Inheritance Flashcards
Emily Remnant lecture series
Who was Mendel and what What did Mendel do?
Mendel-
• Discovered basic principles of inheritance
• Defined laws to explain how genes pass on from one generation to another
o Law of independent assortment
o Law of segregation
• Set the theoretical basis of genetics and modern inheritance before we knew about DNA
• Mendel was a monk who bred pea plants to see how inheritance works, tracking inheritance of discrete traits such as colour through cross-pollination experiments. His success was recognised 50 years after his death
o Controlled crosses with pure-breeding varieties observing simple discrete phenotypes
o Many replicates and test crosses were important for the success of his experiments
What is the law of independent assortment?
alleles of one gene sort into gametes independently of the alleles of another gene (not always true)
What is the law of segregation?
the alleles of a given locus segregate into separate gametes. Each gamete only gets one allele of each gene
What phenotypic ratio does a Dihybrid cross (2 genes) using heterozygous population with dominant/recessive traits independently assorting give?
9:3:3:1
How do you do a dihybrid cross punnet square?
• For dihybrid cross punnet square-
o First find potential gametes that one parent could produce and cross that with potential gametes that other parent could produce
o Probability of each phenotypic class can be calculated based on the frequency of gametes produced by each parent
What phenotypic ratio does a Monohybrid cross using heterozygous population with dominant/recessive traits give?
3:1
What are genes?
physical units of heredity (defined DNA sequences)
What are chromosomes?
long molecules of double-stranded DNA and protein, which contain genes
What is a phenotype?
the observable traits of an organism
What is a genotype?
the genetic constitution of an organism
What is a wild type?
the most common variant in the wild
What are alleles?
alternative (variant) forms of a gene
What is a homozygote?
Contains two copies of the same allele
What is a heterozygote?
Contains two different alleles
What is a dominant trait?
Trait will be expressed in heterozygote
What is a recessive trait?
Trait will be masked by dominant allele
Why do we need statistics in genetics?
• Need statistics as we only have finite number of samples in each experiment- makes conclusions more certain
• Statistics give us a probability value how likely it is that our hypothesis about inheritance is correct
o If probability value obtained is bigger than 0.05, can be often confident that we are correct
What does chi-square hypothesis testing test?
• Tests wow well the observed number of offspring (O) fits with the expected (E) number of offspring
What is the formula for chi square hypothesis testing?
χ^2=
Σ((O-E)^2/E))
How do you perform chi square hypothesis testing?
- Form a hypothesis about the expected offspring based on predicted phenotypic ratios (Null hypothesis/H0)
- Calculate expected values based on total number of observations
Total multiplied by predicted ratio - Calculate the chi-square values for each class
Each different phenotype- one discrete category/class
Add the values together to get final chi square value - Determine degrees of freedom
Calculated as the number of phenotypic categories (or the number of alleles) (n) minus 1
• (n-1) - Determine the probability (p) associated with the chi-square value
Use table of chi-square values
Why do mutant phenotypes arise?
Mutant phenotypes are due to mutations in the DNA sequence of genes
What can variation in amino acid sequences result in?
o Variation in amino acid sequences can result in related proteins with some differences in function
What can variation in gene regulation result in?
o Variation in gene regulation can change the amounts of gene product being made and/or change when and where the gene is expressed
What can variation in intron and exon splicing in mRNA result in?
o Variation in intron and exon splicing of mRNA can result in proteins with different functional domains present or absent
What can novel genes and functions result in?
o Novel genes and functions, although rare, can have a significant impact
What are the seven phenotypes Mendel followed in peas and what are their dominant/recessive versions?
o Flower colour
Purple- dominant
White-recessive
o Flower position
Axial- dominant
Terminal- recessive
o Seed colour
Yellow- dominant
Green- recessive
o Seed shape
Round-dominant
Wrinkled- recessive
o Pod shape
Inflated- dominant
Constricted- recessive
o Pod color
Green-dominant
Yellow- Recessive
o Stem length
Tall- dominant
Dwarf- recessive
What gene is flower colour in peas controlled by and how?
Controlled by basic-helix-loop-helix (bHLH) transcription factor
• bHLH switches on expression of genes involving in making anthocyanin (purple) pigments
o bHLH binds to promoter of gene which leads to transcription of protein that makes anthocyanin pigments
• When it isl mutant, flower is white
o When there is G->A mutation in bHLH gene, bHLH transcription factor is not produced which means that can no longer bind to promoter of gene and protein that makes antocyanin pigment is not produced, turning the flower white
What gene is seed colour in peas controlled by?
Controlled by magnesium dechelatase gene (stay-green gene):
• Positive regulator of the chlorophyll degrading pathway
• When it is mutant, seed colour is green
What gene is seed shape in peas controlled by and how
Controlled by starch-branching enzyme 1 (sbe I)
• Converts amylose to amylopectin, the branched form of starch (starch is a polymer of glucose subunits, some of which are linear and some are branched)
• When it is mutant, peas are wrinkled
o In round peas, starch branching enzyme links all linear glucose molecules together to produce starch, which is a rigid structure making the peas appear round
o The starch branching enzyme is wrinkled in peas, leading to reduced concentrations of amylopectin, the branched form of starch as glucose polymers not joined together
Higher sugar content means less water in peas making it wrinkled
What gene is stem length in peas controlled by?
Controlled by gibberellin 3 beta-hydroxylase:
• Gibberellin is a growth hormone that stimulates cell elongation and cases plants to grow taller
• When it is mutant, plants are dwarves
Are dominance relationships between multiple alleles relative or not?
• Dominance relationships between multiple alleles are relative
What are dominance relationships dependant on?
Context and what you measure
What is co-dominance?
a heterozygote expresses the phenotype of both alleles simultaneously
What is an example of co-dominance?
EG. ABO blood antigens
• IA and IB allele produce A and B antigens and these alleles are co-dominant with each other, leading to an AB phenotype
• Dominant to i allele (basis of O blood) that produces nothing
What is incomplete dominance?
A heterozygote is intermediate between the two homozygous phenotypes
What is an example of incomplete dominance?
cross between a red lower and white flower gives pink flower
What is a loss of function mutation and are they usually dominant or recessive?
Decrease or complete loss of functional gene product
• Partial loss of gene function- leaky or hypomorphic mutation
• Complete loss of gene function- null mutation
These mutations are usually recessive
-Include haplo-sufficient and haplo-insufficient mutations
Describe haplo-sufficient mutations. Are they usually recessive or dominant?
Haplo-sufficient- one wild type allele provides enough normal gene product to produce a wild-type phenotype
• The mutation is recessive
What is an example of haplo-sufficient mutations?
• Example- albinism
o Albinism results from loss of melanin pigmentation in skin cells
o Several genes control the synthesis of melanin in melanocytes
Tyrosinase gene takes tyrosine (unpigmented) and converts it into melanin (brown pigmentation)
Tyrosinase loss of function mutants are recessive
Sufficient enzyme produced in heterozygotes for normal pigmentation
Wild-type allele is haplo-sufficient
What is a haplo-insufficient mutation and are they usually dominant or recessive?
Haplo-insufficient- a single wild-type allele in a heterozygote cannot provide the amount of gene product needed
• The mutation is dominant
What is a gain of function mutation and are they usually dominant or recessive?
Increase in functional gene product or new cellular function
These mutations are usually dominant
What is an example of a gain of function mutation?
EG- drosophila malenogaster
• In drosophila melanogaster, the Antp gene is a homeotic gene (developmental regulator) that promotes leg growth
• Antennapedia mutants arise from the mis-expression of the Antp gene causing legs to grow where antennae should be
• Mutation is dominant because the gain-off-function cannot be compensated by the wild-type allele and heterozygotes show the phenotype
Do genes only have 2 different alleles within a population?
• While diploid individuals can only contain 2 alleles per locus, genes can have many different alleles within a population
What is an allelic series and how is it determined?
• Alleles form an order of dominance or an allelic series, according to phenotypes expressed in the heterozygotes
o Have to figure out relative dominance between multiple alleles
Describe the allelic series of the agouti gene in mice looking at the a^y, a^w-j and a alleles
o The agouti in mice has more than 100 different alleles, including the black a allele, the yellow A^Y allele and the agouti A^w-J allele, which forms an allelic series of A^y>A^w-j>a
Describe how the agouti gene works
the agouti locus produces a molecule that regulates the synthesis of black pigment eumelanin
Different alleles of the agouti locus modify this pattern of bands on a hair shaft in various ways, with pheomelanin appearing when eumelanin production is blocked
Describe the agouti gene allelic sequence in dogs looking at the a^y, a^w, a^t and a alleles
In dogs, the ay allele blocks synthesis of eumelanin so is dominant to all other alleles
Allele a^y>a^w>a^t>a
a lacks the ability to control distribution eumelanin, giving a completely black coat
Describe the allelic sequence of the C gene in rabbits looking at the C, c^ch, c^h and c alleles, what each of these alleles represent and how these mutations occured
o The C gene contains multiple alleles including the C allele (resulting in black rabbits), the c^ch allele (resulting in chinchilla grey rabbits), the c^h allele (resulting in himalayan rabbits) and the c allele, resulting in white rabbits
o The allelic series is C>c^ch>c^h>c
o The c,c^ch and c^h recessive alleles result in a decrease or complete loss of functional gene product
Chinchilla is partial loss of gene function (leaky or hypomorphic mutation)
White is complete loss of gene function (null mutation)
Himalayan is conditional mutation (mutant phenotype is temperature sensitive)
• Enzyme active at low temperatures (black ears, nose…)
• Enzyme inactive at high temperatures (white cores)
What are conditional mutants?
