Summer Bioinformatics Workshop 2008
Sequence Alignments
Chi-Cheng Lin, Ph.D.
Associate Professor
Department of Computer Science
Winona State University – Rochester Center
clin@winona.edu
Summer Bioinformatics Workshop 2008
Sequence Alignments
 Cornerstone of bioinformatics
 What is a sequence?
 Nucleotide sequence
 Amino acid sequence
 Pairwise and multiple sequence alignments
 What alignments can help
 Determine function of a newly discovered gene
sequence
 Determine evolutionary relationships among
genes, proteins, and species
 Predict structure and function of protein
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Summer Bioinformatics Workshop 2008
Why Align Sequences?
 The draft human genome is available
 Automated gene finding is possible
 Gene: AGTACGTATCGTATAGCGTAA
 What does it do?
 One approach: Is there a similar gene in
another species?
 Align sequences with known genes
 Find the gene with the “best” match
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Summer Bioinformatics Workshop 2008
Visualization of Sequence Alignment
 Dot Plot
 One of the simplest and oldest methods for
sequence alignment
 Visualization of regions of similarity




Assign one sequence on the horizontal axis
Assign the other on the vertical axis
Place dots on the space of matches
Diagonal lines means adjacent regions of
identity
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Summer Bioinformatics Workshop 2008
A Simple Example
 Construct a simple
dot plot for
T A G T C G A T G
T *
TAGTCGATG
TGGTCATC
*
G
*
*
*
G
*
*
*
T *
 The alignment is
TAGTCGATG
TGGTC-ATC
*
*
C
A
T *
C
*
*
*
*
*
*
*
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Summer Bioinformatics Workshop 2008
Genes Accumulate Mutations over Time
 Mistakes in gene replication
or repair




Deletions, duplications
Insertions, inversions
Translocations
Point mutations
 Environmental factors
 Radiation
 Oxidation
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Summer Bioinformatics Workshop 2008
Deletions
 Codon deletion:
ACG ATA GCG TAT GTA TAG CCG…
 Effect depends on the protein, position, etc.
 Almost always deleterious
 Sometimes lethal
 Frame shift mutation:
ACG ATA GCG TAT GTA TAG CCG…
ACG ATA GCG ATG TAT AGC CG?…
 Almost always lethal
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Summer Bioinformatics Workshop 2008
Indels
 Comparing two genes it is generally
impossible to tell if an indel is an insertion
in one gene, or a deletion in another,
unless ancestry is known:
ACGTCTGATACGCCGTATCGTCTATCT
ACGTCTGAT---CCGTATCGTCTATCT
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Summer Bioinformatics Workshop 2008
The Genetic Code
Substitutions are
mutations
accepted by
natural selection.
Synonymous:
CGC  CGA
Non-synonymous:
GAU  GAA
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Summer Bioinformatics Workshop 2008
Point Mutation Example: Sickle-cell Disease
 Wild-type hemoglobin
DNA
3’----CTT----5’
 Mutant hemoglobin
DNA
3’----CAT----5’
mRNA
5’----GAA----3’
mRNA
5’----GUA----3’
Normal hemoglobin
------[Glu]------
Mutant hemoglobin
------[Val]-----10
Summer Bioinformatics Workshop 2008
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image credit: U.S. Department of Energy Human Genome Program, http://www.ornl.gov/hgmis.
Summer Bioinformatics Workshop 2008
Comparing Two Sequences
 Point mutations, easy:
ACGTCTGATACGCCGTATAGTCTATCT
ACGTCTGATTCGCCCTATCGTCTATCT
 Indels are difficult, must align sequences:
ACGTCTGATACGCCGTATAGTCTATCT
CTGATTCGCATCGTCTATCT
ACGTCTGATACGCCGTATAGTCTATCT
----CTGATTCGC---ATCGTCTATCT
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Summer Bioinformatics Workshop 2008
Scoring a Sequence Alignment
 Example
 Match score:
 Mismatch score:
 Gap penalty:
+1
+0
–1
ACGTCTGATACGCCGTATAGTCTATCT
||||| |||
|| ||||||||
----CTGATTCGC---ATCGTCTATCT
 Matches: 18 × (+1)
 Mismatches: 2 × 0
 Gaps: 7 × (– 1)
Score = 18 + 0 + (-7) = +11
 Various scoring scheme exist.
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Summer Bioinformatics Workshop 2008
How can we find an optimal alignment?
 Finding the alignment is computationally
hard:
ACGTCTGATACGCCGTATAGTCTATCT
CTGAT---TCG-CATCGTC--T-ATCT
 There are ~888,000 possibilities to align
the two sequences given above.
 Algorithms using a technique called
“dynamic programming” are used – out of
the scope of this workshop.
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Summer Bioinformatics Workshop 2008
Global and Local Alignments
 Global alignments – score the entire alignment
 Local alignment – find the best matching
subsequence
 Why local sequence alignment?
 Global alignment is useful only if the sequences to be
aligned are very similar
 Subsequence comparison between a DNA sequence
and a genome
 Identify


Conserved regions
Protein function domains
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Summer Bioinformatics Workshop 2008
Example
 Compare the two sequences:
TTGACACCCTCCCAATT
ACCCCAGGCTTTACACAG
 Global alignment (does it look good?)
TTGACACCCTCC-CAATT
|| ||
||
ACCCCAGGCTTTACACAG
 Local alignment (does it look good?)
---------TTGACACCCTCCCAATT
|| ||||
ACCCCAGGCTTTACACAG-------16
Summer Bioinformatics Workshop 2008
Where do we get sequences to work with?
 Biological databases
 NCBI Entrez
(http://www.ncbi.nlm.nih.gov/gquery/gquery.fcgi
?term=)




Wet labs
Simulations
Other people’s results
On-line education resources
 BEDROCK (http://www.bioquest.org/bedrock/)
 BLAST results
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