1. Ensembl Database and Web Browser www.ensembl.org
Erin Pleasance
February 21, 2005
Ensembl Database and Web Browser
Stephen Baird
Apoptosis Research Centre
Children’s Hospital of Eastern Ontario
Lecture/Lab 7.3
(c) 2005 CGDN

2. Focus on vertebrates No fungi/plants
Erin Pleasance
February 21, 2005
Focus on vertebrates
No fungi/plants
Brassica/Arabidopsis genome browser is at
Lecture 7.1
(c) 2005 CGDN

3. What is Ensembl? Joint project of EBI and Sanger
Erin Pleasance
What is Ensembl?
February 21, 2005
Joint project of EBI and Sanger
Automated annotation of eukaryotic genomes
Open source software
Relational database system
Web interface
“The main aim of this campaign is to encourage scientists across the world - in academia, pharmaceutical companies, and the biotechnology and computer industries - to use this free information.”
- Dr. Mike Dexter, Director of the Wellcome Trust
Lecture/Lab 7.3
(c) 2005 CGDN

4. Ensembl components Search tools: Data: Chromosomes SNPs and Haplotypes
Erin Pleasance
Ensembl components
February 21, 2005
Search tools:
Data:
Chromosomes
(FeatureView, KaryoView,
Ctyoview, MapView)
SNPs and Haplotypes
(SNPView, GeneSNPView,
HaploView, LDView)
Sequence
Similarity
(BLAST, SSAHA)
Diseases
(DiseaseView)
Genome
Sequence
(ContigView)
Genes
(GeneView, TransView,
ExonView, GeneSeqView)
Markers
(MarkerView)
Functions
(GOView)
Text
(TextView)
Other
Annotations
Protein
(ProtView, DomainView,
FamilyView
Anything
(BioMart/Martview)
Comparative
Genomics
(ContigView, MultiContigView,
SyntenyView, GeneView)
Lecture/Lab 7.3
(c) 2005 CGDN

5. Ensembl Gene Annotation
Erin Pleasance
Ensembl Gene Annotation
February 21, 2005
“Basis for initial analysis and publication of most vertebrate genomes”
Genome assembly from NCBI
Gene build system
Targeted gene builds predict known genes
Similarity gene builds predict novel genes
Lecture/Lab 7.3
(c) 2005 CGDN

6. Curwen et al, Genome Res 14: 942-950, 2004
Erin Pleasance
February 21, 2005
Curwen et al, Genome Res 14: , 2004
Lecture/Lab 7.3
(c) 2005 CGDN

7. Targeted gene build Align known proteins with pmatch and BLAST
Erin Pleasance
Targeted gene build
February 21, 2005
Align known proteins with pmatch and BLAST
Incorporate aligned cDNA sequences to find splice sites, UTRs with genewise
ContigView of best in genome gene with associated evidence
UTRs predicted
Known gene (p53)
Proteins aligned
Unigene clusters aligned
cDNAs aligned
Lecture/Lab 7.3
(c) 2005 CGDN

8. Similarity gene build Identify novel exons ab initio using Genscan
Erin Pleasance
Similarity gene build
February 21, 2005
Identify novel exons ab initio using Genscan
Confirm exons by BLAST to known proteins, mRNAs, UniGene clusters
ContigView of homology gene with associated evidence
Novel gene
GenScan predictions
Proteins aligned
Unigene clusters aligned
Lecture/Lab 7.3
(c) 2005 CGDN

9. Ensembl Gene Annotation
Erin Pleasance
Ensembl Gene Annotation
February 21, 2005
Resulting “Ensembl genes” are highly accurate with low false positive rates
Ensembl human gene identifiers are 95% stable between builds
Ensembl and RefSeq differ with % of the genes
The Consensus CDS (CCDS) project is a collaborative effort between Ensembl/EBI, UCSC and NCBI to identify a core set of human protein coding regions that are consistently annotated and of high quality (~13,000 genes).
Lecture/Lab 7.3
(c) 2005 CGDN

10. Manually curated genes: VEGA
Erin Pleasance
February 21, 2005
Some chromosomes contain manually curated genes from VEGA database
“Otter manual annotation system” allows integration of automatic and manual annotations (eg. from Apollo) into Ensembl by The Human and Vertebrate Annotation (HAVANA) group annotators at the Sanger center
VEGA gene
Lecture/Lab 7.3
(c) 2005 CGDN

11. Ensembl EST genes
Erin Pleasance
February 21, 2005
ESTs not accurate enough to produce Ensembl genes, but important for identifying alternative transcripts
ESTs aligned to genome and merged to create an independent set of “EST genes”
Known gene
EST genes
Unigene clusters aligned
Lecture/Lab 7.3
(c) 2005 CGDN

12. Erin Pleasance
Pseudogenes
February 21, 2005
Processed pseudogenes in annotation identified (lack of introns, frameshifts, presence of multi-exon version elsewhere in genome, etc.)
Pseudogene
Lecture/Lab 7.3
(c) 2005 CGDN

13. Erin Pleasance
Noncoding RNA Genes
February 21, 2005
Genes with no ORFs that are functional (tRNAs, rRNAs, miRNAs …)
7220 annotations from Sean Eddy and Tom Jones
miRNAs
Coding gene
Lecture/Lab 7.3
(c) 2005 CGDN

14. Example 1: Exploring Caspase-3
Erin Pleasance
Example 1: Exploring Caspase-3
February 21, 2005
Aim to demonstrate basic browsing and views
Caspase-3 is a gene involved in apoptosis (cell suicide)
We will look at:
Gene annotation
SNPs
Orthologs and genome alignments
Alternative transcripts and EST genes
Protein Structure
Lecture/Lab 7.3
(c) 2005 CGDN