• Conditional mutants- mutation causes a phenotype only under certain environmental conditions (e.g. temperature, chemical exposure, resource availability)
What 2 different conditions are there in conditional mutants?
o Restrictive condition- mutant phenotype (restricts growth of individual)
o Permissive condition- wild-type phenotype (allow growth of individual/normal phenotype)
What is incomplete penetrance?
• Incomplete penetrance- not every individual with the mutant genotype displays a mutant phenotype
What is penetrance?
o Penetrance- the proportion of individuals of a specific genotype that exhibits the corresponding phenotype
What is an example of incomplete penetrance?
o e.g. polydactyly
Single dominant gene, causing growth of extra fingers and toes
Incomplete penetrance- at least 1 in 4 people with the mutation have 5 digits (penetrance=0.75: ¼ of people have normal phenotype and ¾ of people have mutant phenotype)
What are lethal alleles?
alleles that cause the death of the organism that carries them
What is an example of a lethal allele?
o Example: the agouti AY allele in mice
In mice, A alleles produce wild-type (agouti): dark hair with yellow band
The AY allele in mice is a gain of function mutation resulting in ubiquitous overexpression of the agouti protein (mutants produce solid yellow hair)
Homozygous mutants are embryonic lethal
• Not possible to generate a pure-breeding yellow line
What is the mendelian ratio for dominant lethal alleles?
Dominant lethal alleles
• Homozygous dominant mutant will die
• Offspring ratio= 2:1
What is the mendelian ratio for recessive lethal alleles?
Recessive lethal alleles
• Homozygous recessive mutant will die
• Offspring ratio: 1 (all same dominant phenotype)
What is a pleitropic gene?
• Pleitropic genes- a single gene that affects more than one phenotype or process
What is an example of a pleitropic gene and why is it lethal?
o Example- the agouti AY allele affects both coat colour (yellow dominant), weight (obese dominant), tumor susceptibility, hyperinsulemia (leading to embryonic lethality)
Too much agouti protein results in increased antagonism of melanocortin receptors
How could mutants with the same phenotype occur?
• Mutants with the same phenotype could be alleles of the same gene, or mutations in two different genes
Describe the 5 genes that contribute to coat colour
A gene (agouti)- determines distribution of pigment along the hair
B gene- determines the colour of pigment
C gene- permits colour expression
D gene- controls the intensity of pigment
S gene- controls distribution throughout the body (spotting)
What is the complementation test for and how does it work?
• The complementation test- a functional test between recessive mutants with the same phenotype
Process:
o Two homozygous recessive mutants with the same phenotype are mated
o F1 offspring are analysed
o If the F1 is a wild-type, the two mutations complement each other and represent two different genes
o If the F1 is a mutant, the two mutations fail to complement and represent two different alleles of the same gene
How can we determine if gene interactions are occuring?
• Genes rarely act in isolation
• Can use genotypic and phenotypic ratios of some crosses to determine how many and what type of gene interactions are occurring
o If deviation from expected ratios, means some kind of gene interaction is occurring
What are 5 different types of gene interactions?
- No interaction
- Complementary gene interaction
- Duplicate gene interaction
- Dominant gene interaction
- Epistasis
- —-Recessive epistasis
- —-Dominant epistasis
- —-Dominant suppression epistasis
What is the mendelian ratio for no gene interaction and what does no gene interaction mean?
Two genes, no interaction
Get a 9:3:3:1 ratio
What is complementary gene interaction and what is its mendelian ratio?
Two genes act sequentially to produce a phenotype. Both are required
Get a 9:7 ratio
What is duplicate gene interaction and what is its mendelian ratio?
Two genes perform the same function (redundancy). Either are required
Get a 15:1 ratio
What is dominant gene interaction and what is its mendelian ratio?
Two genes with the same phenotype interact additively (causes enhanced/additive phenotype)
Get a 9:6:1 ratio
What is epistasis?
One gene stops/masks the phenotype of another gene
What is recessive epistasis and what is its mendelian ratio?
• Recessive epistasis- homozygous recessive alleles for one gene masks the phenotypic expression of alleles at a second gene. Most common epistasis version.
o Get a 9:3:4 ratio
What is dominant epistasis and what is its mendelian ratio?
• Dominant epistasis- dominant allele of one gene masks the phenotypic expression of another gene
o Get a 12:3:1 ratio
What is dominant suppression epistasis and what is its mendelian ratio?
• Dominant suppression epistasis- dominant allele of one gene suppress the expression of the dominant allele of another gene
o Get a 13:3 ratio
Describe when chromosomes were first visualised
• Chromosomes visualised as early as 1842
Describe when the process of division was first described
• Process of division described later (1873-82)• Process of division described later (1873-82)
What are the 2 types of cell division, where do they each take place and what is the chromosome number relative to parent cells in their daughter cells?
o Mitosis- Somatic cells: Chromosomes replicate and divide, and chromosome number is maintained in daughter cells
o Meiosis- Germline cells: Chromosomes replicate and undergo 2 rounds of division: chromosome number is halved in daughter cells
What is a karyotype?
• Complete set of chromosomes is called a karyotype
Is chromosome number the same in every species?
No
How many chromosomes do humans have?
o Complete set of human chromosomes is 23 pairs (46 chromosomes)
What is a homologous chromosome pair?
• A homologous chromosome pair has similar DNA sequence and encodes the same genes
What is the chromosome theory of inheritance? (Sutton and Boveri 1902)
Chromosome theory of inheritance states that individual genes are found at specific locations on particular chromosomes, and that the behavior of chromosomes during meiosis can explain why genes are inherited according to Mendel’s laws
Describe what experiment proved the chromosome theory of inheritance
• At the time it was proposed, the chromosome theory of inheritance lacked experimental proof and was wildly controversial
o Morgan and drosophila melanogaster (1910)
Experimental evidence came from one of the first identified mutants in Drosophila melanogaster: a male with white eyes which is unusual as the wild type usually have red-eye colour
Morgan crossed his white type male to his wild type female, and got all wild type offspring (F1 individuals)
Morgan then crossed all his F1 individuals together but found that none of the females were white eyed, but half the males had red eyes and the other half had white eyes (not the 3:1 ratio expected)
What are observations that support the chromosome theory of inheritance?
o This is because chromosomes come in homologous pairs, and one member of the pair comes from the mother and the other from the father
o The members of a homologous pair separate in meiosis, so each sperm or egg receives one- This process matches the segregation of alleles into gametes in Mendel’s law of segregation
o Different chromosome pairs sort into gametes independently of one another in meiosis, like as predicted for genes in Mendel’s law of independent assortment
What are ABC transporters?
o ABC transporters-ATP-Binding cassette transporters which are transmembrane proteins that use ATP to transport molecules across membranes
Describe what role the white gene in drosophila has on drosophila eye colour
• The white gene is an ABC transporter required for eye pigment production
• White gene occurs upstream of many pigmentation genes and delivers pigment precursors guanine and tryptophan to the eye to make the brick red pigment
o Trypotophan, aided or regulated by many genes, eventually makes zanthommatin brown pigment
o Guanine, aided or regulated by many genes, eventually makes drosopterin, orange pigment
• Xw mutation blocks the transport of pigment precursors guanine and tryptophan, resulting in white eyes
When were sex chromosomes first discovered?
• Nettie Stevens (1905) used the mealworm beetle to show that:
o Male gametes had 10 large chromosomes, or 9 large chromosomes and 1 small chromosome
o Female gametes all had 10 large chromosomes
• Observed sex-based differences in chromosomes from a range of insects
Do all organisms have sex chromosomes?
• In many organisms, sex is determined by the presence of sex chromosomes but some organisms do not have sex chromosomes
What is the heterogametic sex?
• Heterogametic sex- the sex with different sex chromosomes
What is the homogametic sex?
• Homogametic sex- the sex with homologous sex chromosomes
What is a hemizygote?
• Hemizygote- Only one copy of an allele is present at a locus instead of two. The phenotype is present regardless of dominance
What are the two human sex chromosomes?
X and Y
Are human males heterogametic or homogametic?
Heterogametic (XY)
Are human females heterogametic or homogametic?
