Genetic Diversity in Eukaryotes

Genetic Diversity in Eukaryotes

• Sexual reproduction generates a high level of species diversity
• Individual offspring will be genetically different from each other and their parents because that diversity is driven by independent assortment of chromosomes and homologous recombination during meiosis

Homologous Pairs of Chromosomes Segregation

• Homologous pairs of chromosomes segregate at meiosis to form haploid gametes (sex cells)

Independent Assortment

• Humans are diploid, containing 23 pairs of chromosomes
• Meiosis randomly incorporates one of each pair into each gamete (sperm or egg cell)
• The potential number of unique gametes from each individual human is 2^23

Crossing Over

• Crossing over of chromatids during metaphase I of meiosis increases genetic diversity
• During metaphase I, homologous chromosomes pair up, and non-sister chromatids cross over
• This crossing over allows exchange of genetic material between the two different parent chromosomes
• Cross-overs are called chiasma (pl. chiasmata)
• Homologous recombination is the process in which an exchange occurs

Homologous Recombination

• Homologous recombination is reciprocal and precise and involves the reciprocal exchange of nearly identical segments between non-sister chromatids (but may carry different alleles of the genes)
• Each chromatid contains a double-stranded DNA molecule
• It ensures precise exchange so that no genes are gained or lost, and no nucleotide added or lost at the crossover point if it occurs within a gene
• Typically, one to three chiasmata form between non-sister chromatids during human meiosis

Recombination

• Homologous recombination generates new combinations of alleles on a chromatid that did not exist in the parent
• Recombination is more likely if the two genes are far apart on the chromosome
• Crossing over during meiosis is a result of 2 out of the 4 gametes
• Genes that are close together are less likely to have a recombination event between them, resulting in resultant alleles not being recombined

Genetic Linkage

• Cross overs can happen at any point on a chromosome, but the closer together two genes are, a cross over will have less chance of occurring during meiosis. The converse of this is the more likely the two alleles of those genes will be inherited together
• When genes are inherited together, they are considered LINKED, also known as genetic linkage

Summary 1

• Genetics diversity in eukaryotes is achieved by independent segregation and genetic recombination
• In meiosis I, parental sister chromatids can form chiasmata and exchange genetic information resulting in recombination
• Recombination between two loci or genes on the same chromosome is more likely the further apart they are
• Loci that are closer together are less likely to recombine and are LINKED

Mapping

• The number of recombination events between two linked genes can estimate their physical distance

Drosophila Mutant Phenotypes

• Mutated variants of Drosophila, a type of fly, include lobed eyes, vestigial wings, and black body color

Dihybrid Test Cross

• A dihybrid test cross is used to calculate distance between 2 genes on the same chromosome
• F1 generation normal wings and normal eyes are the result of wild type/True breeding mutation cross

Results of Dihybrid Test Cross

• W and E genes are linked and on the same chromosome
• Under independent assortment you would get equal numbers of each phenotype (1:1:1:1), and the less frequent phenotypes represent recombinants because crossing over is a result of the alleles having recombined
• Recombination frequency (RF) calculates the physical map distance between 2 genes and is equal to the total number of recombinants divided by the total progeny and expressed as a percentage

Thomas Hunt Morgan

• American Scientist and Nobel Prize winner in medicine in 1933 for his work on chromosomes and heredity
• Used fruit flies (drosophila) as a model organism for genetic studies
• The centiMorgan was named after him by his student
• One centimorgan is the length of DNA where recombination occurs once in every hundred attempts

Summary 2: Dihybrid Test Cross

• In the second generation (F2), progeny of parental types indicate the W and E genes are LINKED on the same chromosome and the W and E genes are segregating together (not independently)
• Less frequent phenotypes mean recombinants are present and the alleles have recombined
• Infrequent recombination events indicate distance can be calculated in order to produce a genetic map of these 2 genes

Calculating Map Distance

• The recombination frequency formula is recombination frequency (RF) = (number of recombinants / total number progeny) x 100
• This expresses the physical map distance, as indicated by map units

Trihybrid Test Cross

• Distance is calculated by working out gene order of three genes
• An example includes defining the location of W or E (5cM) and B (18.5cM) from W

