1

Round and wrinkled peasRound and wrinkled peas

Fig. A, page 22

2

Mutations: Primary tools of genetic Mutations: Primary tools of genetic analysisanalysis

Mutations are heritable changes in base Mutations are heritable changes in base sequences that modify the information sequences that modify the information content of DNAcontent of DNA Forward mutation – changes wild-type to Forward mutation – changes wild-type to

different alleledifferent allele Reverse mutation – causes novel mutation to Reverse mutation – causes novel mutation to

revert back to wild-type (reversion)revert back to wild-type (reversion)

3

General observations of mutation General observations of mutation ratesrates

Mutations affecting phenotype occur very Mutations affecting phenotype occur very rarelyrarely

Different genes mutate at different ratesDifferent genes mutate at different rates Rate of forward mutation is almost always Rate of forward mutation is almost always

higher than rate of reverse mutationhigher than rate of reverse mutation

4

Fig. 7.3

Mutations that cause a phenotypic Mutations that cause a phenotypic change are very rarechange are very rare

5

Classification of mutations by effect Classification of mutations by effect on DNA moleculeon DNA molecule

Substitution – base is replaced by one of the other Substitution – base is replaced by one of the other three basesthree bases

Deletion – block of one or more DNA pairs is lostDeletion – block of one or more DNA pairs is lost Insertion – block of one or more DNA pairs is Insertion – block of one or more DNA pairs is

addedadded Inversion – 180 degree rotation of piece of DNAInversion – 180 degree rotation of piece of DNA Reciprocal translocation – parts of Reciprocal translocation – parts of

nonhomologous chromosomes change placesnonhomologous chromosomes change places Chromosomal rearrangements – affect many Chromosomal rearrangements – affect many

genes at one timegenes at one time

6Fig. 7.2

7

Unequal crossing over creates one Unequal crossing over creates one homologous chromosome with a duplication homologous chromosome with a duplication

and the other with a deletionand the other with a deletion

7.10 a

8

Transposable elements move around the genome Transposable elements move around the genome and are not susceptible to excision or mismatch and are not susceptible to excision or mismatch

repairrepair

Fig. 7.10 e

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Trinucleotide instability causes Trinucleotide instability causes mutationsmutations

FMR-1 genes in FMR-1 genes in unaffected people unaffected people have fewer than have fewer than 50 CGG repeats. 50 CGG repeats.

Unstable Unstable premutation premutation alleles have alleles have between 50 and between 50 and 200 repeats.200 repeats.

Disease causing Disease causing alleles have > 200 alleles have > 200 CGG repeats.CGG repeats.

Fig. B(1) Genetics and Society

10

Trinucleotide repeat in people with Trinucleotide repeat in people with fragile X syndromefragile X syndrome

Fig. A, B(2) Genetics and Society

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Are mutations spontaneous or Are mutations spontaneous or induced?induced?

Most mutations are spontaneous.Most mutations are spontaneous. Luria and Delbruck fluctuation experiments and replica Luria and Delbruck fluctuation experiments and replica

plating - simple ways to tell if mutations are spontaneous plating - simple ways to tell if mutations are spontaneous or if they are induced by a mutagenic agentor if they are induced by a mutagenic agent

12Fig. 7.4

13

Interpretation of Luria-Delbruck Interpretation of Luria-Delbruck fluctuation experimentsfluctuation experiments

Bacterial resistance arises from mutations Bacterial resistance arises from mutations that exist before exposure to bacteriocidethat exist before exposure to bacteriocide

The bacteriocide is a selective agent killing The bacteriocide is a selective agent killing the nonresistant cells, allowing only the the nonresistant cells, allowing only the preexisting mutant cells to survive.preexisting mutant cells to survive.