Homogametic (XX)
Describe the origin of human sex chromosomes
o Sex chromosomes in humans were originally derived from a pair of homologous autosomes (non-sex chromosomes)
1. The Y chromosome acquired a sex determining locus
• Accumulated genes with functions in sex and male fertility
2. Differentiation of X and Y specific regions, with small regions of homology retained in pseudo-autosomal regions (which help X and Y chromosomes to pair up during cell division)
3. Y chromosome lost genes and accumulated mutations
4. Dosage compensation
5. Final heteromorphic chromosomes differ greatly in size and morphology
-Over evolutionary time, they have changed in size and morphology
Describe how many megabases and genes the X chromosome has and how conserved this gene is in placental mammals
o The X chromosome
165 Megabases, about 1000 genes
Highly conserved in placental mammals
Describe how many megabases and genes the X chromosome has, describe its composition and role as well as its evolution
o The Y chromosome
59 Megabases, about 200 genes
Specialised male fertility genes
Many residual and non-functional sequences
Many human sex-linked traits predominantly affect males
Y chromosome has changed due to gene loss over time
Sex determining region on the Y chromosome
What is a pseudoautosomal region?
o Pseudoautosomal region- regions of chromosomes that have maintained homology
What are genes on the X chromosomes called?
X-linked genes
What are genes on the Y chromosome called?
Y-linked genes
Describe the offspring ratio that occurs when there is a Parental cross of homozygous wild-type female with hemizygous mutant male involving a sex-linked trait
• Parental cross of homozygous wild-type female with hemizygous mutant male results in a 2:2:1: ratio when trait is sex linked
o 2 female wild type: 2 male wild-type: 1 mutant male
Describe the offspring ratio that occurs when there is a parental cross of a heterozygous wild-type female with hemizygous mutant male involving a sex-linked trait
• When trait is sex-linked, parental cross of a heterozygous wild-type female with hemizygous mutant male results in a 1:1:1:1 ratio when trait is sex-linked
o 1 female wild type:1 female mutant:1 male wild-type: 1 male mutant
What is criss-cross inheritance?
• Criss-cross inheritance- the trait goes from mother to son, mirroring the inheritance of the X chromosome
o Mothers will only pass on X chromosomes to their sons, and fathers will only pass on X chromosomes to their daughters
What can be inferred of the mother’s genotype if the son’s phenotype is provided
o Mothers that are carriers of the trait are assumed based on the phenotype of sons
Unaffected mothers with affected sons must be heterozygotes
Mothers who are not affected when their sons are is a way of guessing that it is a sex-linked trait
Describe what population is affected by Y-linked inheritance
o Trait occurs in males only
o Occurs in all sons of the affected males
o Daughters of affected males are normal and do not have affected offspring
Describe the consequences of X-linked dominance inheritance in a family tree
(What genotype will result in the trait being phenotypically present in males/females, what will happen phenotypically if an affected heterozygous female is mated to a normal male, and what will happen phenotypically when affected hemizygous males will mate)
Trait is present in males hemizygous for the allele
Trait is present in females homozygous or heterozygous for the allele
Each individual who has the disease has at least one affected parent
• Affected heterozygous females mated to normal males transmit the defect to ½ of their sons and ½ of their daughters
• Affected hemizygous males transmit to all their daughters
• Every affected has at least one affected parent, except in the case of de-novo mutations
Describe the consequences of X-linked recessive inheritance in a family tree
(What genotype will result in the trait being phenotypically present in males/females, what will happen phenotypically if a heterozygous female is mated to a normal male, what will happen phenotypically when affected hemizygous males will mate and what will happen if affected females mate with normal males)
Trait is present in males hemizygous for the allele
Trait is present in females homozygous for the allele
The defect or disease may skip generations
• Normal parents with a female carrier will give no affected females and ½ affected males
• Affected males mated to normal females will have no affected offspring, but all daughters are carriers
• Affected females mated to normal males will have all sons affected and no affected daughter, but all daughters are carriers
Describe the inheritance pattern of autosomal recessive inheritance (if one or all parents have the disorder, what is the phenotype of the children, and is it affected by sex)
o Autosomal recessive inheritance
Individuals who have the disease are often born to parents wo do not
If only one parent has the disorder the risk that a child will have it depends on the genotype of the other parent
If both parents have the disorder, all children will have it
The sex ratio of affected offspring is expected to be equal
The disease is not usually seen in each generation but if an affected child is produced by unaffected parents, the risk to subsequent children is ¼
If the disease is rare in the population, unaffected parents of affected children are likely to be related to one another
Describe the inheritance pattern of autosomal recessive inheritance (if one, all or none of the parents have the disorder, what is the phenotype of the children, and is it affected by sex)
Each individual who has the disease has at least one affected parent
Males and females are affected in equal numbers
Either sex can transmit the disease allele
In crosses where one parent is affected and the other is not, approximately half the offspring have the disease
Two unaffected parents will not have any children with the disease
Two affected parents may produce unaffected children
Describe genetic allelic notation for Drosophila recessive, dominant or sex-linked traits
o Instead of lower case upper case allelic notations, wild-type alleles are indicated by a + superscript. For a recessive trait with allele e • Wild type allele: e+ For a dominant trait with allele B • Wild type allele: B+ For a sex-linked trait with allele w: • Wild type allele: Xw+ o Dominant mutations are given a capital letter and recessive mutations are lower case
Describe the chromosome composition and appearance in Drosophila melanogaster
o Drosophila melanogaster have 4 chromosomes
o 1 X chromosome and 3 autosomes
o Each chromosome has a left and right arm at either side of the centromere except for chromosome 4
Describe sex determination in Drosophila
• Sex determination in Drosophila depends on the number of X chromosomes, not on the presence of a Y chromosome: two X chromosome is a female and one X chromosome is a male
o XX or XXY= female
o XY or XO= male
What are reciprocal crosses and what is their purpose?
- Used to determine sex-linkage: two crosses are performed, where the genotypes of the male and female parents are swapped
- If F1 offspring ratios differ, it indicates that this trait is sex-linked
What are pedigrees and what are they useful for?
- Pedigrees, or family trees, are a way of tracing the inheritance of traits
- They are useful to understand the mode of inheritance of a trait when offspring numbers are low, but family records are available
Describe the symbol used in pedigrees for:
- Males
- Females
- Unaffected
- Affected
- Heterozygous
- Deceased
- Unspecified sex
- Generation
- Parents
- Parents related by blood
- Adoption
- Siblings
- Identical twins
- Fraternal twins
- Generations
- Individuals in a generation
- Males: square
- Females: Circle
- Unaffected: Empty square/circle
- Affected: Filled square/circle
- Heterozygous: Dot inside square/Circle
- Deceased: Diagonal line (from bottom left to top right inside shape)
- Unspecified sex: Diamond
- Generation: Vertical line
- Parents: Horizontal line
- Parents related by blood: 2 horizontal lines
- Adoption: Dotted vertical line
- Siblings: Horizontal line joining vertical lines
- Identical twins: Two lines joined by horizontal line
- Fraternal twins: Two lines
- Generations: Indicated by roman numberals
- Individuals in a generation: Indicated by arabic numerals
Timestamp: 3:40 on the 13/08/2019
What is autosomal dominant?
one mutant allele is sufficient for trait or disease
What is autosomal recessive?
two mutant alleles result in trait or disease
What is X-linked
hemizygous males express the trait, regardless of dominance
Describe the pattern of inheritance of many genetic traits and diseases
- Thousands of human genetic traits and diseases are caused by mutations of single genes which follow mendelian patterns of inheritance
- Can occur on autosomes or sex chromosomes
Why is genetic testing important and what can be done in genetic testing?
• Many deleterious alleles causing genetic diseases are recessive (autosomal or X-linked)
o Detection of carriers is important as carrierxcarrier matings will give affected offspring in autosomal genes at ¼ frequency
• Can do molecular test where alleles of individuals are examined and detect whether the offspring has inherited 2 mutant alleles
o Can also result in employment of personalised medicine where treatment is targeted to a person’s specific genomes and susceptibility
Describe commercial genetic testing
• Commercial genetic testing can report one’s traits and genetic characteristic but results should always be interpreted with a medical background as well, not just a genetic background and commercial genetic testing has many limitations.
What is sexual dimorphism and examples of these
• Many species exhibit sexual dimorphism- physical differences between males and females. For example:
o Colour dimorphism
o Size dimorphism
What are two different mechanisms for sex determination?
• There are lots of different mechanisms for sex determination-
o Both genetic and environmental sex determination mechanisms exist
What are monoecious plants?Give an example
• Monoecious plants- male and female flowers on the same plant such as zucchini
What are hermaphrodites?
o Hermaphrodites- contain both male and female sex organs
What percentage of plants are hermaphrodites?
94% of plants have both male and female organs, often within the same flower
What percentage of animals are hermaphrodites?
About 5% of animals (some worms, snails and fish (such as clownfish)) are hermaphrodites
What are dioecious plants, the percentage of dioecious plants and examples of these?
• Dioecious plants- male and female flowers on different plants
o Only 6% of plants have split into different sexes, including papaya and asparagus
What is genetic sex determination and how does it work?
genetic elements specify whether individuals are female or male
o Individuals release some genetic signal during embryonic development to tell to develop as male or female
What are 3 different types of genetic sex determination and in what animals are they present?
