mutations may, or may not, change the amino acid sequence
glutamine
-C-A-A-
-C-A-Gglutamine
Mutations that change the
corresponding mRNA codon
to another codon for the same
amino acid are called
synonymous substitutions
(“equivalent”)
-- also called silent substitutions
-- common when the change is at the 3rd position
Note:
Mutations may, or may not, change the amino acid sequence
glutamine
-C-A-A-
- synonymous substitutions usually do not
affect the phenotype of the organism
- assumed to be neutral, meaning they
have no effect on fitness
-C-A-Gglutamine
fitness = how much an individual
contributes to the gene pool of
the next generation
Neutral changes have no adaptive value to the organism
Mutations may, or may not, change the amino acid sequence
glutamine
-C-A-A-
HOWEVER – “silent: changes can affect
the phenotype if they introduce a rarely
used codon
Cells make fewer tRNAs for rare codons
-C-A-Gglutamine
Introduction of a rare codon into a highly
transcribed gene can slow translation of
the mRNA by the ribosome (Shields et al. 1988)
If ribosomes stall mid-translation, this can lead to protein
mis-folding that changes the properties of an enzyme
(Kimchi-Sarfaty et al. Science 2007)
So-called “translational selection” can act against silent changes,
which are therefore not truly “neutral” as we often assume
Mutations may, or may not, change the amino acid sequence
glutamine
glutamine
-C-A-A-
-C-A-A-
-C-A-G-
-C-A-C-
glutamine
histidine
Mutations that change the
corresponding mRNA codon
to a codon for a different
amino acid are called
non-synonymous substitutions
(“non-equivalent”)
-- these mutations alter the protein product of the gene
--
Mutations may, or may not, change the amino acid sequence
glutamine
glutamine non-synonymous substitutions
-C-A-A-
-C-A-A-
-C-A-G-
-C-A-C-
glutamine
histidine
alter the protein product,
so often affect the phenotype
of the organism
-- most changes are neutral or detrimental to the organism
-- if the change is adaptive under some circumstance, then
the mutation may lead to a higher fitness
(example: sickle-cell allele and malaria)
Loss-of-function mutations
normal
1-Base
insertion
DNA sequence
ACA-ATG-GTA-CGA
Protein sequence
Cys-Tyr-His-Ala
ACA-GAT-GGT-ACG
Cys-Leu-Pro-Val
Frameshift mutations change all subsequent amino acids,
usually making a dysfunctional protein
Loss-of-function mutations
DNA sequence
ACA-ATG-GTA-CGA
Protein sequence
Cys-Tyr-His-Ala
1-Base
insertion
ACA-GAT-GGT-ACG
Cys-Leu-Pro-Val
1-Base
deletion
ACA-TGG-TAC-GA
Cys-Tyr-Met-Leu
normal
Frameshift mutations change all subsequent amino acids,
usually making a dysfunctional protein
- some viruses rely on frameshifts to make a 2nd functional protein
for “genomic efficiency” (minimize amount of DNA needed)
Loss-of-function mutations
DNA sequence
ACA-ATG-GTA-CGA
Protein sequence
Cys-Tyr-His-Ala
1-Base
insertion
ACA-GAT-GGT-ACG
Cys-Leu-Pro-Val
1-Base
deletion
ACA-TGG-TAC-GA
Cys-Tyr-Met-Leu
normal
Mutation
ACA-ATT-GTA-CGA
to a stop codon
Cys
Loss-of-function mutations
normal
DNA sequence
ACA-ATG-GTA-CGA
Protein sequence
Cys-Tyr-His-Ala
AGG-GGG-CTA
Insertion
ACA-AGG-GGG-CTA-ATG Cys-Ser-Pro-Asp-Tyr
- caused by a transposable element, or “jumping gene”
- transposons inactivate the gene they disrupt, sometimes only
temporarily; they may hop back out at a later date, restoring the
correct coding sequence
- many genomes are littered with transposons or “defunct” former
transposable sequences
Mutation rates vary…
(1) among different species
(2) between the sexes
(3) among individuals
(4) among genes
- generally 10-5 to 10-8 mutations per cell division
- can vary between individuals, depending on their alleles for:
(a) DNA polymerase (some less accurate than others)
(b) DNA repair enzymes (some less efficient than others)

Linkage
When alleles are close together on a chromosome we call them
linked, because crossing over is unlikely to separate them
A
BC
a
BC
a
b c
A
bc
Crossing over between A and B Alleles will tend to get separated
is very likely to happen, so these by recombination during meiosis
loci are not linked
Linkage
When alleles are close together on a chromosome we call them
linked, because crossing over is unlikely to separate them
A
BC
A
BC
a
b c
a
bc
Crossing over between A and B
is very likely to happen, so these
loci are not linked
Alleles will tend to get separated
by recombination during meiosis
Chromosome rearrangements: Inversions
When 2 double-stranded breaks occur in a chromosome, the part
