merger |
It uses a global alignment algorithm (Needleman & Wunsch) to optimally align the sequences and then it creates the merged sequence from the alignment. When there is a mismatch in the alignment between the two sequences, the correct base to include in the resulting sequence is chosen by using the base from the sequence which has the best local sequence quality score. The following heuristic is used to find the sequence quality score:
If one of the bases is a 'N', then the other sequence's base is used, else:
A window size around the disputed base is used to find the local quality score. This window size is increased from 5, to 10 to 20 bases or until there is a clear decision on the best choice. If there is no best choice after using a window of 20, then the base in the first sequence is used.
To calculate the quality of a window of a sequence around a base:
N.B. This heavily discriminates against the iffy bits at the end of sequence reads.
This program was originally written to aid in the reconstruction of mRNA sequences which had been sequenced from both ends as a 5' and 3' EST (cDNA). eg. joining two reads produced by primer walking sequencing.
Care should be taken to reverse one of the sequences (e.g. using the qualifier '-sreverse2') if this is required to get them both in the correct orientation.
Because it uses a Needleman & Wunsch alignment the required memory may be greater than the available memory when attempting to merge large (cosmid-sized or greater) sequences.
The gap open and gap extension penalties have been set at a higher level than is usual (50 and 5). This was experimentally determined to give the best results with a set of poor quality EST test sequences.
% merger Merge two overlapping sequences Input sequence: tembl:v00295 Second sequence: tembl:x51872 Output alignment [v00295.merger]: output sequence [v00295.fasta]: |
Go to the input files for this example
Go to the output files for this example
Typically, one of the sequences will need to be reverse-complemented to put it into the correct orientation to make it join. For example:
% merger file1.seq file2.seq -sreverse2 -outseq merged.seq
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 [*.merger] Output alignment file name [-outseq] seqout [ |
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 filename and optional format, or reference (input USA) | Readable sequence | Required |
[-outfile] (Parameter 3) |
Output alignment file name | Alignment output file | <*>.merger |
[-outseq] (Parameter 4) |
Sequence filename and optional format (output USA) | Writeable sequence | <*>.format |
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 |
-gapopen | Gap opening penalty | Number from 0.000 to 100.000 | @($(acdprotein)? 50.0 : 50.0 ) |
-gapextend | Gap extension penalty | Number from 0.000 to 10.000 | @($(acdprotein)? 5.0 : 5.0) |
Advanced (Unprompted) qualifiers | Allowed values | Default | |
(none) |
ID V00295; SV 1; linear; genomic DNA; STD; PRO; 1500 BP. XX AC V00295; XX DT 09-JUN-1982 (Rel. 01, Created) DT 07-JUL-1995 (Rel. 44, Last updated, Version 4) XX DE E. coli lacY gene (codes for lactose permease). XX KW membrane protein. XX OS Escherichia coli OC Bacteria; Proteobacteria; Gammaproteobacteria; Enterobacteriales; OC Enterobacteriaceae; Escherichia. XX RN [1] RP 1-1500 RX DOI; 10.1038/283541a0. RX PUBMED; 6444453. RA Buechel D.E., Gronenborn B., Mueller-Hill B.; RT "Sequence of the lactose permease