Heteromorphic sex chromosomes- the presence of specific sex chromosomes that are different and visually distinct from each other, and distinct from autosomes
• Heterogametic sex- the sex with different sex chromosomes
• Homogametic sex- the sex with homologous sex chromosomes
• For example, mammals, birds, some reptiles and fish, many invertebrates
Sex-determining genes on homomorphic chromosomes
• Sex-determining/mating type genes are located on autosomes
• Species do not have sex chromosomes
• Amphibians, some reptiles and fish, some invertebrates
o E.g. swordtails: a mating-type gene with 2 alleles determines sex. Females are homozygotes and males are heterozygotes
Chromosome ploidy:
• Males develop from unfertilised eggs and are haploid (n)
• Females develop from fertilized eggs and are diploid (2n)
• Invertebrates (all bees, wasps, ants, most mites and ticks)
What is environmental sex determination?
• Environmental sex determination- external stimuli control sex determination
o Favoured when specific environments are more beneficial to one sex
Describe the order in which sex determination evolved in vertebrates (animal and chromosome type order, not evolution of actual chromosomes)
- Reptiles and birds:
o Birds: ZW, Reptiles: ZW, XY and environmental
o Sex chromosomes have no homology to mammalian XY (come about on different chromosomes present in these organisms). - Mammals
o XY(or variations)
o Monotremes: earliest offshoots of the mammalian lineage
What are the impacts of climate change on temperature sex-determination?
• Sex determination is quite robust-genetic sex determination that can transition in environmental mechanism and vice-versa
Describe sex determination in australian bearded dragons
• Australian bearded dragons use both sex chromosomes and temperature to determine sex
o Normal temperature:
Heteromorphic sex chromosomes and genetic sex determination
• ZW females and ZZ males
o High temperatures higher than 32
Male dragons with ZZ sex chromosomes undergo sex reversal and develop as female
• W chromosome eliminated from population
Must be advantages to having more females as females are favoured in high temperature zones
o Transition to environmental sex determination
What are different types of chromosomal genetic systems?
- XY systems
- XO systems
- ZW systems
- Haplo-diploidy
Describe the genetic XY system:
Male vs female, animals involved, master-sex determining locus
Female: XX
• The absence of SRY allows expression of genes needed for ovary differentiation (the default pathway)
Male: XY
• In mammals, the presence of the Y chromosome determines if an offspring develops as a male
• Male sex depends on a master sex-determining locus, the SRY (sex determining region Y) gene
• SRY encodes a protein, the testis-determining factor (TDF) which leads to testis differentiation and male sexual development
Describe sex determination of monotremes
Monotremes have 10 sex chromosomes
• Females have 10 X chromosomes, males have 5X and 5Y chromosomes
• X1 chromosome has homology to other mammal X chromosomes
Describe the XO genetic system (Males vs female, animals involved)
Female: XX, XXY
Male: XO, XY
The number of X chromosomes determines sex in Drosophila and many other invertebrates
• More than one X chromosome- female
• One X chromosome- male
In many nematodes and other insects, there is no Y chromosome- XX are female and XO are male e.g. C elegans
What is the master-sex determining locus in drosophila and how does it work?
The drosophila chromosome is not involved in sex determination but contains genes required for male fertility. An XO is male but sterile.
• Sex-lethal (Sxl) is the master sex-determining locus in Drosophila
o XX embryo: High sex-lethal expression, which triggers production of the female isoform of the gene doublesex (dsx) via alternate splicing, leading to female development and results in no hypertranscription of X
o XY or XO embryo: low Sxl expression which leads to production of the male isoform of dsx and results in hypertranscription of X
What is the sec-determining locus in C.elegans and how does it work?
- XX means that master sex-determining locus xol-1 is off, which makes a hermaphrodite
- XO means that the master sex-determining locus xol-1 is on, which makes a male.
Describe the ZW genetic system (Males vs females, animals involved, description of chromosomes)
Female: ZW
Male: ZZ
All birds, some reptiles, some insects and fish
Males are the homogametic sex (ZZ) while females are the heterogametic sex (ZW)
• Z chromosome: large, gene rich
• W chromosome: Small, few genes
ZW heteromorphic chromosomes are structurally similar to XY (large vs small) but have completely different gene content
• Dose-dependent Z (need multiple copies of Z to become a male)
What is the master sex-determining gene in birds?
• Master sex-determining gene: dmrt1
Describe the haplo-diploidy genetic system (males vs females)
Female: Diploid
Male: Haploid
Describe honey bee sex determination (chromosome number, sex determiner gene)
Haplo-diploidy
E.g. honey bees
• Females have 32 chromosomes (16 pairs) whilst males have 16 unpaired chromosomes
• Males can only have daughters (if produce diploid malformed males are killed by the colony at development)
• Sons do not have fathers
• Sex is determined by the complementary sex determiner (csd) gene
o Males are haploid and hemizygous for csd
o Females must be heterozygous at the csd locus to develop normally
o Homozygous diploid eggs develop into inviable males
o There are hundreds of different csd alleles
o Honey bee queens mate with over 20 different males to ensure that they have high diversity of alleles at this gene
Will have sperm stored in their ovaries to make sure they can fertilise eggs with lots of csd alleles to make sure they have lots of female heterozygous offspring
Describe 3 examples of environmental sex determination systems
• Environmental systems
o Temperature of egg incubation in reptiles (such as turtles)
Female: Warmer temperatures
Male: Cooler temperatures
o Photoperiod in marine amphipods
o Social factors in many coral reef-dwelling fish
What are sex determination abnormalities that can occur in species where sex is cell autonomous?
o Gynandromorphs- embryos develop as part male and part female, often leading to spectular phenotypes
Chimeras- fusing of two embryos in early development
Polyspermy- multiple sperm fertilise egg, different lineages develop
Errors in mitotic division early in development (sex chromosome loss)
What does cell autonomous sex determination means?
each cell decides its own sexual fate based on its sex chromosome constitution
What are sex determination abnormalities that can occur in species where sex is cell nonautonomous and why?
o Although the SRY gene is considered a master sex-determining locus, other genes are involved in the sex determination cascade
Other genes involved in production of testosterone, some acknowledge testosterone (such as testosterone receptors)
If genes in these other pathways are interrupted, can have changes in sex determination cascades
o Sex-reversal can occur when genetic males develop as females and vice versa due to mutations in key genes in the pathway, for example:
Translocation of the SRY gene to the X chromosome male development with 2X chromosomes
Mutations in genes downstream of SRY XY genetic male but female development
Describe cell nonautonomous sex determination
• Species where sex determination is cell nonautonomous (mammals)
o Sex determining gene controls the fate of the gonads early in development: ovaries or testes
o Gonads direct establishment of phenotypic sex through the production of hormones
o Hormones direct development of sex-specific characteristics throughout the body
Gynandromorphs not possible
Why is there a need for sex chromosome balance?
• In species with heteromorphic sex chromosomes, XX and XY individuals would express their X-linked genes at different levels, unless a special mechanism exists to control the expression of these genes
How is sex chromosome balance achieved? Give an example animal for each mechanism
o In the absence of a special mechanism, XX animals and XY animals would have different levels of X-linked transcription
o Dosage compensation mechanisms have evolved that balance the level of X-linked gene products between the sexes
Up-regulation in the expression of X-linked genes in males; in Drosophila
Down-regulation of X-linked genes in females: in the worm C.elegans
Complete inactivation of one of the two X chromosomes in females: in mammals
Describe how up-regulation in the expression of X-linked genes in male Drosophila flies works
- Drosophila upregulate genes on the X chromosome in males: XY flies have an increased level of X-linked transcription from their single X chromosome (2 fold increase) compared to XX flies
- Dosage compensation complex (DCC), a complex of proteins and RNA molecules, is recruited to the X chromosome in males and is responsible for the increased transcription of X-linked genes
Describe how down-regulation of X-linked genes in females in the worm C.elegans works
- XX worms (females) have a lower level of transcription of each X chromosome compared to XO worms (males)
- The dosage compensation complex (DCC) is recruited to both X chromosomes in XX worms, and both X chromosomes are transcribed at a lower level
What is a Barr body?
condensed, inactivated X chromosome found in female nucleus
How do you visualise a Barr body?
o Visible under the light microscope, with a stain that detects condensed chromatin
How does X chromosome inactivation in a female organism work? (how X chromosome is deactivated, how many are deactivated)
• Only one X chromosome is transcribed in XX individuals- the other X chromosome is condensed into heterochromatin and is no longer transcribed
• Only one X chromosome is expressed in any given cell
• X-linked genes are expressed at the same level in males and females through X-chromosome inactivation dosage compensation
o Most genes are inactivated but not all
o X chromosome inactivation requires Xist
Xist RNA coats the inactive X chromosome and recruits proteins that silence gene expression and result in chromosome condensation
- Inactivation is random (happens in 50/50 ratio)
- Once an X chromosome has been inactivated, it remains inactive through subsequent cell divisions
When does X chromosome inactivation in a female occur?
