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From QTL to QTG: Are we getting closer? Sagiv Shifman and Ariel Darvasi

From QTL to QTG: Are we getting closer? Sagiv Shifman and Ariel Darvasi. The Hebrew University of Jerusalem. Presentation Outline. Overview of fine QTL mapping strategies Inbred strain sequence/haplotype information for fine QTL mapping

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From QTL to QTG: Are we getting closer? Sagiv Shifman and Ariel Darvasi

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  1. From QTL to QTG: Are we getting closer?Sagiv Shifman and Ariel Darvasi The Hebrew University of Jerusalem

  2. Presentation Outline • Overview of fine QTL mapping strategies • Inbred strain sequence/haplotype information for fine QTL mapping • Yin-Yang crosses: A framework for applying inbred strain sequence/haplotype information to fine map QTLs • Simulation analysis using Celera’s sequence information of 4 inbred strains (C57, A/J, DBA, 129)

  3. The Difficulties • Genetics • Genotype/phenotype correlation • Recombination • Functional • The “end-game” (knockout, transgenic, mutation analysis, RNAi, etc.)

  4. Fine Mapping Strategies Genomewide-based strategies: • Large scale BC, F2, half sibs, etc. • Advanced Intercross Lines (AIL) • The heterogeneous stock (HS) Locus-based strategies: • Selective phenotyping • Recombinant progeny testing • Interval specific congenic strains (ISCS) • Recombinant inbred segregation test (RIST)

  5. Sample size required

  6. Ft F1 F2 F3 P Advanced intercross lines (AIL) Semi-random intercrossing CI = CIF2 / (t/2)

  7. AIL - experimental results HDL – QTL Wang et al. 2003

  8. The heterogeneous stock (HS) Eight-way cross of: C57BL/6, BALB/C, RIII, AKR, DBA/2, I, A/J C3H Established 30 years ago (~60 generations) by McClearn et al.

  9. HS - Experimental results Open field activity Talbot et al. 1999 0 0.2 0.4 0.6 0.8 1.0 1.2 1.4 cM

  10. Locus-based strategies:Selective phenotyping (SPh) • Theoretical basis: Only recombinants increase mapping accuracy for a detected QTL. • Procedure: Only individuals recombinant at a QTL-containing interval are subsequently phenotyped.

  11. SPh - Experimental results Lesions density Paigen et al. BC SPh-BC

  12. QTL Recombinant progeny testing Males, recombinant at an interval of interest, are progeny tested to check which QTL allele was retained.

  13. QTL Interval specific congenic strains (ISCS) ISCS are produced by a series of backcrosses and intercrosses

  14. Recombinant inbred segregation test (RIST) P1 RI P2 QTL x x F1,1 F1,2 F2,1 F2,2

  15. RIST - Experimental results D2MIT64 Obesity QTL B. Taylor A. Darvasi D2MIT200 C57L AKXL-16 AKR F21 F22 P=0.02 P=0.41

  16. In silico mapping of complex disease-related traits in mice Grupe et al. 2001 Comment: Chesler et al. 2001 Darvasi 2001

  17. Genome-wide single-nucleotide polymorphism analysis defines haplotype patterns in mouse Wiltshire et al. 2003 Unexpected complexity in the haplotypes of commonly used inbred strains of laboratory mice Yalcin et al. 2004 Multiple Cross and Inbred Strain Haplotype Mapping of Complex-Trait Candidate Genes Park et al. 2003

  18. Yin-Yang Crosses : A framework for Multiple Cross Inbred Strain Haplotype Mapping QTL detection with two inbred lines (P1 and P2) ↓ Estimating QTL map location ↓ Sequence based fine mapping ↓ Selecting an optimal new inbred strain (Pi) ↓ Yin-Yang crosses

  19. QTL mapped in a AxB cross A B

  20. Sequence/Haplotype Information A B

  21. Yin-Yang Crosses A C B  

  22. Simulation Analysis • Initial cross between A/JxC57 • 22,814 SNPs, each at its turn, simulated as the QTN • The strain closest to being half similar to A/J and half to C57 was selected first for Yin-Yang crosses (DBA or 129) • The forth strain was subsequently introduced • At each stage the number of the remaining SNPs that can be the QTN, and the size of the QTL containing interval were estimated

  23. 2 strains 3 strains 4 strains Mapping Resolution: Chromosome 16

  24. Low resolution region High resolution region

  25. Mean Reduction in Interval Length

  26. Distribution of Mapping Resolution with 4 Inbred Strains Proportion Interval length

  27. Conclusions • Using inbred strain sequence/haplotype information combined with multiple crosses (Yin-Yang crosses) can efficiently aid QTL fine mapping • With 4 strains most regions exhibit modest mapping resolution. • The significant haplotype variation observed in 4 strains may suggest that with a large number of strains in hand significant resolution can be achieved. • With sequence information in hand, resolution maps and algorithms can be established to guide the researchers to the optimal strain selection strategy and provide the resolution expected for their region of interest. • A large number of sequenced strains will allow the actual selection of a very small number of strains for additional crosses.

  28. So, Do We Need the 1K RI Set? YES Until we have the 1K RI set any QTL mapping strategy is like riding a bike: it’s fun and we can get very far – Once we get something with an engine though, we will feel the difference! … we are still missing an engine for the “end-game” – Genetics is not everything

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