Mutations do not arise as a direct response Mutations do not arise as a direct response to environmental changeto environmental change

Mutations occur randomly at any timeMutations occur randomly at any time

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Replica plating verifies preexisting Replica plating verifies preexisting mutationsmutations

Fig. 7.5 a

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Fig. 7.5b

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Chemical and physical agents cause Chemical and physical agents cause mutationsmutations

Hydrolysis of a purine Hydrolysis of a purine base, A or G occurs 1000 base, A or G occurs 1000 times an hour in every celltimes an hour in every cell

Deamination removes –NHDeamination removes –NH22 group. Can change C to U, group. Can change C to U, inducing a substitution to an inducing a substitution to an A-T base pair after replicationA-T base pair after replication

Fig. 7.6 a,b

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X rays break the X rays break the DNA backboneDNA backbone

UV light produces UV light produces thymine dimersthymine dimers

Fig. 7.6 c, d

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Oxidation from free radicals formed by irradiation damages individual bases

Fig. 7.6 e

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Repair enzymes fix errors created by Repair enzymes fix errors created by mutationmutation

Excision repair Excision repair enzymes enzymes release release damaged damaged regions of regions of DNA. Repair DNA. Repair is then is then completed by completed by DNA DNA polymerase polymerase and DNA ligaseand DNA ligase

Fig. 7.7a

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DNA polymerase proofreadingDNA polymerase proofreading

Fig. 7.8

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Mutagens increase mutation rate Mutagens increase mutation rate using different mechanismsusing different mechanisms

Fig. 7.12a

22

23Fig. 7.12 b

24Fig. 7.12 c

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Consequences of mutationsConsequences of mutations

Germ line mutations – affect the evolution of Germ line mutations – affect the evolution of speciesspecies

Somatic mutations – affect the survival of an Somatic mutations – affect the survival of an individualindividual Cell cycle mutations may lead to cancerCell cycle mutations may lead to cancer

Conditional mutations – produce changes in Conditional mutations – produce changes in phenotype under one set of conditions but not phenotype under one set of conditions but not under anotherunder another Conditional lethal mutationsConditional lethal mutations

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The Ames test The Ames test for carcinogens for carcinogens

using hisusing his-- mutants of mutants of Salmonella Salmonella

typhimuriumtyphimurium

Fig. 7.13

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What mutations tell us about What mutations tell us about gene gene structurestructure

Complementation testing - are two Complementation testing - are two mutations in the same or different genes?mutations in the same or different genes?

Benzer’s experiments - genes are linear Benzer’s experiments - genes are linear sequences of nucleotide pairs sequences of nucleotide pairs

Some regions of chromosomes mutate at a Some regions of chromosomes mutate at a higher rate than others – hot spotshigher rate than others – hot spots

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Complementation testingComplementation testing

Fig. 7.15 a

29

30

Five complementation groups (different genes) for eye color.Recombination mapping demonstrates distance between genes and alleles.

Fig. 7.15 b,c

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Recombination within a gene can Recombination within a gene can generate wild-typegenerate wild-type

Fig. 7.16

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A gene is a linear sequence of A gene is a linear sequence of nucleotide pairsnucleotide pairs

Seymour Benzer mid 1950s – 1960sSeymour Benzer mid 1950s – 1960s If a gene is a linear set of nucleotides, If a gene is a linear set of nucleotides,

recombination between homologous recombination between homologous chromosomes carrying different mutations chromosomes carrying different mutations within the same gene should generate wild-type within the same gene should generate wild-type

T4 phage as an experimental systemT4 phage as an experimental system Can examine a large number of progeny to detect Can examine a large number of progeny to detect

rare mutation eventsrare mutation events Could allow only recombinant phage to proliferate Could allow only recombinant phage to proliferate

while parental phages diedwhile parental phages died

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Benzer’s experimental procedureBenzer’s experimental procedure

Generated 1612 spontaneous point mutations and Generated 1612 spontaneous point mutations and some deletionssome deletions

Mapped location of deletions relative to one Mapped location of deletions relative to one another using recombinationanother using recombination

Found approximate location of individual point Found approximate location of individual point mutations by deletion mappingmutations by deletion mapping

Performed recombination tests between all point Performed recombination tests between all point mutations known to lie in the same small region of mutations known to lie in the same small region of the chromosomethe chromosome

Result – fine structure map of the Result – fine structure map of the rIIrII gene locus gene locus

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Working with T4 phageWorking with T4 phage

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Phenotypic properties of T4 phagePhenotypic properties of T4 phage

Fig. 7.17 b

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Complementation test for mutations Complementation test for mutations in different genesin different genes

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Detecting recombination between Detecting recombination between two mutations in the same genetwo mutations in the same gene

Fig. 7.17 d

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Deletions for rapid mapping of point Deletions for rapid mapping of point mutations to a region of the chromosomemutations to a region of the chromosome