• X chromosome inactivation in mammals happens early in development as cells divide: one of the two X chromosomes becomes condensed and localised to the nuclear membrane where it is not transcribed
What is the Xist gene and what does it produce?
In placental mammals, the X-inactivation-specific-transcript (Xist gene) is an RNA gene on the X chromosome
Xist is transcribed from the X-inactivation centre of the future inactive X to produce a 17kb long non-coding RNA
Describe X chromosome inactivation results in calico cats
• Example- calico cats
o Cats have an X-linked gene B that affects coat color
o In XBXb heterozygotes, some cells have the chromosome with the B allele inactivated, while other cells have the chromosome with the b allele inactivated
XBXb heterozygotes are a mosaic, with patches of black fur and patches of orange fur (calico)
What is the consequence of X chromosome inactivation in mammal females who have recessive disease traits
• Females that are carriers of X linked disease: half of their cells will have inactivated the mutant X, while half have inactivated wild-type
• In females that are ‘mosaic’ for a disease, the products of both alleles are expressed: sufficient to prevent full expression of X-linked recessive alleles in a heterozygous female
o Will have enough expression of wild-type X chromosome to take over effect of X-linked recessive allele
o Often asymptomatic or only minor symptoms of a given disorder
What is X-inactivation skewing?
• X-inactivation skewing: sometimes a non-random proportion of X’s of one type are inactivated
Where are genes found?
• Genes occupy physical locations (loci) on chromosomes
What characteristic morphology do chromosomes have that can be observed under a microscope?
• Chromosomes have a characteristic morphology that can be observed under a microscope
o Heterochromatin and Euchromatin
o Telomere, centromere, short (p) and long (q) arms
Do all species have the same chromosomes?
• Chromosomes occur in species-specific sets (karyotypes)
o Species have characteristic numbers of chromosomes with distinct sizes and morphology
o All eukaryotes have a defined number of chromosome sets int their genome
o Different species have evolved to have a different number of chromosomes
Describe chromosomal structure (from DNA to Chromosomes)
o DNA is wrapped around chromosomal proteins called histones to form nucleosomes
o Nucleosomes form compact chromatin
Histone H1 helps nucleosomes condense into 30nm fibre
o Chromatin is folded into higher-order structures that form the chromosome
Describe the structure of nucleosomes
Each nucleosome contains 146 bp DNA wrapped around histones (H2A H2B, H3 and H4).
There is an electrostatic interaction between histones and DNA< as histone proteins are positively charged while DNA backbones are negatively charged
This structure needs to be unfolded to allow separation of the DNA strands during gene expression or DNA replication
Describe chromosome eukaryotic anatomy, and why each part is essential
o Telomeres-Mix of protein and DNA sequences at either end of the chromosomes to protect the ends from further degradation
o Centromeres- Spindle microtubules bind to the kinetochore which is formed on centromeric DNA-essential to allow chromosomes to line up during division and not get lost during cell division
Chromosomes will line up on a metaphase plate and will be attached by a spindle formed out of microtubules
o P (short) and Q (long) arms
What are types of chromosomes and each of their distinct features?
• Types of chromosomes-
o Metacentric chromosome-
Centromere in middle
P and Q arms are the same
o Acrocentric chromosomes-
P arm is the short arm
Q arm is the long arm
Centromere is near the top of the chromosome
o Telocentric chromosomes
There is only the q arm
Centromere is at the top of the chromosome
What is the animal with the smallest amount of chromosomes and the number?
Smallest number of chromosomes: Jack jumper ant: 2 for females, 1 for males
What is the animal with the largest number of chromosomes and its number?
Largest number of chromosomes: agrodiae butterfly: 268 chromosomes
What is the monoploid number (n) of chromosomes?
o Number of chromosomes in a basic set is the monoploid number (n)
What is ploidy?
o Ploidy- the number of chromosome sets in an organism’s genome
What is the difference between haploid, diploid and triploid?
Haploid (n)- one set of every chromosome
Diploid (2n)- two sets of every chromosome
Triploid (3n)- three sets of every chromosome
How is chromosomal identity and content recognised and how does this work?
o Chromosomes have a characteristic banding pattern
Euchromatin contains most of the active genes
• Represented by a light stain G-banding pattern achieved through Geimsa staining
Heterochromatin is more condensed
• Represented by a dark stain G-banding pattern achieved through Geimsa staining due to AT rich regions in that DNA that the Giemsa stain stains
What is mitosis?
• Mitosis- somatic cells: chromosomes replicate once and divide once-chromosome number is maintained
What is meiosis?
• Meiosis-germline cells: chromosomes replicate once and undergo 2 rounds of division to make haploid gametes: chromosome number is halved
What are sister chromatids?
• Sister chromatids- basically identical to each other (except for mutations) and both replicated from the original version-DNA replication has occurred
What are homologous pairs of chromosomes?
• Homologous pair- matching pair of chromosomes that have been inherited from each parent
o Chromosomes are identical in size and characteristics, have a lot of differences in their DNA sequence
Describe the phases of meiosis and what happens at each one
o Chromosomes become progressively more condensed and visible during prophase I: chromosomes are synapsed (homologous chromosomes are paired together) and crossing over occurs
o Chromosomes are aligned across the middle during metaphase I
o In Anaphase I, homologues separate but sister chromatids remain together
o In Telophase I, cells separate
o Process starts again in meiosis II, but there is no pairing, no crossing over and the sister chromatids separate to produce 4 haploid products.
What is crossing over in meiosis?
Crossing over- occurs between chromatids of the two homologous chromosomes: sequences from one homologous are now on the other-transfer of genetic material
Does crossing over only happen once on selective pairs of chromosomes?
No.
Every pair of homologues crosses over at least once
The larger the chromosome is, the more chance that recombination events occurs (multiple events can occur on one chromosome)
• Can have recombination at sister homologues, but it wouldn’t be seen because they are identical sequences
Why does crossing over occur? Describe this process.
Homologues are used as the template to repair double strand breaks
• Double stranded breaks important for keeping chromosomes paired up correctly during meiosis
• DNA synthesis to repair a double stranded DNA break
• The newly synthesised DNA is attached to the DNA molecule on the other homologue
• Gaps are repaired
• One sister chromatid has DNA sequences from each homolog
• Recombination important as it allows the chromosomes to remain together at the right times
Where does crossing over occur? Describe why this is.
Crossing over occurs at chiasmata
• Chiasmata- the sites where homologue strands exchange DNA/cross over, which occurs during prophase I
• Homologues are held together by chiasmata while pulled towards opposite poles during chromosome separation at anaphase I
o Allow separation once double stranded breaks have been repaired
How does meiosis uphold Mendel’s law of segregation?
• Mendel’s 1st law, the segregation of alleles, is upheld as homologues separate during meiosis I and sister chromatids separate during Meiosis II: a gamete only has a haploid chromosome number and offspring will have diploid chromosome number
How does meiosis uphold Mendel’s law of independent assortment?
• Each pair of chromosomes segregates independently during meiosis, which is the cellular basis for Mendel’s 2nd law of independent assortment
What is spermatogenesis?
sperm production
What is oogenesis?
egg production
What is the difference beween spermatogenesis and oogenesis in terms of meiosis?
Spermatogenesis- continuous meiosis
Oogenesis- suspended meiosis between first and second stage
Describe meiosis in oogenesis and its products
First stage of meiosis occurs during embryoic development where all the eggs to be used in lifetime are made, while second phase only starts occurring when the female reaches sexual maturity
Females are born with all of their future ova present and arrested at prophase I
During ovulation, primary oocytes complete meiosis I and arrest at metaphase II
Fertilised 2nd oocytes complete Meiosis II
o Only 1 of 4 products becomes an ovum, the rest become polar bodies -one precursor cell will yield one mature ovum
What are polar bodies produced from oogenesis useful for?
• Genetic testing- take polar bodies and test their genotype
o If we know different alleles that are segregating in the mother, can determine whether functional oocyte inherited the good alleles if we can test that the polar body inherited the bad ones
o Polar body from Meiosis II should be the same as her functional egg
o The polar body from Meiosis 1 and the secondary oocyte (future ovum) will have different alleles
What are variations in chromosome number caused by?
• Caused by problems in meiosis and gamete formation
When does polyploidy occur, and what are two types of it?
• Polyploidy occurs when there is variation in the number of sets of chromosomes
- Autopolyploid
- Allopolyploid
What is autopolyploidy and when does it occur?Give an organism example
o Autopolyploid- multiple copies of the same genome: single species chromosome duplication
Occurs when the first division of meiosis fails and gives gametes without reduction in chromosome number
• Unreduced (2n) gametes fuse
• Results in either triploid (3n) or tetraploid (4n)
Common in plants- polyploids have larger cells, hence larger plants, leaves and fruits which is desired in agriculture, but these plants are sterile
What is allopolypoloid? Give an example.
o Allopolyploid- contain two (or more) different genomes after a hybridisation event: the fusion of two or more diploid genomes from different species
Wheat is an allopolyploid of three species (which all have 7 chromosomes)
What is aneuploidy, and two common types of aneuploidy?
