stretcher Wiki The master copies of EMBOSS documentation are available at http://emboss.open-bio.org/wiki/Appdocs on the EMBOSS Wiki. Please help by correcting and extending the Wiki pages. Function Needleman-Wunsch rapid global alignment of two sequences Description stretcher calculates an optimal global alignment of two sequences using a modification of the classic dynamic programming algorithm which uses linear space. The output is a standard alignment file. The substitution matrix, gap insertion penalty and gap extension penalties used to calculate the alignment may be specified. Algorithm The standard sequence global alignment program using the Needleman & Wunsch algorithm, as implemented in the program needle, requires O(MN) space. This is standard computer-science language for it needing an amount of computer memory that is proportional to the product of the two sequences being aligned. So if a 1 kb and a 10 kb sequence take 10 Mega-words of memory to align, you should expect that in order to align a 10 kb sequence and a 1 Mb sequence you will need approximately 10 Giga-words of memory. When using needle computer memory will rapidly be exhausted as the size of the aligned sequences increases. This program implements the Myers and Miller algorithm for finding an optimal global alignment in an amount of computer memory that is only proportional to the size of the smaller sequence - O(N). Usage Here is a sample session with stretcher % stretcher tsw:hba_human tsw:hbb_human Needleman-Wunsch rapid global alignment of two sequences Output alignment [hba_human.stretcher]: Go to the input files for this example Go to the output files for this example Command line arguments Needleman-Wunsch rapid global alignment of two sequences Version: EMBOSS:6.4.0.0 Standard (Mandatory) qualifiers: [-asequence] sequence Sequence filename and optional format, or reference (input USA) [-bsequence] sequence Sequence filename and optional format, or reference (input USA) [-outfile] align [*.stretcher] Output alignment file name (default -aformat markx0) Additional (Optional) qualifiers: -datafile matrix [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. -gapopen integer [12 for protein, 16 for nucleic] Gap penalty (Positive integer) -gapextend integer [2 for protein, 4 for nucleic] Gap length penalty (Positive integer) Advanced (Unprompted) qualifiers: (none) 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 the sequence to be used -send2 integer End of the 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 Input file format stretcher reads any 2 nucleotide or protein sequences. The input is a standard EMBOSS sequence query (also known as a 'USA'). Major sequence database sources defined as standard in EMBOSS installations include srs:embl, srs:uniprot and ensembl Data can also be read from sequence output in any supported format written by an EMBOSS or third-party application. The input format can be specified by using the command-line qualifier -sformat xxx, where 'xxx' is replaced by the name of the required format. The available format names are: gff (gff3), gff2, embl (em), genbank (gb, refseq), ddbj, refseqp, pir (nbrf), swissprot (swiss, sw), dasgff and debug. See: http://emboss.sf.net/docs/themes/SequenceFormats.html for further information on sequence formats. 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: multiple, simple, fasta, msf, clustal, mega, meganon, nexus,, nexusnon, phylip, phylipnon, selex, treecon, tcoffee, debug, srs. The available pairwise alignment format names are: pair, markx0, markx1, markx2, markx3, markx10, match, sam, bam, score, srspair See: http://emboss.sf.net/docs/themes/AlignFormats.html for further information on alignment formats. The default output format is 'markx0'. Output files for usage example File: hba_human.stretcher ######################################## # Program: stretcher # Rundate: Fri 15 Jul 2011 12:00:00 # Commandline: stretcher # [-asequence] tsw:hba_human # [-bsequence] tsw:hbb_human # Align_format: markx0 # Report_file: hba_human.stretcher ######################################## #======================================= # # Aligned_sequences: 2 # 1: HBA_HUMAN # 2: HBB_HUMAN # Matrix: EBLOSUM62 # Gap_penalty: 12 # Extend_penalty: 2 # # Length: 149 # Identity: 65/149 (43.6%) # Similarity: 90/149 (60.4%) # Gaps: 9/149 ( 6.0%) # Score: 277 # # #======================================= 10 20 30 40 HBA_HU MV-LSPADKTNVKAAWGKVGAHAGEYGAEALERMFLSFPTTKTYFPHF-D :: :.: .:. : : :::: . : : ::: :. . .: :. .: : : HBB_HU MVHLTPEEKSAVTALWGKV--NVDEVGGEALGRLLVVYPWTQRFFESFGD 10 20 30 40 50 60 70 80 90 HBA_HU LSH-----GSAQVKGHGKKVADALTNAVAHVDDMPNALSALSDLHAHKLR :: :. .:: ::::: : .. .::.:.. . ::.:: :: HBB_HU LSTPDAVMGNPKVKAHGKKVLGAFSDGLAHLDNLKGTFATLSELHCDKLH 50 60 70 80 90 100 110 120 130 140 HBA_HU VDPVNFKLLSHCLLVTLAAHLPAEFTPAVHASLDKFLASVSTVLTSKYR ::: ::.:: . :. :: : :::: : :. : .: :. : :: HBB_HU VDPENFRLLGNVLVCVLAHHFGKEFTPPVQAAYQKVVAGVANALAHKYH 100 110 120 130 140 #--------------------------------------- #--------------------------------------- Data files For protein sequences EBLOSUM62 is used for the substitution matrix. For nucleotide sequence, EDNAMAT 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 EMBOSS environment variable EMBOSS_DATA. 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: * . (your current directory) * .embossdata (under your current directory) * ~/ (your home directory) * ~/.embossdata Notes A global pairwise alignment is one where it is assumed that the two sequences have diverged from a common ancestor and that the program should try to stretch the two sequences, introducing gaps where necessary, in order to show the alignment over the whole length of the two sequences that best illustrates their similarities. In contrast, a local alignment program like matcher simply finds local, small parts of the two sequences where there is some similarity and makes no assumption about the whole length of the sequence needing to be similar. References 1. E. Myers and W. Miller, "Optimal Alignments in Linear Space," CABIOS 4, 1 (1988), 11-17. Warnings Demonstration of similarity is not evidence of homology! This program will produce a global alignment even if there is no biological justification for thinking that there might be a common ancestor. Diagnostic Error Messages None. Exit status It exits with a status of 0. Known bugs None. See also Program name Description est2genome Align EST sequences to genomic DNA sequence needle Needleman-Wunsch global alignment of two sequences needleall Many-to-many pairwise alignments of two sequence sets Author(s) The original program was written by Gene Myers and Webb Miller in 1989. This application was modified for inclusion in EMBOSS by Ian Longden formerly at: Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge, CB10 1SA, UK. Please report all bugs to the EMBOSS bug team (emboss-bug (c) emboss.open-bio.org) not to the original author. History Completed 13th May 1999. Target users This program is intended to be used by everyone and everything, from naive users to embedded scripts. Comments None