|
|
water |
A local alignment searches for regions of local similarity between two sequences and need not include the entire length of the sequences. Local alignment methods are very useful for scanning databases or other circumsatnces when you wish to find matches between small regions of sequences, for example between protein domains.
Dynamic programming methods ensure the optimal local alignment by exploring all possible alignments and choosing the best. It does this by reading in a scoring matrix that contains values for every possible residue or nucleotide match. water finds an alignment with the maximum possible score where the score of an alignment is equal to the sum of the matches taken from the scoring matrix.
An important problem is the treatment of gaps, i.e., spaces inserted to optimise the alignment score. A penalty is subtracted from the score for each gap opened (the 'gap open' penalty) and a penalty is subtracted from the score for the total number of gap spaces multiplied by a cost (the 'gap extension' penalty).
Typically, the cost of extending a gap is set to be 5-10 times lower than the cost for opening a gap.
There are two ways to compute a penalty for a gap of n positions :
gap opening penalty + (n - 1) * gap extension penalty gap penalty + n * gap length penalty
The first way is used by EMBOSS and WU-BLAST
The second way is used by NCBI-BLAST, GCG, Staden and CLUSTAL.
Fasta used it for a long time the first way, but Prof. Pearson decided
recently to shift to the second.
The two methods are basically equivalent.
The Smith-Waterman algorithm contains no negative scores in the path matrix it creates. The algorithm starts the alignment at the highest path matrix score and works backwards until a cell contains zero.
See the Reference Smith et al. for details.
% water tsw:hba_human tsw:hbb_human Smith-Waterman local alignment. Gap opening penalty [10.0]: Gap extension penalty [0.5]: Output alignment [hba_human.water]: |
Go to the input files for this example
Go to the output files for this example
Standard (Mandatory) qualifiers:
[-asequence] sequence Sequence filename and optional format, or
reference (input USA)
[-bsequence] seqall Sequence(s) filename and optional format, or
reference (input USA)
-gapopen float [10.0 for any sequence] The gap open penalty
is the score taken away when a gap is
created. The best value depends on the
choice of comparison matrix. The default
value assumes you are using the EBLOSUM62
matrix for protein sequences, and the
EDNAFULL matrix for nucleotide sequences.
(Number from 0.000 to 100.000)
-gapextend float [0.5 for any sequence] The gap extension
penalty is added to the standard gap penalty
for each base or residue in the gap. This
is how long gaps are penalized. Usually you
will expect a few long gaps rather than many
short gaps, so the gap extension penalty
should be lower than the gap penalty. An
exception is where one or both sequences are
single reads with possible sequencing
errors in which case you would expect many
single base gaps. You can get this result by
setting the gap open penalty to zero (or
very low) and using the gap extension
penalty to control gap scoring. (Number from
0.000 to 10.000)
[-outfile] align [*.water] Output alignment file name
Additional (Optional) qualifiers:
-datafile matrixf [EBLOSUM62 for protein, EDNAFULL for DNA]
This is the scoring matrix file used when
comparing sequences. By default it is the
file 'EBLOSUM62' (for proteins) or the file
'EDNAFULL' (for nucleic sequences). These
files are found in the 'data' directory of
the EMBOSS installation.
