Pairwise sequence alignment
(practice)
1
Manual Work: Write down
the amino acid sequences
derived from all six
possible reading frames of:
>seq1
ACTGTCGC
2
Open the following website
http://biotools.umassmed.edu/cgi-bin/biobin/transeq
Choose all
frames
Remove X
3
Translate all possible reading frames of the
following sequence
TATGTCTCTCACCAACAAGAACGTCGTTTTCGTGGCCGGTCTGGGCGGCATTGGCCTGGAC
ACCAGCCGGGAGTTGGTCAAGCGTAATCTGAAGAACCTGGTCATCCTGGATCGCATTGAC
AATCCGGCTGCCATTGCCGAACTGAAGGCAATCAATCCCAAGGTGACCATCACCTTCTAT
CCCTACGATGTGACTGTGCCCGTCGCTGAGACCACCAAGCTCCTGAAGACCATCTTTGCC
CAGGTGAAGACAATCGATGTCCTGATCAACGGTGCTGGCATCCTGGACGATCATCAGATC
GAGCGCACCATTGCCGTTAACTACACGGGCCTGGTCAACACCACCACAGCCATTCTGGAC
TTCTGGGACAAGCGCAAGGGCGGCCCAGGCGGCATCATTTGCAACATTGGCTCCGTCACC
GGTTTCAATGCCATCTACCAGGTGCCCGTTTACTCCGGCTCCAAGGCGGCGGTGGTTAAC
TTTACCTCCTCCCTGGCGAAACTGGCTCCCATTACTGGTGTCACTGCTTACACTGTCAAT
CCTGGCATCACCAGGACCACTCTGGTCCACAAATTCAACTCGTGGCTGGATGTGGAGCCC
CGTGTGGCGGAGAAGCTGCTCGAGCATCCCACCCAGACCTCTCAGCAGTGCGCCGAGAAC
TTTGTGAAGGCCATTGAGCTGAACAAGAACGGTGCCATCTGGAAGTTGGATTTGGGCACC
TTGGAGCCCATCACATGGACCCAGCACTGGGACTCGGGCATCTAA
Which reading frame(s) is(are) likely to be the
true reading frame(s)?
4
DOT-PLOT
Give the
coordinates of the
boxes to be filled?
Window size :5
Stringency: 5
A T C G G C A T
A
T
C
G
G
C
A
G
Window size :5
Stringency: 2
A T C G G T A T
A
T
C
G
G
C
A
G
A T C G G T A T
A
T
C
G
G
C
A
G
Nucleic Acid Dot Plots (http://www.vivo.colostate.edu/molkit/dnadot/)
Compare Horse
(NM_001164018)
and Chicken
(NM_001081704)
hemoglobin
Copy DNA
sequences
Window size
must be an odd
number
Number of
mismatches
allowed
Use the Dotplot to compare chicken ovomucoid
(NM_001112662) to itself
GCACCGGCAGCCGCCTGCAGAGCCGGGCAGTACCTCACCATGGCCATGGCAGGCGTCTTCGTGCTGTTCT
CTTTCGTGCTTTGTGGCTTCCTCCCAGATGCTGCCTTTGGGGCTGAGGTGGACTGCAGTAGGTTTCCCAA
CGCTACAGACAAGGAAGGCAAAGATGTATTGGTTTGCAACAAGGACCTCCGCCCCATCTGTGGTACCGAT
GGAGTCACTTACACCAACGATTGCTTGCTGTGTGCCTACAGCATAGAATTTGGAACCAATATCAGCAAAG
AGCACGATGGAGAATGCAAGGAAACTGTTCCTATGAACTGCAGTAGTTATGCCAACACGACAAGCGAGGA
CGGAAAAGTGATGGTCCTCTGCAACAGGGCCTTCAACCCCGTCTGTGGTACTGATGGAGTCACCTACGAC
AATGAGTGTCTGCTGTGTGCCCACAAAGTAGAGCAGGGGGCCAGCGTTGACAAGAGGCATGATGGTGGAT
GTAGGAAGGAACTTGCTGCTGTTGACTGCAGCGAGTACCCTAAGCCTGACTGCACGGCAGAAGACAGACC
TCTCTGTGGCTCCGACAACAAAACATATGGCAACAAGTGCAACTTCTGCAATGCAGTCGTGGAAAGCAAC
GGGACTCTCACTTTAAGCCATTTTGGAAAATGCTGAATATCAGAGCTGAGAGAATTCACCACAGGATCCC
CACTGGCGAATCCCAGCGAGAGGTCTCACCTCGGTTCATCTCGCACTCTGGGGAGCTCAGCTCACTCCCG
ATTTTCTTTCTCAATAAACTAAATCAGCAACAAAAAAAAAA
What do these
parallel lines
represent?
LALIGN - finds multiple matching
subsegments in two sequences
Part of the FASTA package of sequence analysis program.
Lalign - compares two protein or DNA sequences for
local or global similarity and shows the local
sequence alignments.
http://www.ch.embnet.org/software/LALIGN_form.html
11
Choose
method
default matrix
Set scoring
matrix and
gap penalties
Paste your
sequence
12
Open
http://www.ch.embnet.org/software/LALIGN_form.html
Use the sequences below and perform a global alignment with
5 “Opening gap penalty” and 0 “Extending gap penalty”.
>seq1
GCGACTGTTCCTATGAACTGCAGTAGTTATGCCAACACG
ACAAGCGAGGACGGAAAAGTGAGTCTGTGGTACTGATG
GAGTCACCTACGACGCGAGGACGCCAGGTG
>seq2
GCGAGGACGGAAAAGTG
13
GLOBAL DOESN’T ALWAYS
WORK.
GLOBAL
14
SOLUTION: LOCAL
15
Write down the amino acid
sequences derived from all
six possible reading frames
of:
>seq1
ACTGTCGC
>seqRC
GCGACAGT
Forward:
>_1 TV >_2 LS >_3 CR
Reverse
>_1 AT >_2 RQ >_3 DS
16