in between the breaks may flip around and get re-inserted
This results in an inversion, where the gene order is reversed
between the break points relative to the normal chromosome
DNA breaks
reversal of
gene order
Inversions 2
Inverted regions cannot line up properly with the homologous
chromosome during meiosis
A
BC D E F G
a
b c f
e d g
(1) regions will not align during synapsis, when homologs pair
(2) crossing over would cause loss of part of the chromosome
-so(3) alleles f, e and d are now linked, and will be transmitted to
offspring as one big “supergene”
Inversions 3
In natural populations, inversions are very common
Studies on the fruit fly Drosophila have shown that natural selection
favors certain inversions:
- inversions that link alleles for small body size together are
favored in hot, dry areas
- inversions that link alleles for large body size together are
favored in cold, wet climates

Gene Duplication 1
Mutation can create new alleles, but how do you get new genes?
Mistakes during meiosis can result in unequal crossing over,
when a daughter chromosome inherits duplicated regions
of a chromosome
A
B C
A
B C
for instance,
crossing over between two transposons
Gene Duplication 1
Mutation can create new alleles, but how do you get new genes?
Mistakes during meiosis can result in unequal crossing over,
when a daughter chromosome inherits duplicated regions
of a chromosome
A
B C
A
A
B C
B B C
Gene Duplication 2
The viral enzyme reverse transcriptase can create gene copies
from mRNA transcripts, which then get inserted back onto a
chromosome
A
B C
reverse
transcriptase
mRNA
transcript for
the A gene
DNA copy of
the A gene
Gene Duplication 2
The viral enzyme reverse transcriptase can create gene copies
from mRNA transcripts, which then get inserted back onto a
chromosome
A
B C
reverse
transcriptase
mRNA
transcript for
the A gene
inserts anywhere on any
chromosome
DNA copy of
the A gene
Gene Duplication 2
The viral enzyme reverse transcriptase can create gene copies
from mRNA transcripts, which then get inserted back onto a
chromosome
A
B C
=
A
B C A
reverse
transcriptase
mRNA
transcript for
the A gene
DNA copy of
the A gene
- random accidents can thus create functional copies of genes
Gene Duplication 3: polyploidy
In plants, the production of diploid gametes during meiosis can
result in tetraploid offspring
- that is, offspring with 4 copies of each chromosome
Tetraploid plants cannot produce viable offspring with
normal 2N plants, but can breed successfully with other
tetraploid individuals
- new species can thus come into existence, as a population
of plants that only interbreeds with itself
“Directed mutation” controversy
First confirmation that mutation precedes adaptation came from
Luria & Delbruck’s 1943 experiment, called the
Fluctuation Test
- tested two alternative hypotheses about
how E. coli became resistant to a virus:
(1) “acquired hereditary immunity” hypothesis:
each E. coli cell has a small chance of surviving
a virus, but survivors get “acquired immunity”
that can be passed to their offspring
- exposure to virus induces resistance
(= adaptation)
-
“Directed mutation” controversy
First confirmation that mutation precedes adaptation came from
Luria & Delbruck’s 1943 experiment, called the
Fluctuation Test
- tested two alternative hypotheses about
how E. coli became resistant to a virus:
(1) “acquired hereditary immunity” hypothesis:
each E. coli cell has a small chance of surviving
a virus, but survivors get “acquired immunity”
that can be passed to their offspring
- prediction: in each tube, there will be a few
cells that get resistance, but only after they
are plated out with the virus
“Directed mutation” controversy
(2) “mutation” hypothesis: in some tubes, a random
mutation will happen early on & get passed to
most offspring, prior to virus exposure
- will give rise to occasional “jackpot cultures”
that luckily got the resistance mutation early
in their family tree
- prediction: there will be wildly different #’s of
resistant colonies from different starting cultures,
depending on how early the mutation occurred
“Directed mutation” controversy
Lederberg & Lederberg (1952) – replica plating experiment
Put a cloth over a “master plate” covered in bacteria; blotted it onto
many replica plates coated with virus
Virus-resistant colonies appeared in
the same place on each replica plate
master
plate
- thus, those colonies grew from
resistant cells that were already
present on the master plate
- resistance was not induced by the
virus...
