Lecture 3: Molecular Basis of Genetic Polymorphisms and their Detection

Question 1

what is the main source of allelic variation?

Answer 1

mutations

Question 2

define mutation

Answer 2

the process by which genes change from one allelic form to another. the creation of entirely new alleles can occur.

Question 3

describe how mutations arise

Answer 3

• genes mutate randomly, at any time and in any cell of an organism
• mutations can arise spontaneously during normal replication or can be induced by a mutagen

Question 4

how are mutations transmitted to progeny?

Answer 4

• only mutations in gremlin cells can be transmitted to progeny
• somatic mutations cannot be transmitted

Question 5

how do inherited mutations appear in populations of individuals?

Answer 5

as alleles

Question 6

draw a flowchart describing the inheritance of mutations

Answer 6

slide 6

Question 7

define allele frequency

Answer 7

the percentage of the total number of gene copies represented in one allele

Question 8

how can allele frequency be calculated?

Answer 8

by dividing the number of times the allele of interest is observed in a population by the total number of copies of all the alleles at that particular genetic locus in the population

Question 9

wild-type allele

Answer 9

allele whose frequency is greater than or equal to 1%

Question 10

mutant allele

Answer 10

allele whose frequency is less than 1%

Question 11

monomorphic gene

Answer 11

a gene with only one wild-type allele

Question 12

polymorphic gene

Answer 12

a gene with more than one wild-type allele

Question 13

forward mutation

Answer 13

changes wild-type allele to a different allele

Question 14

reverse mutation

Answer 14

causes novel mutation to revert back to wild-type allele (reversion)

Question 15

define a mutagen

Answer 15

a mutation inducer, eg UV light, certain chemicals, etc.

Question 16

mutations affecting phenotype occur

Answer 16

very rarely

Question 17

do all genes mutate at the same rate?

Answer 17

no; different genes mutate at different rates.
- mutation rate varies from 1 in 1000 to 1 in 1,000,000,000 per gene per gamete

Question 18

which rate is higher; rate of forward or rate of reverse mutation?

Answer 18

rate of forward mutation is almost always higher than rate of reverse mutation

Question 19

6 types of mutations (classified by effect on DNA molecule)

Answer 19

• substitution
• deletion
• insertion
• inversion
• reciprocal translocation
• chromosomal rearrangements

Question 20

substitution

Answer 20

base is replaced by one of the three other bases

Question 21

a substitution is a type of

Answer 21

point mutation

Question 22

point mutation

Answer 22

changes in a single nucleotide base within a DNA sequence

Question 23

deletion

Answer 23

block of one or more DNA pairs is lost

Question 24

insertion

Answer 24

block of one or more DNA pairs is added

Question 25

inversion

Answer 25

rotation of piece of DNA

Question 26

reciprocal translocation

Answer 26

parts of non homologous chromosomes change places

Question 27

chromosomal rearrangements

Answer 27

a change in the structure of a chromosome, often involving deletions, duplications, inversions, or translocations; affect many genes at one time

Question 28

single nucleotide polymorphism (SNP)

Answer 28

a variation in a single base pair in a DNA sequence.

Question 29

SNPs are

Answer 29

alleles

Question 30

define a polymorphism

Answer 30

- detectable change (difference) in a given locus/gene - this is what makes an allele an allele

Question 31

how are Mendel's traits encoded?

Answer 31

in DNA: allelic differences at the DNA level can influence mRNA expression and/or protein function, and thus the phenotype DNA -> mRNA -> protein -> organismal traits

Question 32

two ways in which mutations may impact gene expression

Answer 32

1. mutation in exon -> altered transcription -> altered translation -> different amino acid -> folded protein with altered function 2. mutation in promoter sequence -> no transcription -> no RNA -> no function

Question 33

null mutation

Answer 33

completely abolishes the function of a gene

Question 34

leaky/hypomorphic mutation

Answer 34

mutated gene product retains some, but not all, of its normal function

Question 35

silent/synonymous mutation

Answer 35

doesn't alter the amino acid sequence of the protein it encodes

Question 36

give an example of how different types of mutations may result in the same phenotypic consequences

Answer 36

Coding DNA Sequence (CDS) region mutation -> RNA -> protein -> no function Promoter mutant allele -> no RNA -> no protein -> no function

Question 37

is there dominance/recessiveness at the DNA level? why/why not?

