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Gene Linkage and Genetic Mapping

Mendel’s Laws: Chromosomes Homologous pairs of chromosomes: contain genes

whose information is often non-identical =alleles• Different alleles of the same gene segregate at

meiosis I• Alleles of different

genes assort independently in gametes

• Genes on the same chromosome exhibit linkage: inherited together

Gene Mapping

• Gene mapping determines the order of genes and the relative distances between them in map units

• 1 map unit=1 cM (centimorgan)• Alleles of two different genes on the same

chromosome are cis• Alleles of two different genes on different

homologues of the same chromosome are trans

Gene Mapping

• Gene mapping methods use recombination frequencies between alleles in order to determine the relative distances between them

• Recombination frequencies between genes are proportional to their distance apart

• Distance measurement: 1 map unit = 1 percent recombination

Gene Mapping

• Recombination between linked genes located on the same chromosome involves homologous crossing-over = allelic exchange betweenthem

• Recombination changes the allelic arrangement on homologous chromosomes = recombinant

Gene Mapping• Genes with recombination frequencies less

than 50 percent are on the same chromosome (linked)

• Two genes that undergo independent assortment have recombination frequency of 50 percent (or more?) and are located on nonhomologous chromosomes or far apart on the same chromosome (unlinked)

Recombination

• Recombination between linked genes occurs at the same frequency whether alleles are in cis or trans configuration

• Recombination frequency is specific for a particular pair of genes

• Recombination frequency increases with increasing distances between genes

Genetic Mapping

• Map distance between two genes = one half the average number of crossovers in that region

• Map distance=recombination frequency over short distances because all crossovers result in recombinant gametes

• Genetic map = linkage map = chromosome map

Genetic Mapping

• Linkage group = all known genes on a chromosome

• Physical distance does not always correlate with map distance; less recombination occurs in heterochromatin than euchromatin

• Locus=physical location of a gene on chromosome

Gene Mapping: Crossing Over

• Crossing-over between genes on homologous chromosomes changes the linkage arrangement of alleles on a single chromosome

• Two exchanges between the same chromatids result in a reciprocal exchange of the alleles in the region between the cross-over points

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Example: Trihybrid Mapping• Counts from: LSG/lsg x lsg/lsg• n=740

L S G 286 Parentall s g 272 ParentalL s g 4 r (L & SG)l S G 2 r (L & SG)L S g 59 r (G & SL)l s G 44 r (G & SL)L s G 40 r (S & GL)l S g 33 r (S & GL)

&

• Distance L to S: (40+33+4+2)/740 * 100 = 11.2 cM• Interference = 1-[f(doubles)/ f(single1) *f(single2)]

Gene Mapping: Crossing Over

• Cross-overs which occur outside the region between two genes will not alter their arrangement

• Double cross-overs restore the original allelic arrangement

• Cross-overs involving three pairs of alleles specify gene order = linear sequence of genes

Genetic vs. Physical Distance

• Map distances based on recombination frequencies are not a direct measurement of physical distance along a chromosome

• Recombination “hot spots” overestimate physical length

• Low rates in heterochromatin and centromeres underestimate actual physical length

Gene Mapping

• Mapping function: the relation between genetic map distance and the frequency of recombination

• Chromosome interference: cross-overs in one region decrease the probability of second cross-over

• Coefficient of coincidence=observed number of double recombinants divided by the expected number

Gene Mapping: Human Pedigrees

• Methods of recombinant DNA technology are used to map human chromosomes and locate genes

• Genes can then be cloned to determine structure and function

• Human pedigrees and DNA mapping are used to identify dominant and recessive disease genes

Gene Maps: Restriction Endonucleases

• Restriction endonucleases are used to map genes as they produce a unique set of fragments for a gene

• EcoR1 cuts ds DNA at the sequence = 5’-GAATTC-3’ wherever it occurs

• There are >100 restriction endonucleases in use, and each recognizes a specific sequence of DNA bases

Gene Maps: Restriction Enzymes

• Differences in DNA sequence generate different recognition sequences and DNA cleavage sites for specific restriction enzymes

• Two different genes will produce different fragment patterns when cut with the same restriction enzyme due to differences in DNA sequence

Gene Maps: Restriction Enzymes

• Polymorphism= relatively common genetic difference in a population

• Changes in DNA sequence = mutation may cause polymorphisms which alter the recognition sequences for restriction enzymes = restriction fragment length polymorphisms (RFLPs)

Gene Maps: Restriction Enzymes

• RFLPs can map near or in human genes • Genetic polymorphism resulting from a

tandemly repeated short DNA sequence = simple tandem repeat polymorphism (STRP)

• Most prevalent type of polymorphism is a single base pair difference = simple-nucleotide polymorphism (SNP)

• DNA chips can detect SNPs

Human Gene Mapping

• Human pedigrees can be analyzed for the inheritance pattern of different alleles of a gene based on differences in STRPs and SNPS

• Restriction enzyme cleavage of polymorphic alleles differing RFLP pattern produces different size fragments by gel electrophoresis

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Tetrad Analysis

• Meiotic spores held in asci (ascospores)

• Allows recovery of all products of meiosis

• Two types• Unordered tetrads (yeast)

• Usually allows gene to gene map distances

• Under rare circumstances, gene to centromere

• Ordered tetrads (neurospora)• Usually allows gene to centromere map distance

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Unordered Tetrads• Four kinds of tetrads

• Parental ditype (AB, AB, ab, ab)• Non-parental ditype (Ab, Ab, aB, aB)• Tetra-type (AB, Ab, aB, ab)

• When genes tightly linked• only parentals seen

• When genes unliked• parentals and non-parentals equal• tetratypes: gene-centromere X-over• gene-centromere map possible (1 gene @ cen)

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Unlinked Genes in Tetrads

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Linked Genes in Tetrads• Also three tetrad types seen

• parental ditypes: no X-overs (2 str doubles)• non-parental ditypes: 4 str double X-overs• tetratypes more complicated

• single X-overs• 3 strand double X-overs

• Formula for Map distance:• [(1/2 TT’s + 3 NPD’s)/total asci] * 100• applies only to unordered tetrads

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Linkage and Tetrads

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Ordered Tetrads• Neurospora Tetrads: two kinds• First Division Segregation (FDS)

• occurs in absence of recombination• two versions (rotationally equivalent)

• Second Division Segregation (SDS)• occurs with gene-centromere X-overs• four versions (rotationally equivalent)

• Gene-Centromere distance• (1/2 SDS)/total asci * 100• applies only to ordered tetrads

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Ordered Tetrads

Recombination: Holliday Model

Homologous recombination:• single-strand break in homologues pairing

of broken strands occurs • branch migration: single strands pair with

alternate homologue• nicked strands exchange places and gaps

are sealed to form recombinant by Holliday junction-resolving enzyme