gene"; RL Nature 283(5747):541-545(1980). XX CC lacZ is a beta-galactosidase and lacA is transacetylase. CC KST ECO.LACY XX FH Key Location/Qualifiers FH FT source 1..1500 FT /organism="Escherichia coli" FT /mol_type="genomic DNA" FT /db_xref="taxon:562" FT CDS <1..54 FT /codon_start=1 FT /transl_table=11 FT /note="reading frame (lacZ)" FT /db_xref="GOA:P00722" FT /db_xref="PDB:1BGL" FT /db_xref="PDB:1BGM" FT /db_xref="PDB:1DP0" FT /db_xref="PDB:1F49" FT /db_xref="PDB:1F4A" FT /db_xref="PDB:1F4H" FT /db_xref="PDB:1GHO" FT /db_xref="PDB:1HN1" FT /db_xref="PDB:1JYN" FT /db_xref="PDB:1JYV" FT /db_xref="PDB:1JYW" FT /db_xref="PDB:1JYX" FT /db_xref="PDB:1JYY" [Part of this file has been deleted for brevity] FT /db_xref="PDB:2CFP" FT /db_xref="PDB:2CFQ" FT /db_xref="UniProtKB/Swiss-Prot:P02920" FT /protein_id="CAA23571.1" FT /translation="MYYLKNTNFWMFGLFFFFYFFIMGAYFPFFPIWLHDINHISKSDT FT GIIFAAISLFSLLFQPLFGLLSDKLGLRKYLLWIITGMLVMFAPFFIFIFGPLLQYNIL FT VGSIVGGIYLGFCFNAGAPAVEAFIEKVSRRSNFEFGRARMFGCVGWALCASIVGIMFT FT INNQFVFWLGSGCALILAVLLFFAKTDAPSSATVANAVGANHSAFSLKLALELFRQPKL FT WFLSLYVIGVSCTYDVFDQQFANFFTSFFATGEQGTRVFGYVTTMGELLNASIMFFAPL FT IINRIGGKNALLLAGTIMSVRIIGSSFATSALEVVILKTLHMFEVPFLLVGCFKYITSQ FT FEVRFSATIYLVCFCFFKQLAMIFMSVLAGNMYESIGFQGAYLVLGLVALGFTLISVFT FT LSGPGPLSLLRRQVNEVA" FT CDS 1423..>1500 FT /transl_table=11 FT /note="reading frame (lacA)" FT /db_xref="GOA:P07464" FT /db_xref="PDB:1KQA" FT /db_xref="PDB:1KRR" FT /db_xref="PDB:1KRU" FT /db_xref="PDB:1KRV" FT /db_xref="UniProtKB/Swiss-Prot:P07464" FT /protein_id="CAA23572.1" FT /translation="MNMPMTERIRAGKLFTDMCEGLPEKR" XX SQ Sequence 1500 BP; 315 A; 342 C; 357 G; 486 T; 0 other; ttccagctga gcgccggtcg ctaccattac cagttggtct ggtgtcaaaa ataataataa 60 ccgggcaggc catgtctgcc cgtatttcgc gtaaggaaat ccattatgta ctatttaaaa 120 aacacaaact tttggatgtt cggtttattc tttttctttt acttttttat catgggagcc 180 tacttcccgt ttttcccgat ttggctacat gacatcaacc atatcagcaa aagtgatacg 240 ggtattattt ttgccgctat ttctctgttc tcgctattat tccaaccgct gtttggtctg 300 ctttctgaca aactcgggct gcgcaaatac ctgctgtgga ttattaccgg catgttagtg 360 atgtttgcgc cgttctttat ttttatcttc gggccactgt tacaatacaa cattttagta 420 ggatcgattg ttggtggtat ttatctaggc ttttgtttta acgccggtgc gccagcagta 480 gaggcattta ttgagaaagt cagccgtcgc agtaatttcg aatttggtcg cgcgcggatg 540 tttggctgtg ttggctgggc gctgtgtgcc tcgattgtcg gcatcatgtt caccatcaat 600 aatcagtttg ttttctggct gggctctggc tgtgcactca tcctcgccgt tttactcttt 660 ttcgccaaaa cggatgcgcc ctcttctgcc acggttgcca atgcggtagg tgccaaccat 720 tcggcattta gccttaagct ggcactggaa ctgttcagac agccaaaact gtggtttttg 780 tcactgtatg ttattggcgt ttcctgcacc tacgatgttt ttgaccaaca gtttgctaat 840 ttctttactt cgttctttgc taccggtgaa cagggtacgc gggtatttgg ctacgtaacg 900 acaatgggcg aattacttaa cgcctcgatt atgttctttg cgccactgat cattaatcgc 960 atcggtggga aaaacgccct gctgctggct ggcactatta tgtctgtacg tattattggc 1020 tcatcgttcg ccacctcagc gctggaagtg gttattctga aaacgctgca tatgtttgaa 1080 gtaccgttcc tgctggtggg ctgctttaaa tatattacca gccagtttga agtgcgtttt 1140 tcagcgacga tttatctggt ctgtttctgc ttctttaagc aactggcgat gatttttatg 1200 tctgtactgg cgggcaatat gtatgaaagc atcggtttcc agggcgctta tctggtgctg 1260 ggtctggtgg cgctgggctt caccttaatt tccgtgttca cgcttagcgg ccccggcccg 1320 ctttccctgc tgcgtcgtca ggtgaatgaa gtcgcttaag caatcaatgt cggatgcggc 1380 gcgacgctta tccgaccaac atatcataac ggagtgatcg cattgaacat gccaatgacc 1440 gaaagaataa gagcaggcaa gctatttacc gatatgtgcg aaggcttacc ggaaaaaaga 1500 // |
