1. Eric Jorgenson Epidemiology 217 2/21/12
Linkage Analysis
Eric Jorgenson
Epidemiology 217
2/21/12

2. Worldwide Distribution of Human Earwax SNP rs17822931
Yoshiura et al., Nature Genetics 2006

3. Geographic Distribution of PTC Phenotype
High
Low
Wooding Genetics 2006, adapted from Cavalli-Sforza 1994

4. Bimodal Distribution of PTC

5. Your Phenotypes and Genotypes
Taste SNP
Ear wax SNP
Sample Name
taster
ear wax rs rs rs
BU10 Y D CT AG TT
BU12 N
None
BU14 D? AA
BU15 W CC
BU17
BU19 GG
BU20
BU21
mild
BU22
BU23
BU24 W?
BU25
BU26
BU27
BU28
BU29
Y mild
BU30
From Joe Wiemels

6. Types of Genetic Studies
Family Studies
Compare trait values across family members
Linkage Analysis
Compare trait values with inheritance patterns
Association
Compare trait values against genetic variants

7. Family Studies Familial Relationships Phenotype information
Twins
Siblings
Parents/offspring
Phenotype information
Affected/Unaffected (Prostate Cancer)
Quantitative measure (Blood Pressure)
No Genotype information required

8. Why do Family Studies? Is the trait genetic?
What is the mode of transmission?
Dominant
Recessive
Additive
Polygenic (Multiple genes involved)

9. Mutation and Meiosis

10. Recessive trait

11. First PTC Family Study L. H. Snyder Science 1931 Both: 15% One: 32%
Neither: 100%
L. H. Snyder Science 1931

12. Linkage Analysis Narrow down position of disease gene
No biological knowledge needed
Genetic markers (not disease gene)
Recombination

13. Recombination a A a a b B b b A a a a A a a a b b b b B b B b
2 markers-A and B
Know the orientation A a a a A a a a b b b b B b B b

14. Recombination a A a a b B b b R NR NR R A a a a A a a a b b b b B b B

15. Independent Assortmentb B b b
25%
25%
25%
25% A a a a A a a a b b b b B b B b

16. No recombination a A a a b B b b 0% 50% 50% 0% A a a a A a a a b b b b

17. Recombination Fraction61
420
442 77 A a a a A a a a b b b b B b B b

18. Recombination Fraction
Recombination Fraction q =
Recombinants / Total =
/ = 138 / 1000
= 13.8% 61
420
442 77 A a a a A a a a b b b b B b B b

19. Linkage Linkage Analysis Recombination fraction q < 50%
Two traits: PTC and KELL blood group
Two genetic markers
One trait and one genetic marker
Linkage Analysis

20. Human Linkage Analysis
RFLP Markers for Linkage (1980)
Huntington’s Disease Linkage (1983)
Cystic Fibrosis Linkage (1985)
Cystic Fibrosis Gene (1989)
Huntington’s Disease Gene (1993)

21. Genomewide Linkage Analysis
Genetic Markers
q = 10% on average
Genes

22. Linkage Analysis LOD score based on recombination
LOD (q) = log (q)R (1 - q)NR
____________________
(q = 1/2) R + NR

23. Dominant Trait
D d
d d
D d
D d
d d

24. Linkage Analysis R NR NR

25. LOD score LOD (q) = log (q)1 (1 - q)2 q ____________________
0.01
-1.11
0.05
-0.44
0.1
-0.19
0.2
0.3
0.07
0.4
0.06
0.5
0.00

26. IBD Identity by descent Allele Sharing methods
Often used for affected sib pairs

27. Identity By Descent a A a A
25%
25%
25%
25% A A a A A a a a

28. Identity By Descent a A A A a A A a a a Parent 1 Alleles shared IBD A A a A A a a a

29. Identity By Descent a A A A a A A a a a Parent 1 1 1 1 1 2 0% 1 100%
Alleles IBD
Frequency 2 0% 1
100% a A A A a A A a a a