Advanced (Unprompted) qualifiers:
-[no]brief boolean [Y] Brief identity and similarity
Associated qualifiers:
"-asequence" associated qualifiers
-sbegin1 integer Start of the sequence to be used
-send1 integer End of the sequence to be used
-sreverse1 boolean Reverse (if DNA)
-sask1 boolean Ask for begin/end/reverse
-snucleotide1 boolean Sequence is nucleotide
-sprotein1 boolean Sequence is protein
-slower1 boolean Make lower case
-supper1 boolean Make upper case
-sformat1 string Input sequence format
-sdbname1 string Database name
-sid1 string Entryname
-ufo1 string UFO features
-fformat1 string Features format
-fopenfile1 string Features file name
"-bsequence" associated qualifiers
-sbegin2 integer Start of each sequence to be used
-send2 integer End of each sequence to be used
-sreverse2 boolean Reverse (if DNA)
-sask2 boolean Ask for begin/end/reverse
-snucleotide2 boolean Sequence is nucleotide
-sprotein2 boolean Sequence is protein
-slower2 boolean Make lower case
-supper2 boolean Make upper case
-sformat2 string Input sequence format
-sdbname2 string Database name
-sid2 string Entryname
-ufo2 string UFO features
-fformat2 string Features format
-fopenfile2 string Features file name
"-outfile" associated qualifiers
-aformat3 string Alignment format
-aextension3 string File name extension
-adirectory3 string Output directory
-aname3 string Base file name
-awidth3 integer Alignment width
-aaccshow3 boolean Show accession number in the header
-adesshow3 boolean Show description in the header
-ausashow3 boolean Show the full USA in the alignment
-aglobal3 boolean Show the full sequence in alignment
General qualifiers:
-auto boolean Turn off prompts
-stdout boolean Write standard output
-filter boolean Read standard input, write standard output
-options boolean Prompt for standard and additional values
-debug boolean Write debug output to program.dbg
-verbose boolean Report some/full command line options
-help boolean Report command line options. More
information on associated and general
qualifiers can be found with -help -verbose
-warning boolean Report warnings
-error boolean Report errors
-fatal boolean Report fatal errors
-die boolean Report dying program messages
|
| Standard (Mandatory) qualifiers | Allowed values | Default | |
|---|---|---|---|
| [-asequence] (Parameter 1) |
Sequence filename and optional format, or reference (input USA) | Readable sequence | Required |
| [-bsequence] (Parameter 2) |
Sequence(s) filename and optional format, or reference (input USA) | Readable sequence(s) | Required |
| -gapopen | The gap open penalty is the score taken away when a gap is created. The best value depends on the choice of comparison matrix. The default value assumes you are using the EBLOSUM62 matrix for protein sequences, and the EDNAFULL matrix for nucleotide sequences. | Number from 0.000 to 100.000 | 10.0 for any sequence |
| -gapextend | The gap extension penalty is added to the standard gap penalty for each base or residue in the gap. This is how long gaps are penalized. Usually you will expect a few long gaps rather than many short gaps, so the gap extension penalty should be lower than the gap penalty. An exception is where one or both sequences are single reads with possible sequencing errors in which case you would expect many single base gaps. You can get this result by setting the gap open penalty to zero (or very low) and using the gap extension penalty to control gap scoring. | Number from 0.000 to 10.000 | 0.5 for any sequence |
| [-outfile] (Parameter 3) |
Output alignment file name | Alignment output file | <*>.water |
| Additional (Optional) qualifiers | Allowed values | Default | |
| -datafile | This is the scoring matrix file used when comparing sequences. By default it is the file 'EBLOSUM62' (for proteins) or the file 'EDNAFULL' (for nucleic sequences). These files are found in the 'data' directory of the EMBOSS installation. | Comparison matrix file in EMBOSS data path | EBLOSUM62 for protein EDNAFULL for DNA |
| Advanced (Unprompted) qualifiers | Allowed values | Default | |
| -[no]brief | Brief identity and similarity | Boolean value Yes/No | Yes |
ID HBA_HUMAN Reviewed; 142 AA.
AC P69905; P01922; Q96KF1; Q9NYR7;
DT 21-JUL-1986, integrated into UniProtKB/Swiss-Prot.
DT 23-JAN-2007, sequence version 2.
DT 03-APR-2007, entry version 41.
DE Hemoglobin subunit alpha (Hemoglobin alpha chain) (Alpha-globin).
GN Name=HBA1;
GN and
GN Name=HBA2;
OS Homo sapiens (Human).
OC Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi;
OC Mammalia; Eutheria; Euarchontoglires; Primates; Haplorrhini;
OC Catarrhini; Hominidae; Homo.
