needle

 

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Function

Needleman-Wunsch global alignment of two sequences

Description

needle reads two input sequences and writes their optimal global sequence alignment to file. It uses the Needleman-Wunsch alignment algorithm to find the optimum alignment (including gaps) of two sequences along their entire length. The algorithm uses a dynamic programming method to ensure the alignment is optimum, by exploring all possible alignments and choosing the best. A scoring matrix is read that contains values for every possible residue or nucleotide match. Needle finds the 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, minus penalties arising from opening and extending gaps in the aligned sequences. The substitution matrix and gap opening and extension penalties are user-specified.

Algorithm

The Needleman-Wunsch algorithm is a member of the class of algorithms that can calculate the best score and alignment of two sequences in the order of mn steps, where n and m are the sequence lengths. These dynamic programming algorithms were first developed for protein sequence comparison by Needleman and Wunsch, though similar methods were independently devised during the late 1960's and early 1970's for use in the fields of speech processing and computer science.

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.

Penalty for a gap of n positions is calculated using the following formula:

gap opening penalty + (n - 1) * gap extension penalty

In a Needleman-Wunsch global alignment, the entire length of each sequence is aligned. The sequences might be partially overlapping or one sequence might be aligned entirely internally to the other. There is no penalty for the hanging ends of the overlap. In bioinformatics, it is usually reasonable to assume that the sequences are incomplete and there should be no penalty for failing to align the missing bases.

Usage

Here is a sample session with needle


% needle tsw:hba_human tsw:hbb_human 
Needleman-Wunsch global alignment of two sequences
Gap opening penalty [10.0]: 
Gap extension penalty [0.5]: 
Output alignment [hba_human.needle]: 

Go to the input files for this example
Go to the output files for this example

Command line arguments

Needleman-Wunsch global alignment of two sequences
Version: EMBOSS:6.3.0

   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.
                                  (Floating point number from 1.0 to 100.0)
   -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. (Floating
                                  point number from 0.0 to 10.0)
  [-outfile]           align      [*.needle] 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.
   -endweight          boolean    [N] Apply end gap penalties.
   -endopen            float      [10.0 for any sequence] The end gap open
                                  penalty is the score taken away when an end
                                  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.
                                  (Floating point number from 1.0 to 100.0)
   -endextend          float      [0.5 for any sequence] The end gap
                                  extension, penalty is added to the end gap
                                  penalty for each base or residue in the end
                                  gap. This is how long end gaps are
                                  penalized. (Floating point number from 0.0
                                  to 10.0)

   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 first file to standard output
   -filter             boolean    Read first file from standard input, write
                                  first file to 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 and exit. 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
   -version            boolean    Report version number and exit