30. Identity By Descent A A A A a A A a a a Sibling 1 Alleles shared IBD A A a A A a a a

31. Identity By Descent A A A A a A A a a a Sibling 1 2 1 1 0 2 25% 1 50%
Alleles IBD
Frequency 2
25% 1
50% A A A A a A A a a a

32. Identity By Descent IBD can be used for linkage analysis
Expect 50% alleles shared between siblings
Look for IBD > 50% for concordant pairs
Look for IBD < 50% for discordant pairs

33. PTC Linkage Analysis Utah Genetic Reference Project 27 large families
269 subjects

34. PTC Linkage Analysis

35. Human Chromosomes
23 pairs of chromosomes
TAS2R38

36. Fine Mapping
Linkage markers
Genes
Kim et al. Science 2003

37. Linkage Disequilibrium

38. Linkage Disequilibrium

39. Linkage Disequilibrium

40. Linkage Disequilibrium

41. Linkage Disequilibrium
Time

42. Linkage Disequilibrium Mapping
Genetic Markers
Genes

43. PTC Linkage Disequilibrium Mapping
Kim et al. Science 2003

44. TAS2R38 Receptor Structure
Kim et al. J Dent Res 2004

45. 3 SNPs in the TAS2R38 Gene P A V P A I P V V P V I A A V A A I A V V
Haplotype definition
Each individual has two haplotypesdiplotype
Haploytpeallele
diplotypegenotype
A A I
A V V
A V I

46. TAS2R38 Diplotype and PTC Score
2 haplotypes3 diplotypes
AVI = 2
PAV = 10
Heterozygote = 9
A multivariate analysis explains most of the variance with a p-value of < 10-33
Kim et al. Science 2003

47. Confirm Mode of Inheritance
Both: 15%
One: 32%
Neither: 100%
L. H. Snyder Science 1931 47

48. Explain Linkage Signal

49. Geographic Distribution of PTC Phenotype
Wooding Genetics 2006, adapted from Cavalli-Sforza 1994

50. Geographic Distribution of PTC Haplotypes
Kim et al. Science 2003

51. Diplotype and PTC Score
You may notice a 4th diplotype  due to a 3rd, rare haplotype.
Kim et al. Science 2003

52. 3 SNPs form 3 Haplotypes P A V A V I A A V Taster Non-taster Rare
3rd haplotype is the result of recombination.
A of non-taster
AV of taster
Allows us to compare the effect of the 1st SNP vs. the 2nd and 3rd.
Rare-not in all combinations
Rare
A A V

53. Comparing Diplotypes Mean for taster is 9
Mean for rare haplotype is 7.
Difference is P vs. A
Mean for non-taster is 2
Difference is AV vs. VI
Both have an effect, but 2nd and 3rd have a greater effect.

54. Predicted Effect of the 3 SNPs
3 a.a. substitution matrices
Sequence alignment
Scale
Chemical change
1st and 2nd most severe.

55. TAS2R38 Haplotype Function

56. PTC Diplotype and Taste
Sandell and Breslin Current Biology 2006

57. Next Week
Next Generation Sequencing

58. Appendix: Phase Unknown Linkage

59. Phase Unknown ? ? ? ? ? ? ? ? ? ? D d d d 1 2 3 3 D d D d d d 1 3 2 3
? ?
? ?
Phase Unknown
? ?
? ?
? ?
D d
d d
D d
D d
d d

60. Phase Unknown ? ? ?

61. What if we don’t know phase?
We calculate the LOD score for each phase
Divide by 2

62. Phase Unknown + = -0.02 for q = 0.44 LOD (q) = ½ log (q)1 (1 - q)2
____________________
(q = 1/2) 1 + 2 +
½ log (q)2 (1 – q)1
(q = 1/2) 2 + 1
= for q = 0.44