• Aneuploidy occurs when the number of chromosomes of a particular pair is unbalanced
o Monosomy- missing one member of a pair
o Trisomy- with one extra chromosome in a pair
What is non-disjunction?
• Non-disjunction- failure of chromosomes to segregate normally
What is the result of non-disjunction?
o Results in daughter cells with abnormal chromosome numbers (aneuploidy)
When can non-disjunction occur and what is the difference in results at the different times of occurence?
o Can occur at both meiosis I and meiosis II
Non-disjunction at meiosis I results in one gamete that has both homologues, and one that has neither
Non-disjunction at meiosis II yields gametes that are homozygous
On what chromosomes can non-disjunction occur?
o Non-disjunction can occur for any chromosome, but is more common in sex chromosomes
What is an example of monosomy in humans?
Monosomy-one chromosome missing
• E.g. XO: Turner syndrome (2n=45)
What are 4 examples of trisomy in humans?
Trisomy- one chromosome is duplicated (2n=47) • XXX-triple X syndrome: Female o Two barr bodies • XXY-Klinefelter syndrome: Male o One barr body • XYY: Male Trisomy-21: Down syndrome (2n=47)
What are rare types of trisomy, non-existent ones, and why are they rarely in population?
Rare trisomy-
• Rare: trisomy 13 (Patau syndrome)
• Rare: trisomy 18 (Edward syndrome)
• Trisomy 13 and 18 don’t commonly live beyond the age of 1.
• Other trisomy types aren’t really a thing as most would be terminated before birth
Why does incidence of trisomies increase with maternal age?
• Incidence of some trisomies increase with maternal age as there is increased non-disjunction during oogenesis as developing eggs are held in meiosis for a long period of time
o Division only finishes when fertilisation occurs
o The longer the eggs are paused at prophase I, the more likely that non-disjunction occurs
Describe why trisomy 21 can occur
Trisomy-21: Down syndrome (2n=47)
• Most common type of trisomy- 1/2000 live births
• During oogenesis, chromosome 21 usually has a single crossover
o If no chiasma forms, both homologues segregate to same pole at Meiosis I
o Ocasionally, chiasma at extreme ends does not provide enough resistance to tension from spindle, leading to premature pulling to the poles- recombination provides an extra timeframe to prevent premature pulling
• In 5% of cases of trisomy 21, part or all of chromosome 21 is attached to another chromosome
What is a translocation?
o Translocations-partial rearrangements where an individual that carries them could be normal but can have partial sterility (due to unbalanced meiotic products)
How do translocations occur and what are 3 different types of translocations?
When segments of chromosomes break off, and attach to or exchange with other non-homologous chromosomes. Different types of translocation include:
- Reciprocal
- Non-reciprocal
- Robertsonian translocations
What is a reciprocal translocation? Are the gametes always viable?
o A segment from one chromosome is exchanged with a segment from another, non-homologous chromosome, creating two translocation chromosomes
Inviable gametes due to imbalance of genetic material- When translocated and normal chromosomes are mixed
Viable gametes- when two translocated chromosomes or two normal chromosomes are in a gamete
What is a non-reciprocal translocation?
o One piece of chromosome attaches to the other with no exchange
What are Robertsonian translocations?
o Long arms of two telocentric chromosomes fuse, forming a single meta- or acrocentric chromosome
o Translocation of chromosome 29 to chromosome 1 is relatively common in European breed cattle and results in reduced fertility
o Human chromosome 2 is a result of end-to-end fusion of 2 ancestral chromosomes.
All members of Hominidae (great apes) except humans, Neanderthals and denisovans have 24 pairs of chromosomes
What are 2 types of chromosomal rearrangement, and whatis the impact of chromosomal rearrangment?
-Translocation
-Inversions
• Individuals carrying chromosome rearrangements may be phenotypically normal, but have partial sterility
o Produce unbalanced meiotic products that cause zygotes to die
What is the ratio for codominance or incomplete dominance
1:2:1
What are linked genes?
- Genes located on the same chromosome (called syntenic genes)
- Genes that are close together on a chromosome such that their alleles are often inherited together are considered to be linked genes
When are genes unlinked?
-When they are on different chromosome or are far apart enough on one chromosome
How do unliked genes assort?
Independently
How do linked genes assort?
o Linked genes are not assorted independently
What was the first genetic map and who was it created by?
o First genetic map was Drosophila melanogaster X chromosome map created by Sturtevant (Morgan’s student)
How does recombination impact linked genes, gametesand genetic variation?
- Recombination (crossing over) seperates alleles of linked genes
- Recombination increases genetic variation by producing new alleleic combinations
- Crossing over creates parental and recombinant gametes after segregation
- This occurs more frequently between genes that are far apart
When does recombination occur?
- Occurs during prophase I of meiosis
- Crossing over occurs when chromatids between two homologous chromosomes exchange DNA
- This occurs more frequently between genes that are far apart
How do you figure out if genes are linked or unlinked? What do you do?
• Make a test cross to figure out if genes are linked or unlinked
How do you make a dihybrid test cross? What is the process and result?
o Dihybrid test cross- F1 heterozygous X homozygous recessive individual
Only one type of gamete is produced by the double recessive individual- the same recessive alleles are passed on to all offspring
Four types of gametes are produced from the heterozygous parent phenotype of the test cross progeny are derived from the alleles in the gametes of the heterozygous parent
• Two gametes are parental and two are recombinant
What is the results of the dihybrid test cross if the genes are linked?
• Higher ratio of parental phenotypes (resemble the parents) and a lower ratio of recombinant phenotypes (different phenotypes to the parents) if the two genes are linked
o Linked genes don’t have 9:3:3:1 ratio in dihybrid cross, and no 1:1:1:1 ratio in a test dihybrid cross
o The further two genes are apart, the more likely there will be more recombination between them and thus more recombination progeny
What is the result of the dihybrid test cross if two genes are unlinked?
• If two genes are unlinked, expect 1:1:1:1 equal ratio in test cross
How can we measure the genetic distance between linked genes?
Recombination rate
• Recombination frequency can be used to determine the arrangement of elements along a chromosome
What is the formula (and units) for recombination frequency?
• Recombination frequency in map units (m.u) or centiMorgans (cM)= (number of recombinant offspring/Total number of offspring)*100
How do genes more than 50 map units apart assort?
o Genes >50 map units apart have so much recombination between them that they are effectively unlinked an assort independently
• Cannot distinguish recombination frequencies of more than 50% from genes that are assorting independently on separate chromosomes
How do you calculate map distance?
- Generate a heterozygote
- Perform a test cross
- Identify parental and recombinant offspring
- Calculate recombination frequency
What is a genetic linkage map?
o Genetic linkage map- estimation of relative locations of phenotypic or molecular markers along a chromosome
How do we link a DNA sequence to a phenotype?
• Use genetic mapping to link phenotypes back to sequence changes in the genome
• Connect genetic linkage maps to physical maps (DNA sequence)
o Map the gene using genetic techniques then use molecular techniques to find physical DNA sequence responsible
Are molecular markers or phenotypic markers better for greater accuracy in genetic linkage maps?
Molecular markers preferred as there are only so many genes that we can reliably characterise a visible phenotype for -pad out the map more
Why is linkage analysis and genetic mapping useful?
o Identifying genes and biological processes
underlying inherited traits
o Functional genomics to discover why the gene causes the phenotype
o Genetic tests for diseases and potential cures
Linkage mapping is important in understanding human disease because can find genetic causes of familial traits that are passed on through generations
• BRCA1 genetic mapping of breast cancer on q arm of chromosome 17
If know genetic cause, can use that information to find a cure
o A scaffold or map to inform the assembly of whole genome sequences using genetic markers
What are linkage groups and how do they work?
- Diagram of linked markers along a chromosome
- Map distances are additive
- Larger values calculated as a sum of shorter intervals
What percentage can recombination frequency not exceed?
• Recombination frequency cannot exceed 50%
How can we make linkage groups more accurate?
• For greater accuracy, use shorter intervals (more markers) and more offspring
o Many thousands of molecular makers can be used to fill the gaps between the limited number of visible markers due to limited number of genes that provide usable phenotypes for mapping
What are molecular markers, how do they work and why are they useful?
o Are sites of silent DNA variation not associated with any measurable phenotypic variation
o Work the same way as visible markers- can be heterozygous, have recombination between them
o Are critical for understanding human variation and mapping positions of disease genes
Amount of variation present without any phenotypic variation is particularly high in humans compared to other organisms
How useful are molecular markers compared to phenotypic markers in bacteria? Why?
