BCOR 101, WEEK 10
BCOR 101, WEEK 10 BCOR 101
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This 52 page Class Notes was uploaded by Katarina Fielding on Sunday April 24, 2016. The Class Notes belongs to BCOR 101 at University of Vermont taught by Amanda Yonan in Spring 2016. Since its upload, it has received 9 views. For similar materials see Genetics in BioInformatics at University of Vermont.
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Date Created: 04/24/16
Linkage and Chromosome Mapping in Eukaryotes Chapter Five Linkage • Most chromosomes carry a lot of genes • Unit of inheritance through meiosis is a chromosome, not each individual gene • Therefore, sometimes two genes are going to be inherited together because they are so close physically on the same chromosome – Breaking Mendel’s 2Law – Independent assortment of any two genes Complete Linkage • When two genes are always inherited together • Can only produce two gametes: AB (50%) and ab (50%) • Determined by which allele resides on which chromosome in parent • Gametes are known as “parental gametes” because their alleles are exactly the same as the parent’s alleles Complete Linkage YyRr Two Gametes YR (½) yr (½) YR YYRR YyRr Two Phenotypes YyRr (½) ¼ ¼ 3 : 1 yr YyRr yellowgreen (½) roundwrinkled ¼ yyr¼ When genes are transmitted together Crossing Over • Occurs during Prophase I • Homologous chromosomes exchange alleles = Recombination • Crossing over introduces independent assortment between two genes on the same chromosome • However, percentage of crossing over is determined by physical distance between two genes Crossing Over Percent of crossing over is determined by physical distance between two genes: • Closer two genes lay – less likely crossing over can occur – Genes will be linked = inherited together • More distance between two genes – more likely crossing over can occur – Genes may be unlinked – Depends on percent recombination Independent Assortment • With two completely independent genes expect four gametes to be made: AB (25%), Ab (25%), aB (25%), ab (25%) • Two genes on the same chromosome, but that lay far apart will undergo crossing over as often as not – 50% chance of recombination • Thereby producing four gametes in equal amounts = unlinked Independent Assortment YyRr 4 different gametes YR Yr yR yr (¼) (¼) (¼) (¼) Genetic Linkage • Determined by percent recombination • Any recombination less than 50% is considered linked – Only 0% recombination is complete linkage • In reality complete linkage is very rare • Between 0-50% recombination reflects physical distance between genes • Never see more than 50% - why not? Genetic Distance • Frequency of crossing over tells us how far apart two genes are on chromosome • Not exactly the same as physical distance because certain sequences lead to more or less crossing over – Recombination hot spots • Instead, recombination measures genetic distance between two genes – Is correlated to physical distance, but not same Why does the frequency change? Morgan’s hypothesis: two genes located close together on same chromosome were less likely to have a chiasma between them • Therefore chance of crossing over depends on physical distance between genes • This idea allows mapping of genes • Once you know percent of recombination you can determine genetic distance Genetic Mapping • Mapping is determining the order and distance between genes • Genetic mapping is done through breeding and counting offspring • Now a genetic map can be compared to a physical map determined from sequencing – Correlation has been confirmed • Genetic maps calculate chance genes will nd break Mendel’s 2 Law – inherited together Mapping Genetic Distance Recombination seen between genes: 1. yellow-white 0.5% 2. yellow-miniature 35.4% 3. white-miniature 34.5% Obviously: yellow and white genes are closest together • Yellow and miniature are furthest apart • White lays in the middle Mapping Genetic Distance Recombination seen between genes: 1. yellow-white 0.5% 2. yellow-miniature 35.4% 3. white-miniature 34.5% Mapping Genetic Distance • Basic principle still used today to calculate map distance (cM) between genes Based on a few facts: 1. Limited number of chiasmata form during each meiosis 2. Position of chiasmata is random along the length of each chromosome Therefore likelihood of crossing over is less between loci that are close together Likelihood of Crossing Over Recombinant Gametes Out of four gametes produced: • Only two will reflect crossing over event – Producing recombinant offspring Mapping Genetic Distance • Count number of parental and recombinant offspring Recombinant = % recombination Total Offspring • 1% recombination = 1 cM ex blond hair, blue eyes x Brown Brown 3 = Brown blue 44 = blond blue 5 = blond Brown 48 = Brown Brown Single v. Double Crossovers One long chromosome can contain more than one chiasma: • Single crossover (SCO) = one chiasma between two genes • Double crossover (DCO) = two chiasma between three genes • Still only produce two recombinant gametes Product Rule • Product rule determines the chance of two independent events occurring simultaneously • Combined probability is calculated by multiplying individual probabilities • Probability of double crossover (DCO) is equal to probabilities of each single crossover (SCO) multiplied together Mapping Three Genes To discover the order and distance between three genes at once: • Must account for the presence of all: – Single crossovers (SCO) Between each combination of two genes – Double crossovers (DCO) • Must have heterozygous genotypes – Otherwise can’t see crossing over Mapping Three Genes Let’s work through mapping example in Drosophila with three genes: • Don’t know order of genes but have to assume order just to draw out picture – Do know that these genes exist on X chrom. Mapping Three Genes F1: • Males produce two gametes – Cannot have crossing over because Y chromosome doesn’t synapse to X • Females can produce eight different gametes due to SCOs and DCOs Mapping Three Genes • Which gametes do you expect to be most frequent? • Which gametes are least frequent? • How