Results of Trihybrid Test Cross

• The W and E genes for the trihybrid test cross are on the same chromosome
• As progeny inherit a chromosome from the true breeding mutant with only recessive alleles we can consider the chromosome from the heterozygous parent as this will predict the phenotype
• Double recombinants are the result of two cross-over events on the same chromosome
• These occur when there is a smaller number of progeny as these are even rarer than the progeny that result from the single cross over between 2 genes

Gene Order

• Identifying double recombinants assists in defining gene order
• With a double recombinant involving three genes, only the middle gene is switched between chromatids

Summary 3

• Dihybrid test cross: measurement of recombination fraction between pairs of genes estimates the distances
• Limitation: This does not determine the linear order of genes on the chromosome.
• The Trihybrid test cross involves 3 genes to establish their linear order.
• The double recombination events are a result of the middle gene is being swapped between chromatids
• Flanking genes remain in place confirming the order

Calculating Recombination Frequency

• Double recombinants indicate gene order
• The goal is to identify single recombinants for distance calculations
• A single recombination uses a single cross over
• Calculation of distance between E and W: RF is calculated for recombinant progeny with E and W
• Calculation of distance between W and B: RF is calculated for recombinant progeny with W and B
• The middle recombinant indicates that the distance is the total of both distances

Importance of Mapping

• Alfred Sturtevant, a student of Thomas Hunt Morgan, used Drosophila to use variations in the strength of linkage between alleles of different genes to calculate the distance and gene order on a chromosome
• The subsequent paper published in 1913 defined the first genetic map of a chromosome

Drosophila Genome

• The map shows key genes on the maps of the drosophila chromosomes
• Examples include genes for eye, wing, and body color

Human Disease Mapping

• Chromosome mapping has been used in Humans using linkage and recombination frequency principles to find disease genes
• This is much more difficult compared to conducting it on flies because there are fewer progeny, there are limitations to creating experimental crosses and human phenotypes are much more complex being controlled by multiple genes
• However, rare inherited human phenotypes offer a way to study naturally occurring Human mutants

Dihybrid-Cross in Humans

• Sex-determined traits provide a type of dihybrid-cross in humans, such as for phenotypes that occur in males or are X-linked.
• These are relatively easy to map to the X chromosome because the disease phenotype co-segregates with biological gender which are X-linked disorders

Mapping of the X Chromosome

• Haldane and Julia Bell studied red-green colorblindness and haemophilia were inherited together often than expected by chance.
• By observing them they calculated a 10% recombination frequency between the genes for color blindness and hemophilia
• This provided evidence that these traits are genetically linked and close to one another on the X chromosome

Mapping of Disease Genes

• Genetic linkage studies are used to map the location of disease genes in Humans in which polymorphic genetic markers, or sections of genes with variable patterns that follow the same inheritance rules.
• Haldane, Bell and Morgan led to way with modern linkage studies to be used in Humans for things such as cystic fibrosis

Linkage Analysis

• Linkage disease analysis involves sampling DNA samples to identify the alleles of a set of polymorphic genetic markers, indicate the inheritance of the disease, which highlights genes likely impacted by the disease
• This helps paved the way for an 'explosion' in gene discovery findings in the 1980s, with new collaboration and technology to place markers on specific Human chromosomes

Disease Identification

• These first genes identified were for Huntington's disease in 1983, Duchenne Muscular Dystrophy in 1983, Cystic Fibrosisin 1985, Adult Polycystic Kidney Disease in 1985, and Retinoblastoma in 1986
• Huntington's disease (HD) was initially described by George Huntington which was mapped to human chromosome 4 and mutations were seen to be caused with nucleotide repeats of the huntingtin (HTT) gene

The Human Genome Project

• In 2001 the human genome meant the locations and sequences of all 30,000+ genes were publicly available and searchable online
• This helps in finding a disease, where a location and sequences of genes in a family dramatically reduced the time to find a disease
• Whole-genome sequencing, which started in 2005, further reduced the time to find a gene
• Recombination mapping techniques were developed to track the phenotypes in families to finding markers for genes that co-segregation with the X chromosome