“Directed mutation” controversy
Cairns et al. (1988) argued in the journal Nature these experiments
didn’t give cells a fair chance to “try to mutate” to survive, because
the virus killed cells that were not already resistant
Took cells that had a frameshift mutation in their lac gene, needed
for growth on medium where lactose is the only energy source
- normally, cells mutate from lac- to lac+ at a frequency of 1×10-8
3×108 laccells growing
in tube of
normal media
lactose plates – 3 colonies grow
(they’ve mutated to lac+)
no lactose – no colonies grow
“Directed mutation” controversy
Cairns et al. (1988) found that after mutants sat on lactose-containing
plates for 2-4 days, many more colonies suddenly started to appear
- only showed up if cells were sitting on lactose plates
- were 10-100 times more frequent than normal
- no increase in rate of mutation from ValS
ValR, so there
was no across-the-board increase in mutation rate
Day 1
Day 3
Day 5
lactose plates
no lactose
“Directed mutation” controversy
Cairns et al. (1988) proposed that cells were somehow sensing that
they needed to mutate their lactose-metabolizing gene to survive..
... were able to “choose which mutations will occur” by directing
mutation towards the broken lac gene
- from 1988-1994, at least 5 papers argued for “directed mutation”
when cells were grown on a nutrient they couldn’t use
This directly contradicts the fundamental premise of Darwinian
evolution by natural selection, variation precedes adaptation
- make sure you understand why this was so controversial!!
“Directed mutation” controversy
Anderssen et al. (1998) published the following model in Science to
explain the observations of Cairns:
(1) lac- frameshift mutation still produces 1% of b-gal enzyme
encoded by wildtype lac+ allele
(2) Cairns’ expt was done with the lac- mutation on a plasmid,
which could increase the odds of gene duplication events
(3) if a cell acquires a duplication of lac- allele, it can grow a
little more because of the small amount of enzyme produced;
the more duplications that occur, the more growth is possible
lac-
lac- lac-
lac- lac- lac- lac-
“Directed mutation” controversy
Anderssen et al. (1998) published the following model in Science to
explain the observations of Cairns:
(4) this leads to invisible “micro-colonies” of slow-growing cells
with up to 100 copies of the broken gene....
meaning, 100 targets for random mutation
- you’re 100x more likely to get a reverse-mutation to lac+
when you have 100 copies of the broken gene lying around !
(5) once the reverse-mutation to lac+ occurs, the extra broken
copies of lac- are lost and the lac+ cell rapidly forms a colony
Under this model, all mutations happen by chance; duplications help
the cell by producing more b-gal enzyme, which leads to growth &
increases the odds of getting a lucky laclac+ mutation
“Directed mutation” controversy
Anderssen et al. (1998) confirmed experimentally the predictions of
their model, which the “directed mutation” hypothesis did not predict
- residual b-gal enzyme activity was necessary in supposedly
“lac-” mutants, or no late-appearing lac+ colonies ever appeared
- early lac+ cells had up to 50 copies of lac- allele
- the phenomenon required recombination, which is not needed
to fix a frameshift mutation, but is needed for gene duplication
This story is important because it shows that you can challenge the
basic theory of evolution through scientific experimentation –
but careful experiments done for the last 150 years have, so far,
all supported the premise of Darwinian evolution by selection
Bottom line
Natural selection is the sole evolutionary force responsible for
the adaptation of organisms to their environment
Mutation is random with respect to the adaptive “needs” of
individual organisms; you cannot “try” to mutate to survive
Variation precedes adaptation