GENE MAPPINGGENE MAPPING
By A.Arputha SelvarajBy A.Arputha Selvaraj
Comparative GenomicsComparative Genomics
Gene functions have been conservedGene functions have been conserved
across evolutionacross evolution
Nature solves a problem, it rarely solves itNature solves a problem, it rarely solves it
Evolution & GenesEvolution & Genes
The function of many genes is conserved acrossThe function of many genes is conserved across
immense evolutionary distances.immense evolutionary distances.
The function of a gene is affected by its environment.The function of a gene is affected by its environment.
Many gene products function as part of a cascade orMany gene products function as part of a cascade or
“Nothing in Biology Makes Sense Except in the Light
of Evolution” (Theodosius Dobzhansky)
Map Genes in the Context ofMap Genes in the Context of
Uses of Gene MappingUses of Gene Mapping
Identify genes responsible for diseases.Identify genes responsible for diseases.
Heritable diseasesHeritable diseases
Identify genes responsible for traits.Identify genes responsible for traits.
Plants or AnimalsPlants or Animals
Disease resistanceDisease resistance
Meat or Milk ProductionMeat or Milk Production
Types of MapsTypes of Maps
Nucleotide Sequence MapsNucleotide Sequence Maps
complete or partially sequenced organismscomplete or partially sequenced organisms
Cytogenetic MapsCytogenetic Maps
Breakpoints in diseaseBreakpoints in disease
Direct binding of probes to chromosomeDirect binding of probes to chromosome
Genetic Linkage MapsGenetic Linkage Maps
Physical MapsPhysical Maps
Mapping GenesMapping Genes
Essential element -- MarkersEssential element -- Markers
Differences between two members of aDifferences between two members of a
Typically between 1-400 nucleotides inTypically between 1-400 nucleotides in
Can also be gross chromosomalCan also be gross chromosomal
DNA SequencingDNA Sequencing
Two methods originally developed:Two methods originally developed:
Chemical or Maxim and Gilbert method.Chemical or Maxim and Gilbert method.
Dideoxynucleotide or Sanger method.Dideoxynucleotide or Sanger method.
Both Gilber and Sanger won Nobel prizes forBoth Gilber and Sanger won Nobel prizes for
developing these techniques.developing these techniques.
Dideoxynucleotide method is by far theDideoxynucleotide method is by far the
most common employed today.most common employed today.
Automated SequencingAutomated Sequencing
Each nucleotide is labeled with aEach nucleotide is labeled with a
different fluorescent chromophore.different fluorescent chromophore.
Uses Taq polymerase to incorporateUses Taq polymerase to incorporate
fluorescent dideoxy nucleotide.fluorescent dideoxy nucleotide.
Analysis on a polyacrylamide gel orAnalysis on a polyacrylamide gel or
capillary matrix that is can separatecapillary matrix that is can separate
DNA based upon a 1 nucleotideDNA based upon a 1 nucleotide
30-1000 nucleotides in length can30-1000 nucleotides in length can
theoretically be determined.theoretically be determined.
Nucleotide SequencingNucleotide Sequencing
Complete Genome SequencingComplete Genome Sequencing
Bacterial Artificial Chromosomes (BAC)Bacterial Artificial Chromosomes (BAC)
• Can grow in bacteriaCan grow in bacteria
• Have large inserts 100,000-300,000Have large inserts 100,000-300,000
Collect a large library of BACsCollect a large library of BACs
Restriction Endonuclease (RE) MapRestriction Endonuclease (RE) Map
• RE - cuts DNA at specific sitesRE - cuts DNA at specific sites
• EcoRI cuts at GAATTCEcoRI cuts at GAATTC
Genomic SequencingGenomic Sequencing
Once BAC map completeOnce BAC map complete
Sequence ends of BAC clones.Sequence ends of BAC clones.
With dense BAC map, large regions may beWith dense BAC map, large regions may be
covered by overlapping the sequences.covered by overlapping the sequences.
Individual BACs can be completelyIndividual BACs can be completely
Individual BACs can be hybridized toIndividual BACs can be hybridized to
chromosomes to identify chromosome ofchromosomes to identify chromosome of
Permits Mapping by providing:Permits Mapping by providing:
Linkage groupsLinkage groups
During MeiosisDuring Meiosis
Linkage Groups - markers that tend toLinkage Groups - markers that tend to
remain together.remain together.