• In bacteria, vast majority of genome codes for protein- if there is mutations in bacterial DNA not as common to find one that doesn’t provide a phenotypic variation
o Most of the genome is exon, but a change could be a third base change, conservative substitution or redundant information
-Relatively useless
How useful are molecular markers compared to phenotypic markers in worms/flies? Why?
• In worms/flies, only 35% of genome is exon (codes for protein), but a change could be: o A third base change o A conservative substitution o Gene family o Duplicated pathways o Recessive alleles -Relatively useful
How useful are molecular markers compared to phenotypic markers in humans? Why?
• In humans, only 2% of the genome is exon, but a change could be: o A third base change o A conservative substitution o Gene family o Duplicated pathways o Recessive allele -Extremely useful
What are 3 types of common molecular markers?
o SNPs (single nucleotide polymorphisms) o RFLPs (restriction fragment length polymorphisms) o Microsatellites (short tandem repeats)
What are single nucleotide polymorphisms?
Single nucleotide differences in DNA sequence
Are SNPs common?
Most common type of molecular change between individuals
Do SNPs impact phenotype?
• Differences in DNA sequence may or may not be associated with differences in protein structure or phenotypes
How can SNPs be identified?
- Sequencing (e.g. whole genome sequencing) but is expensive
- Microarray chip (SNP array)-most common method
How can the microarray chip be used to detect SNP variation?
o Microarray chip containing spots of DNA oligonucleotide probes, to detect sites of known SNP variation in the genome
1. Attach a single stranded oligonucleotide (40-60 bp in length) corresponding to one variant to one feature on the microarray and single-stranded oligonucleotide corresponding to the other variant to another feature on the microarray
2. Prepare genomic DNA from the individual being tested
3. Label with fluorescent dye
4. Hybridise to the microarray (sites of known variation)
o If there is DNA binding between oligonucleotide and microarray , then there is fluorescence-means that SNPs match up and we know what SNPs the individual has by the number of fluorescent spots they have
What are RFLPs (restriction fragment length polymorphisms) and what do they result from?
Nucleotide differences that generate a change in a restriction enzyme cutting site
SNP variations can destroy or create restriction enzyme recognition sequences
How can restriction fragment length polymorphisms identify SNPs?
Differences in DNA cutting sites results in different sized bands after a restriction digest on agarose gel-> can identify whether SNPs occurred or not based on size of restriction digest
What are microsatellites?
2-5 nucleotides tandemly repeated in a DNA sequence
Why are microsatellites useful as a molecular marker?
These repeats occur in conserved regions of the genome so can use PCR primers to detect this region of the genome but within the microsatellite the repeat sequences will differ in number
Number of repeats in microsatellite DNA sequences are variable within the allele of individuals
Amplify microsatellite alleles with PCR
• Size of PCR product produced changes with number of microsatellite repeats
o Can be used to detect paternity
Describe how to perform a trihybrid test cross and what is involved in each step
o Step 1: Determine if the genes are linked
8 possible phenotypic combination
If not linked, then 1:1:1:1:1:1:1:1
If partially linked, see 4 big number categories and 4 small ones
If linked, then there is deviation in that ratio
• Three categories
o Highest frequency category- parental test cross phenotypes (because recombination is rare event)
o Intermediate frequency category-Single recombination event
o Lowest frequency category-Double recombination event (middle marker has switched chromosomes independent of the ages)
o Step 2: Determine gene order
Look for double crossover classes- the classes with the fewest recombinants (the lowest number)
The marker that is different from the parental configuration is the middle marker
o Step 3: Look at markers in pairwise configurations
Factor in double recombination events between flanking markers
• Double recombinants added in calculation twice
o Don’t do this if there is no clear double-recombinant class
Unlinked genes give 50 or more map units
o Step 4: Put the map together
Why are trihybrid test crosses done?
o Determine if 3 genes are linked, the gene order and the map distance
Describe a trick which may be used in exams through notation of linkage in crosses
Notation in exams-
• Even if notation is linkage location:
o Do not assume all genes are linked
o Do not assume that the order of writing is the order of the genes on the chromosome
Why can recombination rate vary throughout the genome?
o Chromatin structure
o Sex differences
o Chromosome aberrations
What are hotspots?
Hotspots-high levels of recombination
• Greater genetic distance relative to physical distance
What are coldspots?
Coldspots-low levels of recombination
• Shorter genetic distance relative to physical distance
Why do hotspots and coldspots exist?
o DNA in coldspots doesn’t cross over so well due to the proteins surrounding that DNA e.g. genes separated by a centromere
——Pericentric heterochromatin
May be due to the feasibility of DNA in those regions to initiate crossing over
What is pericentric heterochromatin?
Pericentric heterochromatin-condensed chromatin at the centromere which lacks standard meiotic recombination (no recombination here)
Are recombination rates the same for both men and women?
Recombination rates differ between males and females for most animals
Describe the recombination rates in male and female drosophila, and why these rates differ
• In male fruit flies, there is no crossing over at all
o Unusual prophase I of meiosis in which no chiasmata develop presumably due to lack of expression of recombination proteins which usually help this recombination to occur
Describe the recombination rates in male and female humans, and why these rates differ
• Genetic distance and maps on human male and female chromosomes can look different
o Females have slightly more recombination than males
o Probably because males vs females have different tissues and different gamete production rates
What is inversion?
Inversion- a type of chromosome aberration, a structural variation in a chromosome leading to rearrangement of chromosome segments
• Piece of chromosome breaks up and flips around and inserts in the opposite orientation of its original orientation
Describe the impact of inversion on sterility
Heterozygous individuals for inversion chromosomes may be phenotypically normal, but have partial sterility
• Produce unbalanced meiotic products if recombination occurs within the inversion
What are the two types of inversions?
Inversions can be paracentric (include centromere) or pericentric (no centromere)
Describe how recombination occurs in an inversion heterozygote, the products of this recombination and what can change the products
• During meiosis, for genes to align together, a loop needs to form (all markers paired up with each other)
• Products of single recombination mean that there are ½ viable and ½ inviable chromomes
o Dicentric-Have two centromeres
o Acentric-Have no centromeres
• Double recombination events can restore viability to a chromosome
What is a physical map?
• Physical map- position of markers in the DNA sequence (in megabases)
What is a genetic map?
• Genetic map- position of markers in linkage group (in map units)
What is the recombination rate and how do we interpret a graph of recombination rate?
• Recombination rate (megabases per map unit)- differs along chromosomes
o Linear line- no recombination hotspots/coldspots in chromosome
o Non-linear line: recombination results in high rate whilst no recombination results in stationary points
What does genetic mapping of traits use?
• Genetic mapping of traits uses natural genetic variation of individuals in a population
What is a haplotype, how do they happen and what can they be?
• Haplotype- a group of alleles that are inherited together throughout generations
o A block of alleles in a set of linked loci that are inherited together
o These alleles will tend to be passed on together during meiosis, unless separated by recombination
o May consist of any type of genetic variation:
SNPs
Phenotypic markers
RFLPs
Microsatellites
What usually happens to a causal mutation that leads to a trait throughout the generations in terms of inheritance patterns?
• A causal mutation that leads to trait:
o Inherited with other nearby variants on the chromosome
o Recombination can generate blocks of variation that tend to be inherited with a mutation of interest
o Recombination reduces large blocks transmitted through the pedigree
o Variation that happened to be linked to a causative mutation will still be associated with the mutant many generations later
What are ancestry haplogroups and what can be used to detemrine them?
o Ancestry haplogroups- A group of similar haplotypes that share a common ancestor
Indicate likely similar origin or line of descent
Genome-wide SNP analysis can determine ancestry haplogroups
How can the fact that linked alleles and haplotypes stay together for generations be used to identify variation linked to phenotypic traits?
Positional cloning
Genome Wide Association Studies (GWAS)
Describe the purpose and concept of positional cloning
• Technique (used a lot in the 1980s) to locate position of a gene along the chromosome
• Use genetic mapping to connect between the gene that affects the phenotype and the DNA sequence
o DNA sequence for a gene can be identified based on its genetic map position
• Mapping done by observing individuals in a pedigree, checking suite of molecular markers for these individuals and finding area of the genome which always comes up with same markers that are variable in mutated individuals. Then, can look at region of DNA between them
Describe how positional cloning is performed
• Step 1: Gene of interest is mapped between two cloned markers
o Map the phenotype by recombination between known molecular markers
• Step 2: The DNA between the markers is isolated and analysed
• Step 3: The compatible candidate gene is identified based on
o Expression pattern (expression at right tissues at right time)
o Evolutionary comparisons-compare with other organisms to see if gene has the same function in those other organisms
o Sequence analysis-see if it differs from wild type individual
Describe an example of positional cloning performed to map a disease
• First example of positional cloning is mapping of cystic fibrosis gene in 1989
Cystic fibrosis was mapped to a region on 7q31
A region that was conserved in mammals was isolated
A cDNA from this region was isolated
This cDNA was used to probe for transcripts from tissues affected in CF patients
Found that gene expressed in all tissues affected by cystic fibrosis and became candidate for causing the disease
Describe the genetic basis of cystic fibrosis
o Common autosomal recessive disorder, affecting lungs and other organs and tissues that produce sweat, digestive fluids or mucus
o Extensive pedigrees available
o Gene located on chromosome 7 in 1000bp interval
Describe how genome wide association studies are performed and their purpose
- Sequencing is more accessible
- SNPs used
- Compare genome-wide SNP variation in of thousands of affected vs unaffected individuals
- Relative frequencies of each SNP and haplotype are calculated and plotted among affected and unaffected individuals
- Can identify SNPs associated with a disease, but they cannot specify which genes are causal
- SNPs and haplotypes that are more common among affected individuals identify candidate locations for causative alleles for the trait
- Genomic locations can be surveyed for candidate causative genes based on gene function, expression patterns, similarity to mutants in other animals…
How can GWAS-Manhattan plots be interpreted?