Qualifier Type Description Allowed values Default
Standard (Mandatory) qualifiers
[-asequence]
(Parameter 1)
sequence Sequence filename and optional format, or reference (input USA) Readable sequence Required
[-bsequence]
(Parameter 2)
seqall Sequence(s) filename and optional format, or reference (input USA) Readable sequence(s) Required
-gapopen float 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. Floating point number from 1.0 to 100.0 10.0 for any sequence
-gapextend float 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. Floating point number from 0.0 to 10.0 0.5 for any sequence
[-outfile]
(Parameter 3)
align Output alignment file name Alignment output file <*>.needle
Additional (Optional) qualifiers
-datafile matrixf 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
-endweight boolean Apply end gap penalties. Boolean value Yes/No No
-endopen float The end gap open penalty is the score taken away when an end 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. Floating point number from 1.0 to 100.0 10.0 for any sequence
-endextend float The end gap extension, penalty is added to the end gap penalty for each base or residue in the end gap. This is how long end gaps are penalized. Floating point number from 0.0 to 10.0 0.5 for any sequence
Advanced (Unprompted) qualifiers
-[no]brief boolean Brief identity and similarity Boolean value Yes/No Yes
Associated qualifiers
"-asequence" associated sequence qualifiers
-sbegin1
-sbegin_asequence
integer Start of the sequence to be used Any integer value 0
-send1
-send_asequence
integer End of the sequence to be used Any integer value 0
-sreverse1
-sreverse_asequence
boolean Reverse (if DNA) Boolean value Yes/No N
-sask1
-sask_asequence
boolean Ask for begin/end/reverse Boolean value Yes/No N
-snucleotide1
-snucleotide_asequence
boolean Sequence is nucleotide Boolean value Yes/No N
-sprotein1
-sprotein_asequence
boolean Sequence is protein Boolean value Yes/No N
-slower1
-slower_asequence
boolean Make lower case Boolean value Yes/No N
-supper1
-supper_asequence
boolean Make upper case Boolean value Yes/No N
-sformat1
-sformat_asequence
string Input sequence format Any string  
-sdbname1
-sdbname_asequence
string Database name Any string  
-sid1
-sid_asequence
string Entryname Any string  
-ufo1
-ufo_asequence
string UFO features Any string  
-fformat1
-fformat_asequence
string Features format Any string  
-fopenfile1
-fopenfile_asequence
string Features file name Any string  
"-bsequence" associated seqall qualifiers
-sbegin2
-sbegin_bsequence
integer Start of each sequence to be used Any integer value 0
-send2
-send_bsequence
integer End of each sequence to be used Any integer value 0
-sreverse2
-sreverse_bsequence
boolean Reverse (if DNA) Boolean value Yes/No N
-sask2
-sask_bsequence
boolean Ask for begin/end/reverse Boolean value Yes/No N
-snucleotide2
-snucleotide_bsequence
boolean Sequence is nucleotide Boolean value Yes/No N
-sprotein2
-sprotein_bsequence
boolean Sequence is protein Boolean value Yes/No N
-slower2
-slower_bsequence
boolean Make lower case Boolean value Yes/No N
-supper2
-supper_bsequence
boolean Make upper case Boolean value Yes/No N
-sformat2
-sformat_bsequence
string Input sequence format Any string  
-sdbname2
-sdbname_bsequence
string Database name Any string  
-sid2
-sid_bsequence
string Entryname Any string  
-ufo2
-ufo_bsequence
string UFO features Any string  
-fformat2
-fformat_bsequence
string Features format Any string  
-fopenfile2
-fopenfile_bsequence
string Features file name Any string  
"-outfile" associated align qualifiers
-aformat3
-aformat_outfile
string Alignment format Any string srspair
-aextension3
-aextension_outfile
string File name extension Any string  
-adirectory3
-adirectory_outfile
string Output directory Any string  
-aname3
-aname_outfile
string Base file name Any string  
-awidth3
-awidth_outfile
integer Alignment width Any integer value 0
-aaccshow3
-aaccshow_outfile
boolean Show accession number in the header Boolean value Yes/No N
-adesshow3
-adesshow_outfile
boolean Show description in the header Boolean value Yes/No N
-ausashow3
-ausashow_outfile
boolean Show the full USA in the alignment Boolean value Yes/No N
-aglobal3
-aglobal_outfile
boolean Show the full sequence in alignment Boolean value Yes/No Y
General qualifiers
-auto boolean Turn off prompts Boolean value Yes/No N
-stdout boolean Write first file to standard output Boolean value Yes/No N
-filter boolean Read first file from standard input, write first file to standard output Boolean value Yes/No N
-options boolean Prompt for standard and additional values Boolean value Yes/No N
-debug boolean Write debug output to program.dbg Boolean value Yes/No N
-verbose boolean Report some/full command line options Boolean value Yes/No Y
-help boolean Report command line options and exit. More information on associated and general qualifiers can be found with -help -verbose Boolean value Yes/No N
-warning boolean Report warnings Boolean value Yes/No Y
-error boolean Report errors Boolean value Yes/No Y
-fatal boolean Report fatal errors Boolean value Yes/No Y
-die boolean Report dying program messages Boolean value Yes/No Y
-version boolean Report version number and exit Boolean value Yes/No N

Input file format

needle reads any 2 sequence USAs of the same type (DNA or protein).