Distance - the further apart two markersDistance - the further apart two markers
lie, the more often recombination willlie, the more often recombination will
occur between those markers.occur between those markers.
Markers on the same chromosome can beMarkers on the same chromosome can be
so far apart that they appear in differentso far apart that they appear in different
linkage groups.linkage groups.
Cytogenetic MappingCytogenetic Mapping
Philadelphia ChromosomePhiladelphia Chromosome
Chronic myelogenous leukemia (CML) isChronic myelogenous leukemia (CML) is
characterized by a reciprocal translocationcharacterized by a reciprocal translocation
between chromosomes 9 and 22 thatbetween chromosomes 9 and 22 that
produces the Philadelphia chromosome.produces the Philadelphia chromosome.
Invariably there is disease progression,Invariably there is disease progression,
with loss of the capacity for terminalwith loss of the capacity for terminal
differentiation by the hematopoietic stemdifferentiation by the hematopoietic stem
cell, resulting in an acute leukemia.cell, resulting in an acute leukemia.
Genetic Linkage MapsGenetic Linkage Maps
Polymorphism - an allele present in aPolymorphism - an allele present in a
population that exhibits multiple forms.population that exhibits multiple forms.
Monomorph - Single form in a population.Monomorph - Single form in a population.
Typically result in no effect on survival ofTypically result in no effect on survival of
Types of MarkersTypes of Markers
Single Nucleotide Polymorphisms (SNPs)Single Nucleotide Polymorphisms (SNPs)
Occur approximately every 100-300 bp inOccur approximately every 100-300 bp in
Microsatellites or Tandem RepeatsMicrosatellites or Tandem Repeats
Physical MapsPhysical Maps
Sequence Tagged Sites (STSs)Sequence Tagged Sites (STSs)
Use Polymerase Chain Reaction (PCR).Use Polymerase Chain Reaction (PCR).
Primers based upon:Primers based upon:
• random sequencerandom sequence
• expressed sequence tagexpressed sequence tag
Amplify from library of clones containingAmplify from library of clones containing
large inserts (BAC).large inserts (BAC).
Relate to BAC map.Relate to BAC map.
If more than one on same clone, then closeIf more than one on same clone, then close
Types of questionsTypes of questions
Where does gene X exist within the genome of organism Y?Where does gene X exist within the genome of organism Y?
What are some flanking markers?What are some flanking markers?
Which genes exist on a chromosome, and in what order do theyWhich genes exist on a chromosome, and in what order do they
Show the genes that exist in region R of the chromosome. ShowShow the genes that exist in region R of the chromosome. Show
me the corresponding sequence data for that region.me the corresponding sequence data for that region.
Display the region of a chromosome between points A and B.Display the region of a chromosome between points A and B.
Show both the cytogenetic and sequence map for that region,Show both the cytogenetic and sequence map for that region,
aligned to each other based on markers that have been placedaligned to each other based on markers that have been placed
on both maps.on both maps.
What is the distance between two genes? (Note: scale dependsWhat is the distance between two genes? (Note: scale depends
on the type of map on which those genes have been placed.)on the type of map on which those genes have been placed.)
I know the cytogenetic location of gene X. What is theI know the cytogenetic location of gene X. What is the
corresponding physical location?corresponding physical location?
Genes and MapsGenes and Maps
Map LinkMap Link
OMIM LinkOMIM Link
Syntenic Relationships - tendency ofSyntenic Relationships - tendency of
closely linked genes to remain linkedclosely linked genes to remain linked
during evolution.during evolution.
KEGG pathwaysKEGG pathways
Examine Cell Adhesion MoleculesExamine Cell Adhesion Molecules
-What does JAM3 appear to interact with?-What does JAM3 appear to interact with?
-What information is available if you click on-What information is available if you click on
What motifs are found in JAM2?What motifs are found in JAM2?
Click on Tight JunctionClick on Tight Junction
-What molecules appear to interact with the JAM--What molecules appear to interact with the JAM-
JAM dimer? What is ZO-1JAM dimer? What is ZO-1