Genomic Analyses: QTLs, etc.


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Genomic Analyses: QTLs, etc.

  1. 1. Direct and correlated responses to selection. 1. EGR <ul><li>Primary correlated response to selection for fast growth to 42 days of age is still EGR 14 . </li></ul><ul><li>Correlated response to selection for EGR 14 is still EGR 14/42 . </li></ul>
  2. 2. Correlated responses to selection 2. Body Weight <ul><li>Direct response to selection for fast EGR 14 still makes a large, lean bird at 42 days of age. </li></ul><ul><ul><li>Divergence between 14H and 42H birds at 42 days of age is largely fat! </li></ul></ul><ul><li>14L chicks largest at hatch, smallest at 14 days of age, but getting bigger by 42 days of age. </li></ul>
  3. 3. 14L and 14H Lines at 14 and 42 Days (S 14 ) 14L 14H 38 g 319 g 172 g 861 g
  4. 4. Life Cycle of Selection Lines
  5. 5. QTL Analysis F 2 segregating generations Substantial heterosis or dominance for early growth
  6. 6. QTL Analysis F 2 segregating generations Substantial additive variation for late growth
  7. 7. Types of Molecular Markers <ul><li>Restriction Fragment Length Polymorphisms </li></ul><ul><ul><li>RFLP </li></ul></ul><ul><ul><li>Restriction enzyme in conjunction with probe. </li></ul></ul><ul><li>Randomly Amplified Polymorphic DNA </li></ul><ul><ul><li>RAPD </li></ul></ul><ul><ul><li>PCR using random primer sequences </li></ul></ul><ul><li>Microsatellite DNA </li></ul><ul><ul><li>Short sequences of tandemly repeated DNA </li></ul></ul><ul><ul><li>PCR with resultant different length cDNA </li></ul></ul>
  8. 8. RFLP A a AA Aa aa Resulting gel
  9. 9. RAPD A a AA Aa aa Resulting gel No PCR product
  10. 10. Microsatellite A a AA Aa aa Resulting gel GCC GCC GCC GCC GCC GCC GCC GCC
  11. 11. Molecular marker loci properties Rafalski and Tingey, 1993
  12. 12. Molecular mechanism for crossing-over (Robin Holliday, 1960s): <ul><li>Homologous chromosomes “recognize” and align. </li></ul><ul><li>Single strands of each DNA (one on each chromosome) break and anneal to the opposite chromosome forming Holliday intermediate. </li></ul><ul><ul><li>As chromosome ends pull apart, branch point migrations occur to create a 4-arm intermediate structure. </li></ul></ul><ul><ul><li>4-arm intermediate is cut by by endonucleases in one of 2 planes. </li></ul></ul><ul><ul><li>DNA seals the gaps. </li></ul></ul><ul><li>Model predicts that physical exchange between two gene loci at the ends of the chromosomes should occur about 50% of the time. One pattern (intermediate cut in one plane) yields the parental arrangement, the other (cut in the other plane) is recombinant. </li></ul>
  13. 13. Holliday model of chromosome recombination
  14. 14. Experimental designs for QTL detection <ul><li>Inbred lines </li></ul><ul><ul><li>Back cross </li></ul></ul><ul><ul><li>Test cross </li></ul></ul><ul><ul><li>F 2 </li></ul></ul><ul><ul><li>Recombinant Inbred Lines </li></ul></ul><ul><li>Segregating populations </li></ul><ul><ul><li>Full-sibs </li></ul></ul><ul><ul><li>Half-sibs </li></ul></ul><ul><ul><li>Grand-daughter </li></ul></ul><ul><ul><li>Animal model </li></ul></ul>
  15. 15. Backcross design Parental strains X X F 1 strains Backcross Progeny Non-recombinants Recombinants Frequency = 1 - r Frequency = r m q m q M Q M Q m q M Q M Q M Q m q M Q M Q M Q m Q M Q M q M Q
  16. 16. Bulked segregant analysis 14L 14H F 1 F 2 Bulk 1 Mostly qq ; enriched for m , depleted for M Bulk 2 Mostly QQ ; enriched for M , depleted for m m q m q M Q M Q M Q m q
  17. 17. Recombinant inbred lines (by selfing)
  18. 18. Recombinant inbred lines (by sibling mating)
  19. 19. Advantages of RI lines <ul><li>Each strain is an eternal resource. </li></ul><ul><ul><li>Only need to genotype once. </li></ul></ul><ul><ul><li>Reduce individual variation by phenotyping multiple individuals from each strain. </li></ul></ul><ul><ul><li>Study multiple phenotypes on the same genotype. </li></ul></ul><ul><li>Greater mapping precision. </li></ul><ul><ul><li>More dense breakpoints on the RI chromosomes. </li></ul></ul>
  20. 20. Grand-daughter design X Grandsire & Grandam(s) X X Sons x random dams (genotyped) Son Son Grand-daughters M 1 Q 1 M 2 Q 2 M x Q x M x Q x M x Q x M x Q x M 1 Q 1 M x Q x M 2 Q 2 M x Q x M x Q x M x Q x M 1 Q 1 M x Q x M x Q x M x Q x M x Q x M x Q x M 2 Q 2 M x Q x
  21. 21. Determining the genome sequence Sequence assembly Partial digest Library of bacterial artificial chromosomes (BACs) with genomic fragments Alignment of BAC clones > contigs and anchoring On the molecular gene map Sequencing of overlapping BAC clone fragments “ Tiling path” > selection of BAC clones to be sequenced Annotation of genome sequence > prediction of genes
  22. 22. Genomes of human and mouse compared human: 22 chromosomes + X or Y mouse: 19 chromosomes + X or Y homologous segments are color-coded (total >300) human chromosomes correspond to segments of different mouse chromosomes e.g.. Segments of chr. 10, 11, 15, 16 and 19 correspond to mouse chromosome 7
  23. 23. Historical perspective: from genetic map to genome sequence
  24. 24. Chicken Chromosome 4 25 cM LEI122 * LEI76 LEI81 ** 270 cM SPP1 Murine SPP1 56.0 cM Chicken Chromosome 4 Murine Chromosome 5 (Late Growth QTL) (Cluster 1) (Cluster 2) Cluster 1 - Protein kinase II, glucokinase, VDR, galactosyltransferase, Ubiquitin C and FGF5 Cluster 2 - Acads, Phkg and Asl Cluster 3 - ZP3, Zonadhesin , Epo and Actin (Cluster 3)
  25. 25. Different markers corresonding to the same genes in prior slide.