Input files for usage example

'tsw:hba_human' is a sequence entry in the example protein database 'tsw'

Database entry: tsw:hba_human

ID   HBA_HUMAN               Reviewed;         142 AA.
AC   P69905; P01922; Q1HDT5; Q3MIF5; Q53F97; Q96KF1; Q9NYR7; Q9UCM0;
DT   21-JUL-1986, integrated into UniProtKB/Swiss-Prot.
DT   23-JAN-2007, sequence version 2.
DT   15-JUN-2010, entry version 86.
DE   RecName: Full=Hemoglobin subunit alpha;
DE   AltName: Full=Hemoglobin alpha chain;
DE   AltName: Full=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; DOI=10.1073/pnas.77.12.7054;
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; DOI=10.1073/pnas.78.8.5041;
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.";


  [Part of this file has been deleted for brevity]

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     137    137       L -> R (in Toyama).
FT                                /FTId=VAR_035242.
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/Ethiopia; O(2) affinity
FT                                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   CONFLICT     10     10       N -> H (in Ref. 13; BAD97112).
FT   HELIX         5     16
FT   HELIX        17     21
FT   HELIX        22     36
FT   HELIX        38     43
FT   HELIX        54     72
FT   HELIX        74     76
FT   HELIX        77     80
FT   HELIX        82     90
FT   HELIX        97    113
FT   TURN        115    117
FT   HELIX       120    137
FT   TURN        138    140
SQ   SEQUENCE   142 AA;  15258 MW;  15E13666573BBBAE CRC64;
     MVLSPADKTN VKAAWGKVGA HAGEYGAEAL ERMFLSFPTT KTYFPHFDLS HGSAQVKGHG
     KKVADALTNA VAHVDDMPNA LSALSDLHAH KLRVDPVNFK LLSHCLLVTL AAHLPAEFTP
     AVHASLDKFL ASVSTVLTSK YR
//

Database entry: tsw:hbb_human

ID   HBB_HUMAN               Reviewed;         147 AA.
AC   P68871; A4GX73; B2ZUE0; P02023; Q13852; Q14481; Q14510; Q45KT0;
AC   Q549N7; Q6FI08; Q6R7N2; Q8IZI1; Q9BX96; Q9UCD6; Q9UCP8; Q9UCP9;
DT   21-JUL-1986, integrated into UniProtKB/Swiss-Prot.
DT   23-JAN-2007, sequence version 2.
DT   15-JUN-2010, entry version 88.
DE   RecName: Full=Hemoglobin subunit beta;
DE   AltName: Full=Hemoglobin beta chain;
DE   AltName: Full=Beta-globin;
DE   Contains:
DE     RecName: Full=LVV-hemorphin-7;
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 [GENOMIC DNA].
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   [2]
RP   NUCLEOTIDE SEQUENCE [GENOMIC DNA].
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   [3]
RP   NUCLEOTIDE SEQUENCE [GENOMIC DNA], AND VARIANT LYS-7.
RX   PubMed=16175509; DOI=10.1086/491748;
RA   Wood E.T., Stover D.A., Slatkin M., Nachman M.W., Hammer M.F.;
RT   "The beta-globin recombinational hotspot reduces the effects of strong
RT   selection around HbC, a recently arisen mutation providing resistance
RT   to malaria.";
RL   Am. J. Hum. Genet. 77:637-642(2005).
RN   [4]
RP   NUCLEOTIDE SEQUENCE [GENOMIC DNA].
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   [5]
RP   NUCLEOTIDE SEQUENCE [GENOMIC DNA], AND VARIANT ARG-113.
RA   Cabeda J.M., Correia C., Estevinho A., Cardoso C., Amorim M.L.,
RA   Cleto E., Vale L., Coimbra E., Pinho L., Justica B.;
RT   "Unexpected patterns of globin mutations in thalassemia patients from
RT   north of Portugal.";


  [Part of this file has been deleted for brevity]

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   CONFLICT     26     26       Missing (in Ref. 15; ACD39349).
FT   CONFLICT     42     42       F -> L (in Ref. 13; AAR96398).
FT   HELIX         6     16
FT   TURN         21     23
FT   HELIX        24     35
FT   HELIX        37     42
FT   HELIX        44     46
FT   HELIX        52     57
FT   HELIX        59     77
FT   TURN         78     80
FT   HELIX        82     94
FT   TURN         95     97
FT   HELIX       102    119
FT   HELIX       120    122
FT   HELIX       125    142
FT   HELIX       144    146
SQ   SEQUENCE   147 AA;  15998 MW;  A31F6D621C6556A1 CRC64;
     MVHLTPEEKS AVTALWGKVN VDEVGGEALG RLLVVYPWTQ RFFESFGDLS TPDAVMGNPK
     VKAHGKKVLG AFSDGLAHLD NLKGTFATLS ELHCDKLHVD PENFRLLGNV LVCVLAHHFG
     KEFTPPVQAA YQKVVAGVAN ALAHKYH
//

Output file format

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.

