1. Next-Generation Sequencing Eric Jorgenson Epidemiology 217 2/28/12

2. Outline Overview of Sequencing Example Next Generation Sequencing Study: Whole Genome, Exome, Families (IBD), Cancer PTC Taste Sensitivity

3. http://www.bloomberg.com/video/84364498/ http://www.bloomberg.com/video/84364540/ http://www.bloomberg.com/video/86406762/ http://www.bloomberg.com/news/2012-01-17/search-genome-as- tennis-thrice-weekly-no-barrier-to-decoded-dna.html http://www.bloomberg.com/news/2012-02-15/harvard-mapping-my- dna-turns-scary-as-threatening-gene-emerges.html Links to videos and articles

4. Huntington’s Disease Testing Almqvist AJHG 1999

5. Number of Genetic Markers for Genetic Studies Genome-wide Linkage Studies 300-400 Microsatellite Markers Genome-wide Association Studies 100,000-2,500,000 SNPs Exome Sequencing Studies 30,000,000 Basepairs Gene-based Studies 22,000 Genes Whole Genome Sequencing Studies 3,200,000,000 Basepairs

6. Variant detection through next generation sequencing Meyerson et al. NRG 2010

7. ANNOVAR: Using Annotation to Narrow the Search Space openbioinformatics.org/annovar

8. Outline Overview of Sequencing Example Next Generation Sequencing Study: Whole Genome, Exome, Families (IBD), Cancer PTC Taste Sensitivity

9. Sequencing of a Single Individual with Family Data Lupski et al. NEJM 2010

10. CMT Subtypes: Many Genes

11. Phenotypes in Unsequenced Family Members

12. SNP Distribution in Proband

13. The First 8 Human Genomes

14. Nonsynonymous SNPs in Known Disease Genes

15. Family Pedigree

16. Putative Causal Variant at a Conserved Amino Acid

17. Exome Sequencing Identifies a Tibetan Adaptation Yi et al. Science 2010

18. Family Sequencing for Rare Diseases Roach et al. Science 2010

19. Sequence Data Improves Identity By Descent Resolution Su and Jorgenson under review

20. Cancer: Tumor vs. Normal Lee et al. Nature 2010

21. Molenaar et al. Nature 2012 Nonsynonymous Somatic Mutations in Neuroblastoma

22. Molenaar et al. Nature 2012 Mutation count associated with age, stage, and survival

23. Outline Overview of Sequencing Example Next Generation Sequencing Study: Whole Genome, Exome, Families (IBD), Cancer PTC Taste Sensitivity

24. Distribution of PTC Phenotype PTC Score Number of Subjects

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

26. 3 SNPs Form 3 Haplotypes PAVPAV AVIAVI AAVAAV Taster Non-taster Rare

27. PTC Phenotype by TAS2R38 Diplotype PTC Score Number of Subjects

28. Outliers After Adjusting for TAS2R38 Diplotype PTC Score Number of Subjects

29. Unusual PTC Phenotypes (AVI Homozygotes in Green) 11 8 108993912 9

30. Unusual PTC Phenotypes (AVI Homozygotes in Green) 14109114102 11

31. 10 Genomes, 5 Hard Drives

32. Summary of Variation Utah Sample 1Sample 2Sample 3Sample 4Sample 5 GenderFemale MaleFemale Total Sequence (Gb)214220218243219 Percent fully called0.950.96 0.970.96 Coverage (X fold)5355536354 SNPs3,270,9203,269,4873,278,5573,355,2663,341,154 Insertions184,763190,633197,830210,805206,120 Deletions195,419200,495208,031221,532216,578 Synonymous SNPs9,6669,5479,80810,0049,981 Missense SNPs9,2539,1359,3509,4869,581 Nonsense SNPs9097828892 Frameshift Insertions10310297112127 Frameshift Deletions9910191108116 Novel SNPs0.04 Novel Insertions0.18 0.190.200.19 Novel Deltions0.22 0.23

33. Quality Control: 99.8% Concordance Sample 1 Genotyping SequencingHomozygous Reference HeterozygousHomozygous Variant Homozygous Reference 479,773429422 Heterozygous 426234,156293 Homozygous Variant 65168172,479

34. Variant Distribution in Utah

36. Using Relatedness 11 8 108993912 9

37. Identity By Descent

39. Nonsynonymous Variants

40. How can whole genome sequence influence treatment? Identify Genes with Protein Altering Mutations Determine Variation in Specific Genes

41. Genes with Protein Altering Variants

42. ABO Blood Group

43. Determination of ABO Type

44. Appendix: Study Design Considerations in Sequencing Extreme Sampling Families

45. Sampling from the Extremes of a Quantitative Distribution Su and Jorgenson under revision

46. Relative Power of Sampling from Various Phenotype Deciles Su and Jorgenson under revision

47. Roach et al. Science 2010 Families can reduce error rates

48. Roach et al. Science 2010 Families can reduce error rates