Physical and transcript mapping

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Physical and transcript mapping

• Physical mapping
• Transcript identification

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Physical and transcript mapping

• Somatic Cell Hybrids
• Monochromosomal
• Polychromosomal
• Radiation hybrids

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Physical and transcript mapping

• Somatic Cell Hybrids
• Subchromosomal assignment by PCR of hybrids with
  different chromosomal content.

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Physical and transcript mapping

• Radiation hybrid maps
• Random breakpoints.
• Ordering of markers.

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Physical and transcript mapping

• Fluorescence in situ hybridization
• YAC hybridization.
• Translocation breakpoint 3q26 and 17q23.

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Physical and transcript mapping

• Fluorescence in situ hybridization
• Chromosome painting.
• Chromosome 8 paint.

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Physical and transcript mapping

• Flow sorting chromosomes
• Fluorescent dye labeled chromosomes.
• Flow karyogram.
• Fractionate whole chromosomes Yeast/YACs

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Physical and transcript mapping

• Pulsed field electrophoresis
• Rotating magnetic field.
• Large DNA fragments.

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Physical and transcript mapping

• Clone contig
• Overlapping DNA fragments.

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Physical and transcript mapping

• Chromosome walking
• Clone to clone hybridization.

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Physical and transcript mapping

• Inverse PCR
• Clone flanking unique DNA sequences.

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Physical and transcript mapping

• Bubble PCR
• Amplification of uncharacterized sequences
  flanking known DNA.

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Physical and transcript mapping

• PCR based screening
• Amplification of row, column and plate pools for
  clone identification.

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Physical and transcript mapping

• Contig assembly
• YAC contig assembly by STS content mapping.

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Physical and transcript mapping

• Inter-SINE PCR
• Amplify unique sequences between repeats.
• Species specific (Alu in primates).

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Physical and transcript mapping

• Zoo blot
• Study highly conserved sequences.
• Species specific.

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Physical and transcript mapping

• Exon trapping

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Physical and transcript mapping

• cDNA selection using magnetic bead capture

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Genetic mapping

• Genetic markers
• Recombinants
• Genetic vs. Physical distances

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Genetic mapping

• Marker Number Features
• RFLPs gt105 diallelic max het 0.5
• VNTRs mini gt104 many alleles high het
• VNTRs mic gt105 many alleles high het
• SNPs gt106 less informative than VNTRs

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Genetic mapping

• Defining recombinants

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Genetic mapping

• Single vs. Double recombinants

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Genetic mapping

• Relationship between genetic and physical maps
  for human chromosome 19

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Genetic mapping

• Recognizing recombinants
• AD disorder
• III-6 is recombinant (disorder A2 allele)
• III-1 III-5 are non recomb and III-6 recomb or
  III-1 II-5 are recomb and III-6 is non-recomb
• III-7 and III-8 have disorder but the A1 allele
  may not be identical by descent

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Genetic mapping

• LOD score ratio of likelihood linked
  recombination fraction theta)/not linked
  (recombination fraction 0.5)
• Positive in favor of linkage (3)
• Negative scores against linkage (-2)
• Curve 1 linkage (Zgt3) no recombinants
• Curve 2 linkage (Zgt3) recombination fraction 0.23
• Curve 3 no linkage (Zlt-2) for recombination
  fractions below 0.12
• Curve 4 inconclusive

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Genetic mapping

• Multipoint mapping
• Highest peaks are most likely locations
• Odds in favor of a position are measured by the
  degree to which the highest peak overtops its
  rivals
• LOD scores dip near markers that show
  recombinants with the disease

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Genetic mapping

• Double recombinants suggest errors
• Probability of a true double recombinant with
  markers 5cM apart is small (below 0.05 x 0.05
  0.0025)

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Genetic mapping

• Autozygosity mapping
• Mapping for markers that are identical by
  descent
• Congenital deafness
• Affected members all homozygous for AFMa052yb5
  and D2S158
• Gene between AFMb346ye5 and D2S174

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