OX NCBI_TaxID=9606;
RN [1]
RP NUCLEOTIDE SEQUENCE [GENOMIC DNA] (HBA1).
RX MEDLINE=81088339; PubMed=7448866; DOI=10.1016/0092-8674(80)90347-5;
RA Michelson A.M., Orkin S.H.;
RT "The 3' untranslated regions of the duplicated human alpha-globin
RT genes are unexpectedly divergent.";
RL Cell 22:371-377(1980).
RN [2]
RP NUCLEOTIDE SEQUENCE [MRNA] (HBA2).
RX MEDLINE=80137531; PubMed=6244294;
RA Wilson J.T., Wilson L.B., Reddy V.B., Cavallesco C., Ghosh P.K.,
RA Deriel J.K., Forget B.G., Weissman S.M.;
RT "Nucleotide sequence of the coding portion of human alpha globin
RT messenger RNA.";
RL J. Biol. Chem. 255:2807-2815(1980).
RN [3]
RP NUCLEOTIDE SEQUENCE [GENOMIC DNA] (HBA2).
RX MEDLINE=81175088; PubMed=6452630;
RA Liebhaber S.A., Goossens M.J., Kan Y.W.;
RT "Cloning and complete nucleotide sequence of human 5'-alpha-globin
RT gene.";
RL Proc. Natl. Acad. Sci. U.S.A. 77:7054-7058(1980).
RN [4]
RP NUCLEOTIDE SEQUENCE [GENOMIC DNA].
RX PubMed=6946451;
RA Orkin S.H., Goff S.C., Hechtman R.L.;
RT "Mutation in an intervening sequence splice junction in man.";
RL Proc. Natl. Acad. Sci. U.S.A. 78:5041-5045(1981).
RN [5]
RP NUCLEOTIDE SEQUENCE [GENOMIC DNA], AND VARIANT LYS-32.
RX MEDLINE=21303311; PubMed=11410421;
RA Zhao Y., Xu X.;
RT "Alpha2(CD31 AGG-->AAG, Arg-->Lys) causing non-deletional alpha-
RT thalassemia in a Chinese family with HbH disease.";
RL Haematologica 86:541-542(2001).
RN [6]
[Part of this file has been deleted for brevity]
FT /FTId=VAR_002840.
FT VARIANT 131 131 A -> D (in Yuda; O(2) affinity down).
FT /FTId=VAR_002842.
FT VARIANT 131 131 A -> P (in Sun Prairie; unstable).
FT /FTId=VAR_002841.
FT VARIANT 132 132 S -> P (in Questembert; highly unstable;
FT causes alpha-thalassemia).
FT /FTId=VAR_002843.
FT VARIANT 134 134 S -> R (in Val de Marne; O(2) affinity
FT up).
FT /FTId=VAR_002844.
FT VARIANT 136 136 V -> E (in Pavie).
FT /FTId=VAR_002845.
FT VARIANT 137 137 L -> M (in Chicago).
FT /FTId=VAR_002846.
FT VARIANT 137 137 L -> P (in Bibba; unstable; causes alpha-
FT thalassemia).
FT /FTId=VAR_002847.
FT VARIANT 139 139 S -> P (in Attleboro; O(2) affinity up).
FT /FTId=VAR_002848.
FT VARIANT 140 140 K -> E (in Hanamaki; O(2) affinity up).
FT /FTId=VAR_002849.
FT VARIANT 140 140 K -> T (in Tokoname; O(2) affinity up).
FT /FTId=VAR_002850.
FT VARIANT 141 141 Y -> H (in Rouen; O(2) affinity up).
FT /FTId=VAR_002851.
FT VARIANT 142 142 R -> C (in Nunobiki; O(2) affinity up).
FT /FTId=VAR_002852.
FT VARIANT 142 142 R -> H (in Suresnes; O(2) affinity up).
FT /FTId=VAR_002854.
