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Comparative Genomics
Bioinformatic Tools for Comparative Genomics of Vectors
Overview
 Comparing Genomes
 Homologies and Families
 Sequence Alignments
Bioinformatic Tools for Comparative Genomics of Vectors
Comparative Genomics
 Allows us to achieve a greater understanding of vertebrate
evolution
 Tells us what is common and what is unique between different
species at the genome level
 The function of human genes and other regions may be
revealed by studying their counterparts in lower organisms
 Helps identify both coding and non-coding genes and
regulatory elements
Bioinformatic Tools for Comparative Genomics of Vectors
Sequence Conservation Over Time
Bioinformatic Tools for Comparative Genomics of Vectors
Non Coding Regions
 Large stretches of non-coding regions in vertebrates
 Regulatory regions of:
Developmental genes
Transcription factors
miRNA
Bioinformatic Tools for Comparative Genomics of Vectors
Kikuta et al., Genome Research, May 2007
Methods of Alignment- Ensembl

BLASTZ-net (comparison on nucleotide level) is used for species
that are evolutionary close, e.g. human – mouse
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Translated BLAT (comparison on amino acid level) is used for
evolutionary more distant species, e.g. human – zebrafish

PECAN global alignment used for multispecies alignments
Bioinformatic Tools for Comparative Genomics of Vectors
Why Compare Genomes?
We can better understand evolution/ speciation
We can find important, functional regions of the sequence (codons,
promoters, regulatory regions)
It can help us locate genes in other species that are missing or not welldefined (also through comparison and alignments).
Quality control!
Bioinformatic Tools for Comparative Genomics of Vectors
Evolution at the DNA Level
Deletion
Mutation
…ACTGACATGTACCA…
Sequence edits
…AC----CATGCACCA…
Rearrangements
Inversion
Translocation
Duplication
Bioinformatic Tools for Comparative Genomics of Vectors
Comparing Genomes
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Mammals have roughly 3 billion base pairs in their genomes
Over 98% human genes are shared with primates, with more than 9598% similarity between genes.
Even the fruit fly shares 60% of its genes with humans! (March 2000)
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Compare human & Mouse
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40% of human genome align with mouse
24% of human genome missing in mouse (also mouse-specific sequences)
Bioinformatic Tools for Comparative Genomics of Vectors
Improving Gene Quality
Comparative genomics
predicts one long
transcript.
Bioinformatic Tools for Comparative Genomics of Vectors
Pseudogene recovery
chr 3
human
mouse
rat
dog
cow
chr X
We find 67
confident cases
where a human
protein is closer to
the ancestor than
any extant species
in the alignment
Bioinformatic Tools for Comparative Genomics of Vectors
How Does Ensembl Predict Homology?
• Uses all the species
• Prediction pipeline: Begins with
BLAST and sequence clustering
• Compares gene relationships to
species relationships
Bioinformatic Tools for Comparative Genomics of Vectors
BSR: Blast Score Ratio. When 2 proteins P1 and P2 are compared, BSR=scoreP1P2/max(selfscoreP1 or self-scoreP2). The default threshold used in the initial clustering step is 0.33.
Orthologue / Paralogue Prediction Algorithm
(1) Load the longest translation of each gene from all species used in Ensembl.
(2) Run WUBLASTp+SW of every gene against every other (both self and non-self
species) in a genome-wide manner.
(3) Build a graph of gene relations based on Best Reciprocal Hits (BRH) and Blast Score
Ratio (BSR) values.
(4) Extract the connected components (=single linkage clusters), each cluster
representing a gene family.
(5) For each cluster, build a multiple alignment based on the protein sequences using
MUSCLE.
(6) For each aligned cluster, build a phylogenetic tree using PHYML. An unrooted tree is
obtained at this stage.
(7) Reconcile each gene tree with the species tree to call duplication event on internal
nodes and root the tree (TreeBeSt).
(8) From each gene tree, infer gene pairwise relations of orthology and paralogy types.
Bioinformatic Tools for Comparative Genomics of Vectors
Species Tree
Anopheles gambiae
Aedes aegypti
Drosophila melanogaster
Dasypus novemcinctus
Loxodonta africana
Echinops telfairi
Tupaia belangeri
Homo sapiens
Pan troglodytes
Macaca mulatta
Otolemur garnettii
Mus musculus
Rattus norvegicus
Spermophilus tridecemlineatus
Cavia porcellus
Oryctolagus cuniculus
Erinaceus europaeus
Myotis lucifugus
Canis familiaris
Felis catus
Bos taurus
Monodelphis domestica
Ornithorhynchus anatinus
Gallus gallus
Xenopus tropicalis
Gasterosteus aculeatus
Oryzias latipes
Takifugu rubripes
Tetraodon nigroviridis
Danio rerio
Ciona intestinalis
Ciona savignyi
Caenorhabditis elegans
Saccharomyces cerevisiae
Bioinformatic Tools for Comparative Genomics of Vectors
Species and Gene Trees
Phylogenetic Tree Reconciliation: the Species/Gene Tree Problem
Dufayard et al. ERCIM News No. 43 October 2000
Bioinformatic Tools for Comparative Genomics of Vectors
Genes/Species Tree reconciliation: TreeBeST
Bioinformatic Tools for Comparative Genomics of Vectors
Reconciliation
M
M
Duplication node
Speciation node
R
R
H
species tree
H
M
M
R’
H
H
R
H’
unrooted gene tree
M’
R
Viewing Trees in Ensembl

