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Physical and transcript mapping

Physical and transcript mapping. Physical mapping Transcript identification. Physical and transcript mapping. Somatic Cell Hybrids Monochromosomal Polychromosomal Radiation hybrids. Physical and transcript mapping. Somatic Cell Hybrids

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Physical and transcript mapping

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  1. Physical and transcript mapping • Physical mapping • Transcript identification

  2. Physical and transcript mapping • Somatic Cell Hybrids • Monochromosomal • Polychromosomal • Radiation hybrids

  3. Physical and transcript mapping • Somatic Cell Hybrids • Subchromosomal assignment by PCR of hybrids with different chromosomal content.

  4. Physical and transcript mapping • Radiation hybrid maps • Random breakpoints. • Ordering of markers.

  5. Physical and transcript mapping • Fluorescence in situ hybridization • YAC hybridization. • Translocation breakpoint 3q26 and 17q23.

  6. Physical and transcript mapping • Fluorescence in situ hybridization • Chromosome painting. • Chromosome 8 paint.

  7. Physical and transcript mapping • Flow sorting chromosomes • Fluorescent dye labeled chromosomes. • Flow karyogram. • Fractionate whole chromosomes Yeast/YACs

  8. Physical and transcript mapping • Pulsed field electrophoresis • Rotating magnetic field. • Large DNA fragments.

  9. Physical and transcript mapping • Clone contig • Overlapping DNA fragments.

  10. Physical and transcript mapping • Chromosome walking • Clone to clone hybridization.

  11. Physical and transcript mapping • Inverse PCR • Clone flanking unique DNA sequences.

  12. Physical and transcript mapping • Bubble PCR • Amplification of uncharacterized sequences flanking known DNA.

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

  14. Physical and transcript mapping • Contig assembly • YAC contig assembly by STS content mapping.

  15. Physical and transcript mapping • Inter-SINE PCR • Amplify unique sequences between repeats. • Species specific (Alu in primates).

  16. Physical and transcript mapping • Zoo blot • Study highly conserved sequences. • Species specific.

  17. Physical and transcript mapping • Exon trapping

  18. Physical and transcript mapping • cDNA selection using magnetic bead capture

  19. Genetic mapping Genetic markers Recombinants Genetic vs. Physical distances

  20. Genetic mapping Marker Number Features RFLPs >105 diallelic max het 0.5 VNTRs mini >104 many alleles high het VNTRs mic >105 many alleles high het SNPs >106 less informative than VNTRs

  21. Genetic mapping Defining recombinants

  22. Genetic mapping Single vs. Double recombinants

  23. Genetic mapping Relationship between genetic and physical maps for human chromosome 19

  24. 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

  25. 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 (Z>3) no recombinants Curve 2 linkage (Z>3) recombination fraction 0.23 Curve 3 no linkage (Z<-2) for recombination fractions below 0.12 Curve 4 inconclusive

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