FT VARIANT 142 142 R -> L (in Legnano; O(2) affinity up).
FT /FTId=VAR_002853.
FT VARIANT 142 142 R -> P (in Singapore).
FT /FTId=VAR_002855.
FT HELIX 4 15
FT HELIX 16 20
FT HELIX 21 35
FT HELIX 37 42
FT HELIX 53 71
FT HELIX 73 75
FT HELIX 76 79
FT HELIX 81 89
FT HELIX 96 112
FT TURN 114 116
FT HELIX 119 136
FT TURN 137 139
SQ SEQUENCE 142 AA; 15258 MW; 15E13666573BBBAE CRC64;
MVLSPADKTN VKAAWGKVGA HAGEYGAEAL ERMFLSFPTT KTYFPHFDLS HGSAQVKGHG
KKVADALTNA VAHVDDMPNA LSALSDLHAH KLRVDPVNFK LLSHCLLVTL AAHLPAEFTP
AVHASLDKFL ASVSTVLTSK YR
//
|
ID HBB_HUMAN Reviewed; 147 AA.
AC P68871; P02023; Q13852; Q14481; Q14510; Q45KT0; Q6FI08; Q8IZI1;
AC Q9BX96; Q9UCP8; Q9UCP9;
DT 21-JUL-1986, integrated into UniProtKB/Swiss-Prot.
DT 23-JAN-2007, sequence version 2.
DT 20-MAR-2007, entry version 43.
DE Hemoglobin subunit beta (Hemoglobin beta chain) (Beta-globin).
GN Name=HBB;
OS Homo sapiens (Human).
OC Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi;
OC Mammalia; Eutheria; Euarchontoglires; Primates; Haplorrhini;
OC Catarrhini; Hominidae; Homo.
OX NCBI_TaxID=9606;
RN [1]
RP NUCLEOTIDE SEQUENCE.
RX MEDLINE=81064667; PubMed=6254664; DOI=10.1016/0092-8674(80)90428-6;
RA Lawn R.M., Efstratiadis A., O'Connell C., Maniatis T.;
RT "The nucleotide sequence of the human beta-globin gene.";
RL Cell 21:647-651(1980).
RN [2]
RP NUCLEOTIDE SEQUENCE.
RX MEDLINE=77126403; PubMed=1019344;
RA Marotta C., Forget B., Cohen-Solal M., Weissman S.M.;
RT "Nucleotide sequence analysis of coding and noncoding regions of human
RT beta-globin mRNA.";
RL Prog. Nucleic Acid Res. Mol. Biol. 19:165-175(1976).
RN [3]
RP NUCLEOTIDE SEQUENCE.
RA Lu L., Hu Z.H., Du C.S., Fu Y.S.;
RT "DNA sequence of the human beta-globin gene isolated from a healthy
RT Chinese.";
RL Submitted (JUN-1997) to the EMBL/GenBank/DDBJ databases.
RN [4]
RP NUCLEOTIDE SEQUENCE, AND VARIANT DURHAM-N.C. PRO-115.
RC TISSUE=Blood;
RA Kutlar F., Abboud M., Leithner C., Holley L., Brisco J., Kutlar A.;
RT "Electrophoretically silent, very unstable, thalassemic mutation at
RT codon 114 of beta globin (hemoglobin Durham-N.C.) detected by cDNA
RT sequencing of mRNA, from a Russian women.";
RL Submitted (AUG-1999) to the EMBL/GenBank/DDBJ databases.
RN [5]
RP NUCLEOTIDE SEQUENCE, AND VARIANT LOUISVILLE LEU-43.
RC TISSUE=Blood;
RA Kutlar F., Harbin J., Brisco J., Kutlar A.;
RT "Rapid detection of electrophoretically silent, unstable human
RT hemoglobin 'Louisville', (Beta; Phe 42 Leu/TTT to CTT) by cDNA
RT sequencing of mRNA.";
RL Submitted (JAN-1999) to the EMBL/GenBank/DDBJ databases.
