1. Sean Davis, M.D., Ph.D. Genetics Branch, Center for Cancer Research National Cancer Institute National Institutes of Health Genomics for Pediatrics An Overview of Technologies for Observing the Human Genome

4. The Human Genome Project

5. The Central Dogma

6. phenotype Gene Copy Number Sequence Variation Chromatin Structure and Function Gene Expression Transcriptional Regulation DNA Methylation Patient and Population Characteristics

8. Comparative genomic hybridization

10. Normal Karyotype Tumor Karyotype

12. Integral to life as we know it

13. Can be leveraged to build systems for interrogating biologic processes

14. ~30,000 genes 1 2 3 4 5

16. Gene Expression Microarrays Golub et al., Science 286:531-537. (1999). Spellman et al., Molecular Biology of the Cell 9, 3273-3297. (1999).

18. Breast cancer precursor or benign lesion?

19. Histologic grading used to help define treatment

20. Low grade and high grade clear-cut, but intermediate grade problematic

22. RNA hybridized to DNA, multiple probes per gene

23. RNA hybridized to DNA, one or more probes per exon

26. Useful for determining relative copy number of DNA across the entire genome

27. Not useful (directly) for determining genome structure

28. Array Comparative Genomic Hybridization (aCGH) Tumor DNA Normal DNA Hybridize (Tumor Suppressors) (Oncogenes) Normal Copy Number DNA Loss Amplification

29. Tumor Chromosome Normal Chromosome

30. A Genome View of Copy Number

31. Frequency of Copy Number Changes Summary of copy number changes from 46 breast cancer cell lines

33. Typically requires mutation screening, FISH, and some small deletions may be missed

34. Use array comparative genomic hybridization to get very high-resolution view of the region

37. Comparative Genomic Hybridization (CGH)

39. Again, DNA hybridized to DNA, but designed to detect the differences in hybridization due to a SNP

40. Can be used for measuring copy number, finding stretches of uniparental disomy, risk alleles for certain conditions, and in linkage and association studies

42. Comparative Genomic Hybridization (CGH)

43. Single Nucleotide Polymorphisms

44. DNA methylation

45. MicroRNA expression

46. “Tiling array” applications

47. Others....

48. Tiling Array Technology

49. Simultaneous Gene Expression and Copy Number on Tiling Arrays Annotated Genes Expression Copy Number, Sample 1 Copy Number, Sample 2 Simultaneous measurement of copy number in two samples and gene expression in one sample overlayed on map of genes in the region

50. Simultaneous Gene Expression and Copy Number on Tiling Arrays Annotated Genes Expression Copy Number, Sample 1 Copy Number, Sample 2 Increased expression in the small amplicon does not include all genes, giving clues as to the biologically important genes in the region

51. Simultaneous Gene Expression and Copy Number on Tiling Arrays Annotated Genes Expression Copy Number, Sample 1 Copy Number, Sample 2 Spikes of expression at exons of ERBB2

52. Simultaneous Expression and Copy Number Copy Number Expression Copy Number Evolutionary Conservation Expression Copy Number Opposite Strand Expression Evolutionary Conservation Expression Copy Number

54. Why did the chicken cross the road? Darwin1: It was the logical next step after coming down from the trees. Darwin2: The fittest chickens cross the road.

59. Next-generation Sequencing

63. These “accessible” regions have been associated with open chromatin

64. Regions of open chromatin are necessary for transcriptional and regulatory machinery to have access to gene neighborhoods and facilitate transcription

67. Transcription start sites

68. Early replicating regions

69. Transcription factor binding sites (experimentally determined by ChIP/chip, etc.) Identification and analysis of functional elements in 1% of the human genome by the ENCODE pilot project. The ENCODE Consortium. Nature , 2007.

70. DNAse-Seq Method Crawford, G.E., Davis, S., Scacheri, P.C., Renaud, G., Halawi, M.J., Erdos, M.R., Green, R., Meltzer, P.S., Wolfsberg, T.G., and Collins, F.S. Nat Methods , 2006

75. DNAse is known to cut preferentially in the minor groove, which is exposed every 10.4 bases when wrapped around a nucleosome

76. A nucleosome is wrapped by 147 base pairs when complexed with DNA

77. Implication: Nucleosomes are positioned in a highly organized, precise manner Nucleosome Positioning

79. Phenotype Gene Copy Number Sequence Variation Chromatin Modification Gene Expression Transcriptional Regulation DNA Methylation

81. Databases of gene and protein function and interactions

83. These technologies will see increasing use in clinical medicine, but such use must be tempered by sound clinical hypotheses and testing

84. The data generated by many of these technologies necessitates improved clinical, translational, and research information systems

87. Designing studies that evaluate the operating characteristics of new testing methods is costly and requires the appropriate patient populations

88. There are many ethical concerns associated with the enormous amounts of personal information that might be gleaned from genomic technologies applied in the clinical setting

89. The Biggest Challenge? How do we integrate all the disparate pieces of information, collected longitudinally and by many sources, to improve the health of the individual?

91. Sven Bilke

92. Yuan Jiang

96. Les Biesecker

97. One day the zoo-keeper noticed that the orangutan was reading two books - the Bible and Darwin's The Origin of Species. In surprise he asked the ape, "Why are you reading both those books"? "Well," said the orangutan, "I just wanted to know if I was my brother's keeper or my keeper's brother." [email_address]