Output files for usage example

File: hba_human.needle

########################################
# Program: needle
# Rundate: Fri 15 Jan 2010 12:00:00
# Commandline: needle
#    [-asequence] tsw:hba_human
#    [-bsequence] tsw:hbb_human
# Align_format: srspair
# Report_file: hba_human.needle
########################################

#=======================================
#
# Aligned_sequences: 2
# 1: HBA_HUMAN
# 2: HBB_HUMAN
# Matrix: EBLOSUM62
# Gap_penalty: 10.0
# Extend_penalty: 0.5
#
# Length: 149
# Identity:      65/149 (43.6%)
# Similarity:    90/149 (60.4%)
# Gaps:           9/149 ( 6.0%)
# Score: 292.5
# 
#
#=======================================

HBA_HUMAN          1 MV-LSPADKTNVKAAWGKVGAHAGEYGAEALERMFLSFPTTKTYFPHF-D     48
                     || |:|.:|:.|.|.||||  :..|.|.|||.|:.:.:|.|:.:|..| |
HBB_HUMAN          1 MVHLTPEEKSAVTALWGKV--NVDEVGGEALGRLLVVYPWTQRFFESFGD     48

HBA_HUMAN         49 LS-----HGSAQVKGHGKKVADALTNAVAHVDDMPNALSALSDLHAHKLR     93
                     ||     .|:.:||.|||||..|.::.:||:|::....:.||:||..||.
HBB_HUMAN         49 LSTPDAVMGNPKVKAHGKKVLGAFSDGLAHLDNLKGTFATLSELHCDKLH     98

HBA_HUMAN         94 VDPVNFKLLSHCLLVTLAAHLPAEFTPAVHASLDKFLASVSTVLTSKYR    142
                     |||.||:||.:.|:..||.|...||||.|.|:..|.:|.|:..|..||.
HBB_HUMAN         99 VDPENFRLLGNVLVCVLAHHFGKEFTPPVQAAYQKVVAGVANALAHKYH    147


#---------------------------------------
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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).

Data files

For protein sequences EBLOSUM62 is used for the substitution matrix. For nucleotide sequence, EDNAFULL is used. Others can be specified.

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:

Notes

needle is a true implementation of the Needleman-Wunsch algorithm and so produces a full path matrix. It therefore cannot be used with genome sized sequences unless you've a lot of memory and a lot of time.

References

  1. Needleman, S. B. and Wunsch, C. D. (1970) J. Mol. Biol. 48, 443-453.
  2. Kruskal, J. B. (1983) An overview of squence comparison In D. Sankoff and J. B. Kruskal, (ed.), Time warps, string edits and macromolecules: the theory and practice of sequence comparison, pp. 1-44 Addison Wesley.

Warnings

needle is for aligning two sequences over their entire length. This works best with closely related sequences. If you use needle to align very distantly-related sequences, it will produce a result but much of the alignment may have little or no biological significance.

A true Needleman Wunsch implementation like needle 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 stretcher instead.

Diagnostic Error Messages

Uncaught exception
 Assertion failed
 raised at ajmem.c:xxx

Probably means you have run out of memory. Try using stretcher if this happens.

Exit status

0 upon successful completion.

Known bugs

None.

See also

Program name Description
est2genome Align EST sequences to genomic DNA sequence
needleall Many-to-many pairwise alignments of two sequence sets
stretcher Needleman-Wunsch rapid global alignment of two sequences

When you want an alignment that covers the whole length of both sequences, use needle.

When you are trying to find the best region of similarity between two sequences, use water.

stretcher is a more suitable program to use to find global alignments of very long sequences.

Author(s)

Alan Bleasby
European Bioinformatics Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge CB10 1SD, UK

Please report all bugs to the EMBOSS bug team (emboss-bug © emboss.open-bio.org) not to the original author.

History

Completed 8th July 1999.

Modified 26th July 1999 - scoring tweaked.

Modified 22 Oct 2000 - %ID and %Similarity scores added.

Target users

This program is intended to be used by everyone and everything, from naive users to embedded scripts.

Comments

None