RN [6]
RP NUCLEOTIDE SEQUENCE, AND VARIANT TY GARD GLN-125.
[Part of this file has been deleted for brevity]
FT VARIANT 141 141 A -> T (in St Jacques: O(2) affinity up).
FT /FTId=VAR_003081.
FT VARIANT 141 141 A -> V (in Puttelange; polycythemia; O(2)
FT affinity up).
FT /FTId=VAR_003082.
FT VARIANT 142 142 L -> R (in Olmsted; unstable).
FT /FTId=VAR_003083.
FT VARIANT 143 143 A -> D (in Ohio; O(2) affinity up).
FT /FTId=VAR_003084.
FT VARIANT 144 144 H -> D (in Rancho Mirage).
FT /FTId=VAR_003085.
FT VARIANT 144 144 H -> P (in Syracuse; O(2) affinity up).
FT /FTId=VAR_003087.
FT VARIANT 144 144 H -> Q (in Little Rock; O(2) affinity
FT up).
FT /FTId=VAR_003086.
FT VARIANT 144 144 H -> R (in Abruzzo; O(2) affinity up).
FT /FTId=VAR_003088.
FT VARIANT 145 145 K -> E (in Mito; O(2) affinity up).
FT /FTId=VAR_003089.
FT VARIANT 146 146 Y -> C (in Rainier; O(2) affinity up).
FT /FTId=VAR_003090.
FT VARIANT 146 146 Y -> H (in Bethesda; O(2) affinity up).
FT /FTId=VAR_003091.
FT VARIANT 147 147 H -> D (in Hiroshima; O(2) affinity up).
FT /FTId=VAR_003092.
FT VARIANT 147 147 H -> L (in Cowtown; O(2) affinity up).
FT /FTId=VAR_003093.
FT VARIANT 147 147 H -> P (in York; O(2) affinity up).
FT /FTId=VAR_003094.
FT VARIANT 147 147 H -> Q (in Kodaira; O(2) affinity up).
FT /FTId=VAR_003095.
FT HELIX 5 15
FT TURN 20 22
FT HELIX 23 34
FT HELIX 36 41
FT HELIX 43 45
FT HELIX 51 56
FT HELIX 58 76
FT TURN 77 79
FT HELIX 81 93
FT TURN 94 96
FT HELIX 101 118
FT HELIX 119 121
FT HELIX 124 141
FT HELIX 143 145
SQ SEQUENCE 147 AA; 15998 MW; A31F6D621C6556A1 CRC64;
MVHLTPEEKS AVTALWGKVN VDEVGGEALG RLLVVYPWTQ RFFESFGDLS TPDAVMGNPK
VKAHGKKVLG AFSDGLAHLD NLKGTFATLS ELHCDKLHVD PENFRLLGNV LVCVLAHHFG
KEFTPPVQAA YQKVVAGVAN ALAHKYH
//
|
The output is a standard EMBOSS alignment file.
The results can be output in one of several styles by using the command-line qualifier -aformat xxx, where 'xxx' is replaced by the name of the required format. Some of the alignment formats can cope with an unlimited number of sequences, while others are only for pairs of sequences.
The available multiple alignment format names are: unknown, multiple, simple, fasta, msf, trace, srs
The available pairwise alignment format names are: pair, markx0, markx1, markx2, markx3, markx10, srspair, score
See: http://emboss.sf.net/docs/themes/AlignFormats.html for further information on alignment formats.