• Interpreting GWAS-Manhattan plots
o Each dot represents a haplotype, a region containing variable SNPs
o Each number on the X axis represents a chromosome
o Each colour indicates when chromosome changes
o Probability that a haplotype is associated with the trait is located on the y axis
o SNPs/haplotypes with a significant association with the trait are highlighted in a different colour (green)
How can genes be surveyed as candidate causative genes?
o Expression pattern- from transcript analysis such as RNA sequencing and microarrays
Which genes are expressed in the appropriate time and location
o Evolutionary comparison
Has the gene been identified with a phenotype in other closely related organisms?
o Mutations and polymorphisms- Do people affected by the trait share a DNA sequence variation in one of these genes?
What is the use of insecticides and how do they work?
• Modern agricultural practices rely on chemical methods of pest control
• Compounds that act as insecticides have diverse chemical structures with different modes of action but a lot of them target nervous system of insects
o E.g. pyrethrins, neonicotinoids, DDT, dieldrin
Disrupt nervous system of arthropods, causing paralysis and death
What are the problems of insecticide use?
• Problem of insecticide use-
o Puts pest species under intense selective pressure
o Variants containing mutations in genes that provide resistance (ability to tolerate insecticide) are favoured
o Resistance is selected for in a population and becomes widespread
Describe dieldrin and the problem with its use
o First used in 1950s
o Insect-specific neurotoxic chemical
o Dieldrin- a synthetic chemical with high insecticidal activity
o Resistant populations developed quickly
Present in pests of medical and agricultural significance
Crop pests and vectors of disease
Describe the resistance capability of insects with the Rdl gene depending on their genotype
o Unknown cause of the resistance phenotype
Resistant homozygotes survive much higher doses (4000 fold higher doses) than wild-type homozygotes
Heterozygotes survive at intermediate levels
What is the downside of an insect being resistant to dieldrin?
o Resistance to this chemical shows temperature sensitivity
When exposed to high temperatures (380C), resistant homozygotes (RR) exhibit higher sensitivity than wild-type homozygotes (SS)
• Heterozygotes (RS) have intermediate levels of temperature sensitivity
Resistant flies will go into heat shock (go to sleep in a while and eventually die)
Recovery times of resistant mutants is much longer (10 minutes) than wild type flies (5 minutes)
Describe how the mutation responsible for dieldrin resistance was found
• Backcross and linkage mapping with 2 markers
o Bristle phenotype- stubble (position 58.5)
o Lyra wings (position 40.5)
• Rdl (resistance to dieldrin) was mapped on position 25 on chromosome 3
Further refining of location of causal mutation with positional cloning
• Screened lots of different markers along chromosome to lead to identification of region containing the resistance gene
• Resistance mapped to a locus containing the first identified insect ligand-gated chloride channel gene
Where is the Rdl gene and what does it code for?
- Rdl (resistance to dieldrin) was mapped on position 25 on chromosome 3
- Resistance mapped to a locus containing the first identified insect ligand-gated chloride channel gene
Describe the molecular function of Rdl gene in wild-type individuals
Normal-
• Ligand-gated chloride channel with large extracellular domain that binds to GABA (an excitatory ligand) and allows for chloride ion influx to flow through
o Five Rdl subunits join together to make a chloride channel pore in the membrane
What is the impact of insecticide dieldrin on the the Rdl gene?
• Insecticide dieldrin blocks chloride influx
Why are mutants in the Rdl gene resistant to dieldrin?
• Single nucleotide polymorphism (SNP)-> G to T mutation which results in an amino acid change in transmembrane region of the protein (alanine to serine substitution at position 301)
• Mutant receptor no longer binds to dieldrin
o Chloride channel is not blocked by insecticide
o Insects survive
Describe how it can be observed whether insect populations have the Rdl mutant gene for dieldrin resistance.
o Mutation screening-see how many populations around the world have this mutation
G to T SNP that leads to resistance generates an RFLP and removes HaeII recognition site
PCR performed on fragment
Restriction enzyme HaeII cuts twice in wild type individuals but in resistance mutants only gets cut once: therefore different banding pattern can be seen
• Uncut fragment- 268 bp
• Homozygous resistant- 205 and 63 bp
• Homozygous susceptible mutant- 3 bands
• Heterozygote- all bands present in either allele
HaeII method used to recognise mutants around the world
Describe the Tasmanian devil in terms of:
- type of mammal
- location
- chromosomes
• Tasmanian devil-
o World’s largest carnivorous marsupial, distribution now restricted to Tasmania
o Has 7 different chromosomes
What is the Tasmanian devil face tumour disease and how and when was it spread?
• Tasmanian devil face tumour disease-
o Transmissible cancer that can spread like a contagious disease
o Spread by the transfer of living cancer cells by biting
o Spread throughout Tasmania since first identified in 1996
How do we known that the tasmanian devil face tumour disease is a transmissable cancer?
o Fatal transmissible cancer is decimating the devil population in Tasmania
All DFTD cells are genetically identical to each other and genetically distinct from their hosts own cell
• Seen by using genetic markers and karyotyping
How did the tasmanin devil face tumour disease originate?
Cancer originated from a single individual and spread, rather than arising repeatedly
• First strain in 1996 (female origin)
• New strain identified in 2015 (male origin)
o Tumor cells must have arisen from a female because they contain X-chromosome fragments
Describe the karyotype of tumour cells in tasmanian devil face tumour disease when compared to normal cells
o There are translocations, deletions, rearrangements…
o Chromosome 1 and sex chromosomes missing from tumor karyotypes
Translocations and joining from different chromosomes
o Chromosome 5 aneuploid
o Four additional marker chromosomes (M1-M4) are present, made up of chromosome fragment translocations, probably inversions and other chromosome abnormalities
Not additional chromosomes-just have been formed from bits of pre-existing chromosomes
How were karyotype differences highlighted in tumour cells in tasmanian devil face tumour disease?
o Fluorescence labelling of chromosomes highlights karyotype differences
Label then look under a microscope
Describe the process of Crawford et al’s 2017 study:
Loci associated with skin pigmentation identified in African populations
• Genome-wide association study and functional characterisation of novel genes involved in human skin pigmentation
• Studies often focus on European lineages
• Large diversity of pigmentation phenotypes within Africa
o Melanin index- skin tone measurements
o Erythemal dose- level of radiation required to turn skin red
• Compared populations to each other using those phenotypes
• Look at Africans as baseline for rest of population
o Measure phenotypic diversity (pigmentation levels)
o From blood, isolate DNA
Analysed 1,570 DNA samples of Africans with diverse pigmentation phenotypes
o Genotyping using SNP array
Illumina Omni5M exome array about 4.3 million SNPs
• SNPs from international HapMap and 1000 genomes projects
o Association between genetic variation and the trait
Manhattan plot of GWAS
Then functionally characterised each gene to make sure they had function for skin pigmentation-functional gene experiments necessary to confirm causative genes once associations have been found
Describe the results of Crawford et al’s 2017 study
• GWAS identified novel variants associated with skin pigmentation
• Significant variants clustered in 4 genomic regions that together account for about 30% of the phenotypic variation
• Four regions identified account f about 30% of variation in skin pigmentation phenotype
o SLC24A5, MFSD12, DDBI and HERC2
• Proved phenotype of candidate genes with functional experiments in model organisms
o MFSD12- codes for a protein that modifies pigmentation in human melanocytes
Decreased MFSD12 expression associated with darker pigmentation
Genetic knockouts of MFSD12 orthologues affect pigmentation in mice
• Identified previously uncharacterised genes associated with skin pigmentation in ethically diverse Africans
What are gene interactions possibly involved in skin pigmentation?
o Gene interactions possibly involved:
Complementary gene interactions (both genes needed for phenotype)
Duplicate gene interaction (either gene needed for phenotype/redundancy)
Dominant gene interaction (genes with same phenotype interact additively)
Epistasis (one gene stops/masks the phenotype of another)
• These genes have diverse functions
o Repairing UV damage: melanogenesis in skin cells
What is the type of evidence necessary to confirm causative genes once an association has been identified?
- Gene expression analysis
- Molecular function of candidate gene
- Functional experiments in model organisms