will we determine map distance? And order of 3 genes? Mapping Three Genes Total: 10,000 Mapping Three Genes • Determine order of three genes based on the DCO compared to NCO’s phase – w must be in middle • Determine map distance by % recombinant Mapping Example in Maize • Another thing that needs to be determined in linkage mapping is phase Phase: which alleles are inherited together on the same homologue • Heterozygotes have two different phases: – AB/ab – Ab/aB • Also known as chromosomal alignment Determining Phase • How can we determine phase? • Look at results of cross: most common = NCO Determining Order • How do we determine the order of genes? • Look at results of cross: least common = DCO Determining Order DCO = pr v bm + + + Possible order (knowing phase): + v bm v + bm v bm + pr + + + pr + + + pr Determining Genetic Distance • How do we calculate cM between genes? • Look at results of cross: cM = % Recombinant (161 + 86)/1109= 0.223 or 22.3 cM (395 + 86)/1109= 0.433 or 43.4 cM Mapping Example in Maize • Which genes are 22.3 cM apart and which genes are 43.4 cM apart? v + bm + v pr + pr + v + bm + pr bm + pr + Undetected Crossovers • Some crossovers are impossible to detect • A small number of offspring might make it impossible to see a rare phenotypic class • Genetic maps are most accurate between genes that are closely linked Interference • A crossover event in one region inhibits a second crossover in nearby regions • Because of physical constraints that prevent chiasmata forming close together • Quantified by comparing expected double crossovers to observed DCOs: Observed DCO Expected DCO Interference • Observed DCO – simply count offspring • Expected DCO = SCO x SCO – Assuming chance of each SCO is independent of other SCO – Use product rule to determine chance of DCO • This ratio gives you an idea of the amount of expected crossovers that were seen • Interference = 1 – Observed DCO Expected DCO Synteny Mapping • Without sequencing entire genomes human genes were mapped to chromosomes • Produce a hybrid cell line that contains some human chromosomes – Know which human chromosomes present • Check to see if cell line produces a protein – If so: then gene must be on remaining chrom’s – If not: then gene must not be on these chrom’s Synteny Mapping • Use data below to map Genes A and B: Gene A must be? Gene B must be? Gene D must be? Mapping in Haploids • Haploid organisms usually replicate through mitosis exact copies • Some go through a diploid zygote stage where crossing over can be seen • Diploid zygote undergoes meiosis into a spore (acsi) containing four haploid cells – Known as a tetrad • Examining the alleles in each cell (of four) allows genetic mapping Mapping Haploids w/Tetrads Three types of tetrads can be made: 1. Parental type (PD) 2. Non-parental type (NPD) 3. Tetratrype (T) PD = same allele combinations as parents NPD = all genotypes are different than parents (recombination of chromosomes) T = two genotypes are parental, two are recombinant Tetrad Analysis Tetrad Analysis If PD = NPD genes are unlinked If PD >> NPD genes are linked • Calculate map distance: NPD + ½ T RF = Total number of tetrads • RF is a percent, multiply by 100 to get cM Questions? Discussion Thomas Hunt Morgan • Columbia University • Drosophila melanogaster • Saw different percentages of offspring phenotypes would occur depending on which two genes he was studying Questions: 1. What is causing different phenotypes? 2. Why does the frequency change? What is causing different (recombinant) phenotypes? • Crossing over between genes • Introduces new allele combination • New phenotypes not seen in parents • Morgan was the first person to describe crossing over Between synapsed homologues during Prophase I LOD score • Log of Odds ratio • Measures the probability that two traits are linked in humans (Can’t set up cross, small number of offspring) • Data taken from multiple, large pedigrees (likelihood of genotype/phenotype data assuming linkage) LOD = lo10 (likelihood of genotype/phenotype data assuming no linkage) • LOD ≥ 3 indicates linkage between genes Physical Mapping • With recombinant DNA technology mapping genes done on a physical map DNA markers: polymorphisms whose location in the genome is known • Use linkage testing to discover a marker that is linked to a gene • Since you know the location of the marker, you now know the location of the gene Physical Mapping • Linked genes that are too close together to follow Mendel’s 2 ndlaw Disease • They will always go into the Marker same gamete in meiosis • Use linkage to known markers to identify disease causative genes Common Markers • RFLP – Restriction Fragment Length Polymorphism – Polymorphic sequence at cut sites • STR – Short tandem repeats (Microsatellites) – Different number of repeating units • SNP – Single nucleotide polymorphism – Different base at one specific location Sequencing a Genome • The ultimate gene mapping experiment • Determine the exact sequence of an entire genome for an organism • Produce a physical map with location and physical distance between all genes – Compare/contrast to genetic map • Sequenced genomes: – Human, Drosophila, mouse, etc Mendel and Linkage • Linkage breaks Mendel’s 2ndLaw – Independent assortment of any two genes • Why didn’t Mendel ever see linkage? • Studied 7 traits in an organism with 7 chromosomes – was he lucky? • No - physical maps show us that some of his traits were on same chromosome Mendel and Linkage • Genes v and le are physically and genetically linked (Chromosome 4) • Mendel did not report that v-le showed independent assortment – Did not study every gene combination • Do you think: – This is a gene combination he did not study? – Or that Mendel saw these two genes didn’t show independence and left out the results? Sister Chromatid Crossing Over • Can crossing over happen between sister chromatid – held together by centromere? • Experiments have shown yes • Because chromatids have same exact alleles you can’t see the effect in offspring • Does actually matter at all?
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