15. Text Search Species-specific homepage caspase-3 Gene Erin Pleasance
February 21, 2005
Gene
caspase-3
Lecture/Lab 7.3
(c) 2005 CGDN

16. GeneView GeneSpliceView GeneRegulationView ContigView GeneSNPView
Erin Pleasance
February 21, 2005
GeneSpliceView
GeneRegulationView
ContigView
GeneSNPView
TransView of transcript
ExonView
ExportView
ProteinView
Orthologs predicted by sequence similarity and synteny
Lecture/Lab 7.3
(c) 2005 CGDN

17. GeneView DAS - Distributed Annotation System
Erin Pleasance
GeneView
February 21, 2005
DAS - Distributed Annotation System
- external annotation of splicing, transcripts, array expression, pubmed links, associated phenotypes, Protonet, Reactome, UniProt.
Information for each Transcript
- similarity matches, links to RefSeq, OMIM, PDB, Array probes, GO, InterPro, Protein FamilyView, transcript structure, protein properties.
Lecture/Lab 7.3
(c) 2005 CGDN

18. GeneView GeneSNPView Erin Pleasance February 21, 2005 Lecture/Lab 7.3
(c) 2005 CGDN

19. GeneSNPView Erin Pleasance February 21, 2005 Lecture/Lab 7.3
(c) 2005 CGDN

20. Other SNP/Haplotype tools
Erin Pleasance
Other SNP/Haplotype tools
February 21, 2005
SNPView – info on a single SNP
ProteinView (protein sequence with SNP markup)
LDView: View linkage disequilibrium (only limited regions)
HaploView: View haplotypes (only limited regions)
Lecture/Lab 7.3
(c) 2005 CGDN

21. GeneView Click Back to Erin Pleasance February 21, 2005
Lecture/Lab 7.3
(c) 2005 CGDN

22. ContigView Chromosome and bands Sequence contigs To Detailed view
Erin Pleasance
February 21, 2005
Chromosome and bands
Sequence contigs
To Detailed view
Lecture/Lab 7.3
(c) 2005 CGDN

23. ContigView: Detailed View
Erin Pleasance
February 21, 2005
See other tracks, options in menus
Genscan predictions
Targetted gene predictions
(2 alternative transcripts)
Gene annotations
EST genes
Other tracks: Aligned sequences etc.
Base View Region
Lecture/Lab 7.3
(c) 2005 CGDN

24. ContigView- Features menu
Erin Pleasance
February 21, 2005
Export image (ps, pdf, svg) or fasta file
Click on ‘close menu’
Lecture/Lab 7.3
(c) 2005 CGDN

25. MultiContigView
Conserved regions
Rat ortholog
Lecture/Lab 7.3

26. Other Comparative Genomics Tools
Up to 6 genome alignments with MLAGAN in AlignSliceView
Other view is SyntenyView
Also access comparative genomics through EnsMart
Lecture/Lab 7.3

27. DAS-Distributed Annotation System
Lecture/Lab 7.3

28. Data Mining with BioMart
Allows very fast, cross-data source querying
Search for genes (features, sequences, etc.) or SNPs based on
Position; function; domains; similarity; expression; etc.
Accessible from Ensembl website (MartView) as well as stand-alone
Extremely powerful for data mining
Lecture/Lab 7.3

29. Erin Pleasance
Example 2: BioMart
February 21, 2005
A new disease locus has been mapped between markers D21S1991 and D21S171. It may be that the gene involved has already been identified as having a role in another disease. What candidates are in this region?
Lecture/Lab 7.3
(c) 2005 CGDN

30. BioMart: Choosing your dataset
Lecture/Lab 7.3

31. BioMart: Filtering 21
D21S1991
D21S171
Lecture/Lab 7.3

32. Note you can output different types of information
BioMart: Output
Note you can output different types of information
Lecture/Lab 7.3

33. BioMart: Output
Lecture/Lab 7.3

34. Sequence Similarity Searching
Use SSAHA for exact matches (fast)
Use BLAST for more distant similarity (slow)
Lecture/Lab 7.3

35. Looking for Help?
Lecture/Lab 7.3

36. DAS: Getting your Own Data in Ensembl
DAS (Distributed Annotation System)
Anyone can load data into Ensembl and allow others to view it in the same view (eg. ContigView) as other Ensembl annotations
Click on ‘Manage sources’ in DAS dropdown menu
Lecture/Lab 7.3

37. Other Ways to Access Ensembl
MySQL database directly accessible
APIs for Perl and Java
Other software
Apollo Java genome annotation viewer/editor
Sockeye Java viewer
You can get your own local version of Ensembl: software and data freely available
Sockeye
Lecture/Lab 7.3

38. Exercises
Erin Pleasance
February 21, 2005
Ex 1. Homologues of human genes are often present in Fugu rubripes in more condensed form (with shorter introns). Is this true for the gene PTEN, a tumor suppressor often mutated in advanced cancers?
Try MultiContigView; can you think of another way to get this information as well?
Ex 2. The microRNA bantam regulates the Drosophila (fruitfly) gene hid by binding the 3’ UTR. Hid is involved in apoptosis, and it is possible that binding sites for bantam could be found in the 3’ UTR of other apoptosis genes as well. Obtain the 3’ UTR sequence of all Drosophila genes known to be involved in apoptosis.
Using BioMart, the GO term for apoptosis is GO: , evidence code TAS
Ex 3. The file “PCR_product.txt” on the webserver contains the sequence of a PCR product amplified from a mouse cDNA library. What gene does the product correspond to? Does it contain the complete coding sequence of that gene?
Would it be better to use BLAST or SSAHA?
Lecture/Lab 7.3
(c) 2005 CGDN