GeneView page

GeneTreeView
Bioinformatic Tools for Comparative Genomics of Vectors
Types of Homologues
Orthologs : any gene pairwise relation where the ancestor node is a
speciation event
Paralogs : any gene pairwise relation where the ancestor node is a
duplication event
Bioinformatic Tools for Comparative Genomics of Vectors
Orthologue and Paralogue Types
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ortholog_one2one
ortholog_one2many
ortholog_many2many
apparent_ortholog_one2one
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within_species_paralog
between_species_paralog
Bioinformatic Tools for Comparative Genomics of Vectors
Ortholog and Paralog types
Ortholog and Paralog types
Bioinformatic Tools for Comparative Genomics of Vectors
Orthologues on GeneView
What is ‘1 to 1’?
What is ‘1 to many’?
Bioinformatic Tools for Comparative Genomics of Vectors
Protein Families
 How: Cluster proteins for every isoform
(transcript) in every species.
 Why: Predict a function for ‘novel’
genes/proteins
Understand gene relationships
Bioinformatic Tools for Comparative Genomics of Vectors
Protein Dataset
More than 1,800,000 proteins clustered:
 All Ensembl protein predictions from
all species supported
895,070 protein predictions
 All metazoan (animal) proteins in UniProt:
96,030 UniProtKB/Swiss-Prot
892,0208 UniProtKB/TrEMBL
Bioinformatic Tools for Comparative Genomics of Vectors
Clustering Strategy
 BLASTP all-versus-all comparison
 Markov clustering
 For each cluster:
 Calculation of multiple sequence alignments with
ClustalW
 Assignment of a consensus description
Bioinformatic Tools for Comparative Genomics of Vectors
Where are Families
shown? ProtView
Link to
FamilyView
Bioinformatic Tools for Comparative Genomics of Vectors
Where are Families shown?
FamilyView
JalView multiple
alignments
Ensembl family
members within
human
Ensembl family
members in other
species
Bioinformatic Tools for Comparative Genomics of Vectors
 Comparing Genomes
 Homologies and Families
 Sequence alignments
Bioinformatic Tools for Comparative Genomics of Vectors
Aligning Whole Genomes- Why?
• To identify homologous regions
• To spot trouble gene predictions
• Conserved regions could be functional
• To define syntenic regions (long regions of DNA sequences
where order and orientation is highly conserved)
Bioinformatic Tools for Comparative Genomics of Vectors
Aligning large genomic sequences
 Should find all highly similar regions between two sequences
 Should allow for segments without similarity, rearrangements
etc.
 Issues
 Heavy process
 Scalability, as more and more genomes are sequenced
 Time constraint
Bioinformatic Tools for Comparative Genomics of Vectors
Whole Genome Multiple Alignments
 Enredo
 Defines orthology map (co-linear regions)
 Supports segmental duplications
 Pecan
 Consistency based multiple aligner
 Optimized to cope with long DNA sequences
 Ortheus
 Ancestral sequences reconstructor
 Inferring the history of insertion and deletions
In ContigView...
Bioinformatic Tools for Comparative Genomics of Vectors
Multiple Alignments using
PECAN
 Currently 2 sets:
 10 amniota vertebrates:
 7 eutherian mammals:
To come… the fish!
Bioinformatic Tools for Comparative Genomics of Vectors
Alignment Strategy
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Use all coding exons
Get sets of best reciprocal hits
Create orthology maps
Build multiple global alignments
Bioinformatic Tools for Comparative Genomics of Vectors
View Alignments: ContigView
In the Detailed View Panel:
Bioinformatic Tools for Comparative Genomics of Vectors
View Conservation: ContigView
Click on a Pink Bar for AlignSliceView… export alignments
Bioinformatic Tools for Comparative Genomics of Vectors
AlignSliceView
Bioinformatic Tools for Comparative Genomics of Vectors
GeneSeqalignView
Bioinformatic Tools for Comparative Genomics of Vectors
GeneSeqalignView
Bioinformatic Tools for Comparative Genomics of Vectors
MultiContigView
Comparison of chromosomes in
multiple species.
(Links from SyntenyView, ContigView,
CytoView)
Bioinformatic Tools for Comparative Genomics of Vectors
Export Alignments in BioMart
Choose ‘Compara pairwise alignments’
Bioinformatic Tools for Comparative Genomics of Vectors
Syntenic Regions
 Genome alignments are compiled into larger syntenic
regions
 Alignments are clustered together when the relative
distance between them is less than 100 kb and order
and orientation are consistent
 Any clusters less than 100 kb are discarded
Bioinformatic Tools for Comparative Genomics of Vectors
Enredo
Anchors
500.000 anchors
for mammals
--more than 1 anchor
per 10Kb
Supports segmental
duplications!!
Covers 90% of the human
protein coding genes
(Hsap-Mmus-Rnor-Cfam-Btau)
Bioinformatic Tools for Comparative Genomics of Vectors
SyntenyView
Human
chromosome
Orthologues
Mouse
chromosomes
Mouse
chromosomes
Bioinformatic Tools for Comparative Genomics of Vectors
CytoView
Syntenic
blocks
Bioinformatic Tools for Comparative Genomics of Vectors
Summary
 View Homology in pages such as GeneView, ProtView,
SyntenyView, GeneTreeView, or BioMart
 View Protein Family information in FamilyView
 View Alignments in ContigView, GeneSeqAlign View,
through BioMart
Bioinformatic Tools for Comparative Genomics of Vectors