1. 1 of 25 Sequence Variation in Ensembl

2. 2 of 25 Outline SNPs SNPs in Ensembl Linkage disequilibrium SNPs in BioMart DAS sources

3. 3 of 25 Single nucleotide polymorphisms (SNPs) Two human genomes differ by ~0.1% Polymorphism: a DNA variation in which each possible sequence is present in at least 1% of people Most polymorphisms (~90%) take the forms of SNPs: variations that involve just one nucleotide ~1 out of every 300 bases in the human genome ~10 million in the human genome

4. 4 of 25 Functional Consequences SNPs in coding area that alter aa sequence SNPs in coding areas that don’t alter aa sequence SNPs in promoter or regulatory regions SNPs in other regions Cause of most monogenic disorders, e.g: Hemochromatosis (HFE) Cystic fibrosis (CFTR) Hemophilia (F8) May affect splicing May affect the level, location or timing of gene expression No direct known impact on phenotype, useful as markers

5. 5 of 25 Practical Applications Disease diagnosis Association studies Pharmacogenomics Forensic testing Population genetics and evolutionary studies Marker-assisted selection

6. 6 of 25 Practical Applications

7. 7 of 25 SNPs in Ensembl Most SNPs imported from dbSNP (rs……): Imported data: alleles, flanking sequences, frequencies, …. Calculated data: position, synonymous status, peptide shift, …. For human also: HGVbase TSC Affy GeneChip 100K and 500K Mapping Array Affy Genome-Wide SNP array 6.0 Ensembl-called SNPs (from Celera reads and Jim Watson’s and Craig Venter’s genomes) For mouse, rat, dog and chicken also: Sanger- and Ensembl-called SNPs (other strains / breeds)

8. 8 of 25 dbSNP Central repository for simple genetic polymorphisms: single-base nucleotide substitutions small-scale multi-base deletions or insertions retroposable element insertions and microsatellite repeat variations http://www.ncbi.nlm.nih.gov/SNP/index.html For human (dbSNP build 128): 34,434,159 submissions (ss#’s) 11,883,685 RefSNP clusters (rs#’s) 6,262,709 validated 737,679 with frequency

9. 9 of 25 SNPs in Ensembl - Types Non-synonymousIn coding sequence, resulting in an aa change Synonymous In coding sequence, not resulting in an aa change FrameshiftIn coding sequence, resulting in a frameshift Stop lostIn coding sequence, resulting in the loss of a stop codon Stop gainedIn coding sequence, resulting in the gain of a stop codon Essential splice site In the first 2 or the last 2 basepairs of an intron Splice site1-3 bps into an exon or 3-8 bps into an intron UpstreamWithin 5 kb upstream of the 5'-end of a transcript Regulatory regionIn regulatory region annotated by Ensembl 5' UTRIn 5' UTR IntronicIn intron 3' UTRIn 3' UTR DownstreamWithin 5 kb downstream of the 3'-end of a transcript IntergenicMore than 5 kb away from a transcript

10. 10 of 25 SNPs in Ensembl - Species Human Chimp Mouse Rat Dog Cow Platypus Chicken Zebrafish Tetraodon Mosquito

11. 11 of 25 Caveat For human, mouse and rat Ensembl defines all SNP alleles respective to the + strand of the genome assembly! (to be able to merge dbSNP data with Sanger resequencing data) Exceptions: Those cases where SNPs are shown as part of a sequence

12. 12 of 25 A missense SNP, C1858T, in PTPN22 (Tyrosine-protein phosphatase non-receptor type 22) has been identified as a genetic risk factor for rheumatoid arthritis. This SNP is also referred to as R620W. 1.Find the SNPView page for this SNP. 2.Why are the alleles on this page given as A/G? 3.What is the minor allele of this SNP in Caucasians? 5 MINUTE EXERCISE

13. 13 of 25 SNPs in Ensembl GeneSNPView (1) SNP alleles Transcript InterPro domains

14. 14 of 25 SNPs in Ensembl GeneSNPView (2)

15. 15 of 25 SNPs in Ensembl TranscriptSNPView (1) Shows SNP alleles in different: Individuals (human): Celera HuAA, HuCC, HuDD and HuFF, Craig Venter, Jim Watson Strains (mouse, rat) Breeds (chicken, dog)

16. 16 of 25 SNPs in Ensembl TranscriptSNPView (2) Resequencing coverage Alleles in different individuals SNP alleles Different individuals

17. 17 of 25 SNPs in Ensembl TranscriptSNPView (3)

18. 18 of 25 1.Find the TranscriptSNPView page for human PTPN22. 2.Do all individuals (HuAA, HuCC, HuDD, HuFF, Venter and Watson) have resequence coverage at the position of the C1858T (R620W) SNP? 3.Has any of the individuals a higher risk to get rheumatoid arthritis based on its genotype at this position? 4.Is there an individual that is heterozygote at this position? 5 MINUTE EXERCISE

19. 19 of 25 Haplotypes and Linkage Disequilibrium A haplotype is a set of SNPs on a single chromatid that are statistically associated Linkage disequilibrium describes a situation in which some combinations of SNP alleles occur more or less frequently in a population than would be expected from a random formation of haplotypes from alleles based on their frequencies

20. 20 of 25 Measures of LD D = P(AB) – P(A)P(B) D ranges from – 0.25 to + 0.25 D = 0 indicates linkage equilibrium dependent on allele frequencies, therefore of little use D’ = D / maximum possible value D’ = 1 indicates perfect LD estimates of D’ strongly inflated in small samples r 2 = D 2 / P(A)P(B)P(a)P(b) r 2 = 1 indicates perfect LD measure of choice

21. 21 of 25 Linkage Disequilibrium LDView It is also possible to export SNP information for upload into the HaploView software tool

22. 22 of 25 Linkage Disequilibrium LDTableView

23. 23 of 25 Retrieve all non-synonymous SNPs for the human CFTR gene using BioMart and export their id, genomic position, alleles and peptide shift (hint: which dataset should you start with?). 5 MINUTE EXERCISE

24. 24 of 25 DAS Sources For human, data from the following DAS Sources can be visualised on ContigView: DGV and DGV loci: Structural variations from the Database of Genomic Variations (CNVs, InDels, inversions etc.) RedonCNV regions and RedonCNV loci: Copy number variations from Redon et al. paper SegDup Washu: Segmental Duplications, University of Washington

25. 25 of 25 Q & A Q U E S T I O N S A N S W E R S