########################################
# Program: water
# Rundate: Sun 15 Jul 2007 12:00:00
# Commandline: water
# [-asequence] tsw:hba_human
# [-bsequence] tsw:hbb_human
# Align_format: srspair
# Report_file: hba_human.water
########################################
#=======================================
#
# Aligned_sequences: 2
# 1: HBA_HUMAN
# 2: HBB_HUMAN
# Matrix: EBLOSUM62
# Gap_penalty: 10.0
# Extend_penalty: 0.5
#
# Length: 145
# Identity: 63/145 (43.4%)
# Similarity: 88/145 (60.7%)
# Gaps: 8/145 ( 5.5%)
# Score: 293.5
#
#
#=======================================
HBA_HUMAN 3 LSPADKTNVKAAWGKVGAHAGEYGAEALERMFLSFPTTKTYFPHF-DLS- 50
|:|.:|:.|.|.|||| :..|.|.|||.|:.:.:|.|:.:|..| |||
HBB_HUMAN 4 LTPEEKSAVTALWGKV--NVDEVGGEALGRLLVVYPWTQRFFESFGDLST 51
HBA_HUMAN 51 ----HGSAQVKGHGKKVADALTNAVAHVDDMPNALSALSDLHAHKLRVDP 96
.|:.:||.|||||..|.::.:||:|::....:.||:||..||.|||
HBB_HUMAN 52 PDAVMGNPKVKAHGKKVLGAFSDGLAHLDNLKGTFATLSELHCDKLHVDP 101
HBA_HUMAN 97 VNFKLLSHCLLVTLAAHLPAEFTPAVHASLDKFLASVSTVLTSKY 141
.||:||.:.|:..||.|...||||.|.|:..|.:|.|:..|..||
HBB_HUMAN 102 ENFRLLGNVLVCVLAHHFGKEFTPPVQAAYQKVVAGVANALAHKY 146
#---------------------------------------
#---------------------------------------
|
The Identity: is the percentage of identical matches between the two sequences over the reported aligned region (including any gaps in the length).
The Similarity: is the percentage of matches between the two sequences over the reported aligned region (including any gaps in the length).
EMBOSS data files are distributed with the application and stored in the standard EMBOSS data directory, which is defined by the EMBOSS environment variable EMBOSS_DATA.
To see the available EMBOSS data files, run:
% embossdata -showall
To fetch one of the data files (for example 'Exxx.dat') into your current directory for you to inspect or modify, run:
% embossdata -fetch -file Exxx.dat
Users can provide their own data files in their own directories. Project specific files can be put in the current directory, or for tidier directory listings in a subdirectory called ".embossdata". Files for all EMBOSS runs can be put in the user's home directory, or again in a subdirectory called ".embossdata".
The directories are searched in the following order:
water is for aligning the best matching subsequences of two sequences. It does not necessarily align whole sequences against each other; you should use needle if you wish to align closely related sequences along their whole lengths.
A true Smith Waterman implementation like water needs memory proportional to the product of the sequence lengths. For two sequences of length 10,000,000 and 1,000 it therefore needs memory proportional to 10,000,000,000 characters. Two arrays of this size are produced, one of ints and one of floats so multiply that figure by 8 to get the memory usage in bytes. That doesn't include other overheads. Therefore only use water and needle for accurate alignment of reasonably short sequences.
If you run out of memory, try using supermatcher or matcher.
Uncaught exception Assertion failed raised at ajmem.c:xxx
Probably means you have run out of memory. Try using supermatcher or matcher if this happens.
| Program name | Description |
|---|---|
| matcher | Finds the best local alignments between two sequences |
| seqmatchall | All-against-all comparison of a set of sequences |
| supermatcher | Match large sequences against one or more other sequences |
| wordfinder | Match large sequences against one or more other sequences |
| wordmatch | Finds all exact matches of a given size between 2 sequences |
matcher is a local alignment program that gives as good an alignment as
water
but it uses far less memory. However,water
runs twice as fast as matcher.supermatcher is designed for local alignments of very large sequences. It gives good results as long as there is not significant amounts of insertion or deletion in the alignment.
supermatcher Finds a match of a large sequence against one or more sequences matcher Finds the best local alignments between two sequences
Modified 27th July 1999 - tweaking scoring.
Modified 22 Oct 2000 - added ID and Similarity scores.