Fig. 7.18 a

39

Recombination Recombination mapping to identify mapping to identify the location of each the location of each

point mutation point mutation within a small within a small

regionregion

Fig. 7.18 b

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Fine structure map of Fine structure map of rIIrII gene gene regionregion

Fig. 7.18 c

41

What mutations tell us about What mutations tell us about gene gene functionfunction

One gene, one enzyme hypothesis - a gene contains One gene, one enzyme hypothesis - a gene contains the information for producing a specific enzymethe information for producing a specific enzyme Beadle and Tatum used Beadle and Tatum used auxotrophicauxotrophic and and prototrophicprototrophic

strains of strains of NeurosporaNeurospora to test hypothesis to test hypothesis Genes specify the identity and order of amino Genes specify the identity and order of amino

acids in a polypeptide chainacids in a polypeptide chain The sequence of amino acids in a protein The sequence of amino acids in a protein

determines its three-dimensional shape and determines its three-dimensional shape and functionfunction

42

Beadle and Tatum – One gene, one Beadle and Tatum – One gene, one enzymeenzyme

1940s – isolated mutants that disrupted synthesis 1940s – isolated mutants that disrupted synthesis of arginineof arginine AuxotrophAuxotroph – needs supplement to grow on minimal – needs supplement to grow on minimal

mediamedia PrototrophPrototroph –needs no supplement –needs no supplement

Recombination analysis - mutations in four Recombination analysis - mutations in four distinct regions of genomedistinct regions of genome

Complementation tests - each of four regions Complementation tests - each of four regions correlated with a different complementation correlated with a different complementation group (4 different genes)group (4 different genes)

43Fig. 7.20 a

44

Fig. 7.20 b

45

Interpretation of Beadle and Tatum Interpretation of Beadle and Tatum experimentsexperiments

Each gene controls the synthesis of an Each gene controls the synthesis of an enzyme involved in catalyzing the enzyme involved in catalyzing the conversion of an intermediate into arginineconversion of an intermediate into arginine

46

Some proteins are multimeric, containing subunits Some proteins are multimeric, containing subunits composed of more than one polypeptidecomposed of more than one polypeptide

Fig. 7.24

47

How do genotypes and phenotypes How do genotypes and phenotypes correlate?correlate?

Alteration of amino acid composition of a Alteration of amino acid composition of a proteinprotein

Alteration of the amount of normal protein Alteration of the amount of normal protein producedproduced

Changes in different amino acids at Changes in different amino acids at different positions have different effectsdifferent positions have different effects

48

Summary of dominance Summary of dominance relationshipsrelationships

Fig. 3.2

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Codominant blood group allelesCodominant blood group alleles

Fig. 3.4b

50

Dominant or recessive alleles depend on the Dominant or recessive alleles depend on the relationship between protein function and relationship between protein function and

phenotypephenotype

Recessive – alleles that produce nonfunctional proteins (loss-of Recessive – alleles that produce nonfunctional proteins (loss-of function)function) Null mutationsNull mutations – no synthesis of protein or promote synthesis of protein – no synthesis of protein or promote synthesis of protein

incapable of carrying out any functionincapable of carrying out any function Hypomorphic mutationsHypomorphic mutations – produce much less protein or a protein with – produce much less protein or a protein with

weak functionweak function

Incomplete dominance – phenotype varies in proportion to Incomplete dominance – phenotype varies in proportion to amount of proteinamount of protein

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Recessive mutationsRecessive mutations

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Incomplete dominanceIncomplete dominance

53

Dominant or recessive alleles depend on the Dominant or recessive alleles depend on the relationship between protein function and relationship between protein function and

phenotypephenotype

Dominant – reflect several different occurrences (generally gain-Dominant – reflect several different occurrences (generally gain-of-function) of-function)

Hypermorphic mutationsHypermorphic mutations – produces more protein or same amount of a – produces more protein or same amount of a more effective proteinmore effective proteinHaploinsufficiency Haploinsufficiency –one allele does not provide enough protein–one allele does not provide enough proteinDominant negativeDominant negative – mutant protein negatively affects the normal – mutant protein negatively affects the normal productproductNeomorphic mutationsNeomorphic mutations – generate a novel phenotype – generate a novel phenotype

54

Dominant mutationsDominant mutations