Answer 37

- there is no dominance and recessiveness at the DNA level, since it is the underlying genotypic basis of inheritance (codominance). - dominance/recessiveness can only be assessed at the phenotypic level

Question 38

describe the gene basis behind Mendel's pea shape

Answer 38

Starch branching enzyme 1 R: Sbe1 r: no Sbe1 Case 1: RR, Rr -> Sbe1 produced - this causes amylose to be converted into amylopectin, which is a branched starch required to give the round pea shape Case 2: rr -> no Sbe1 produced - amylose is not converted into amylopectin, thus giving a wrinkled pea shape

Question 39

describe the gene basis behind Mendel's stem length

Answer 39

GA = gibberellin acid plant growth hormone gibberellin (GA) 3β-hydroxylase is an enzyme required for the conversion of GA20 (inactive form) into GA1 (bioactive form) Case 1: dominant allele LE -> GA 3β-hydroxylase produced and there is rapid conversion of GA20 into GA1, leading to long stems Case 2: recessive allele le -> GA 3β-hydroxylase produced and there is very low conversion of GA20 into GA1, leading to short stems this is an example of how a SNP change in one amino acid disrupts the activity of enzymes

Question 40

describe PKU

Answer 40

- normally, phenylalanine (found in food) is usually converted into tyrosine by the enzyme phenylalanine hydroxylase (PAH) - in people with PKU, mutations in both exons and introns for the PAH gene can inactivate the gene - this causes inability to convert phenylalanine into tyrosine, instead converting it into phenylpyruvic acid - build-up of phenylpyruvic acid can interfere with nervous system development, so must be caught early in order to change diet accordingly

Question 41

BRCA1

Answer 41

- a tumour-suppressor gene involved in repairing DNA damage - mutations in this gene interfere with DNA repair, leading to cancer risk (breast + ovarian in women, breast + prostate in men) - hundreds of mutations in BRCA1 have been found that increase the risk of breast cancer - there is a 12% risk in the general public, 60% risk in those with harmful BRCA1 mutations

Question 42

draw a table for pea genotype and amount of functional, starch-producing protein

Answer 42

25

Question 43

is haplosufficiency or haploinsufficiency more common?

Answer 43

for many genes, 50% of the protein product is sufficient to give a wild-type phenotype (haplosufficient)

Question 44

haplosufficiency

Answer 44

- A single copy of a functional allele (wild-type) (=50% of the protein product) is sufficient to produce a normal phenotype. - If one wild-type allele is present, the individual will typically have a normal phenotype, even if the other allele is a mutant.

Question 45

haploinsufficiency

Answer 45

- A single copy of a functional allele (wild-type) is not sufficient to produce a normal phenotype. - Individuals with one wild-type and one mutant allele may show a mutant phenotype because the remaining functional allele cannot compensate for the loss of the other.

Question 46

what is the similarity between haploinsufficiency and dominant negative situations?

Answer 46

Haploinsufficiency and dominant negative effects are two different ways in which a single mutated gene can cause a dominant phenotype.

Question 47

describe the difference between haploinsufficiency and dominant negative situations

Answer 47

1. Haploinsufficiency - When a single functional copy of a gene is not enough to produce a normal phenotype. - Mechanism: One allele is inactivated or deleted → 50% of normal gene product → insufficient for normal function. 2. Dominant negative - A mutated gene product interferes with the function of the normal (wild-type) protein. - Mechanism: Mutant protein binds or competes with the normal protein → disrupts function of the protein complex.

Question 48

how can we distinguish haploinsufficiency with dominant negative using protein levels?

Answer 48

Haploinsufficiency: ~50% - One functional allele → reduced dosage Dominant Negative: 0–50% - Mutant protein disrupts normal protein

Question 49

how can we detect allelic polymorphisms at the molecular level?

Answer 49

- PCR and DNA sequencing - new technologies

Question 50

principles of allele detection

Answer 50

- ultimately resides at the level of DNA sequence - can detect polymorphism from DNA to protein level - analysis performed on diploid nuclear genome

Question 51

what provides the most comprehensive picture of allelic polymorphisms?

Answer 51

PCR amplification and DNA sequencing,

Question 52

2 ways to screen for BRCA1

Answer 52

- gene sequencing: most comprehensive but most expensive (BRCA1 is >80,000 bp) - SNP detection of 1-3 more common mutations known to cause breast cancer - chapter but much less comprehensive

Question 53

suggested screening strategy for relatives of patients with breast cancer

Answer 53

- for a patient with cancer, use gene sequencing to screen for causal mutation. possible results: causal SNP identified, SNP of uncertain significance, or no causal SNP - if causal SNP identified, use SNP detection approaches at identified SNP to screen at-risk relatives

Question 54

new technologies for disease screening

Answer 54

next generation sequencing, eg Illumina, allows for massive amounts of sequencing at much lower costs