ID X51872; SV 1; linear; genomic DNA; STD; PRO; 1832 BP. XX AC X51872; XX DT 17-APR-1990 (Rel. 23, Created) DT 05-JUL-1999 (Rel. 60, Last updated, Version 5) XX DE Escherichia coli lacA gene for thiogalactoside transacetylase XX KW lac operon; lacA gene; lacY gene; thiogalactoside transacetylase. XX OS Escherichia coli OC Bacteria; Proteobacteria; Gammaproteobacteria; Enterobacteriales; OC Enterobacteriaceae; Escherichia. XX RN [1] RC (1-1832) RP 1-1832 RX PUBMED; 3901000. RA Hediger M.A, Johnson D.F., Nierlich D.P., Zabin I.; RT "DNA sequence of the lactose operon: the lacA gene and the transcriptional RT termination region."; RL Proc. Natl. Acad. Sci. U.S.A. 82(19):6414-6418(1985). XX FH Key Location/Qualifiers FH FT source 1..1832 FT /organism="Escherichia coli" FT /mol_type="genomic DNA" FT /db_xref="taxon:562" FT CDS <1..18 FT /codon_start=1 FT /transl_table=11 FT /product="lacY gene product" FT /protein_id="CAA36161.1" FT /translation="VNEVA" FT CDS 82..693 FT /transl_table=11 FT /gene="lacA" FT /product="thiogalactoside transacetylase" FT /db_xref="GOA:P07464" FT /db_xref="InterPro:IPR001451" FT /db_xref="InterPro:IPR011004" FT /db_xref="PDB:1KQA" FT /db_xref="PDB:1KRR" FT /db_xref="PDB:1KRU" FT /db_xref="PDB:1KRV" FT /db_xref="UniProtKB/Swiss-Prot:P07464" FT /protein_id="CAA36162.1" FT /translation="MNMPMTERIRAGKLFTDMCEGLPEKRLRGKTLMYEFNHSHPSEVE FT KRESLIKEMFATVGENAWVEPPVYFSYGSNIHIGRNFYANFNLTIVDDYTVTIGDNVLI FT APNVTLSVTGHPVHHELRKNGEMYSFPITIGNNVWIGSHVVINPGVTIGDNSVIGAGSI FT VTKDIPPNVVAAGVPCRVIREINDRDKHYYFKDYKVESSV" XX SQ Sequence 1832 BP; 519 A; 510 C; 450 G; 353 T; 0 other; gtgaatgaag tcgcttaagc aatcaatgtc ggatgcggcg cgacgcttat ccgaccaaca 60 tatcataacg gagtgatcgc attgaacatg ccaatgaccg aaagaataag agcaggcaag 120 ctatttaccg atatgtgcga aggcttaccg gaaaaaagac ttcgtgggaa aacgttaatg 180 tatgagttta atcactcgca tccatcagaa gttgaaaaaa gagaaagcct gattaaagaa 240 atgtttgcca cggtagggga aaacgcctgg gtagaaccgc ctgtctattt ctcttacggt 300 tccaacatcc atataggccg caatttttat gcaaatttca atttaaccat tgtcgatgac 360 tacacggtaa caatcggtga taacgtactg attgcaccca acgttactct ttccgttacg 420 ggacaccctg tacaccatga attgagaaaa aacggcgaga tgtactcttt tccgataacg 480 attggcaata acgtctggat cggaagtcat gtggttatta atccaggcgt caccatcggg 540 gataattctg ttattggcgc gggtagtatc gtcacaaaag acattccacc aaacgtcgtg 600 gcggctggcg ttccttgtcg ggttattcgc gaaataaacg accgggataa gcactattat 660 ttcaaagatt ataaagttga atcgtcagtt taaattataa aaattgcctg atacgctgcg 720 cttatcaggc ctacaagttc agcgatctac attagccgca tccggcatga acaaagcgca 780 ggaacaagcg tcgcatcatg cctctttgac ccacagctgc ggaaaacgta ctggtgcaaa 840 acgcagggtt atgatcatca gcccaacgac gcacagcgca tgaaatgccc agtccatcag 900 gtaattgccg ctgatactac gcagcacgcc agaaaaccac ggggcaagcc cggcgatgat 960 aaaaccgatt ccctgcataa acgccaccag cttgccagca atagccggtt gcacagagtg 1020 atcgagcgcc agcagcaaac agagcggaaa cgcgccgccc agacctaacc cacacaccat 1080 cgcccacaat accggcaatt gcatcggcag ccagataaag ccgcagaacc ccaccagttg 1140 taacaccagc gccagcatta acagtttgcg ccgatcctga tggcgagcca tagcaggcat 1200 cagcaaagct cctgcggctt gcccaagcgt catcaatgcc agtaaggaac cgctgtactg 1260 cgcgctggca ccaatctcaa tatagaaagc gggtaaccag gcaatcaggc tggcgtaacc 1320 gccgttaatc agaccgaagt aaacacccag cgtccacgcg cggggagtga ataccacgcg 1380 aaccggagtg gttgttgtct tgtgggaaga ggcgacctcg cgggcgcttt gccaccacca 1440 ggcaaagagc gcaacaacgg caggcagcgc caccaggcga gtgtttgata ccaggtttcg 1500 ctatgttgaa ctaaccaggg cgttatggcg gcaccaagcc caccgccgcc catcagagcc 1560 gcggaccaca gccccatcac cagtggcgtg cgctgctgaa accgccgttt aatcaccgaa 1620 gcatcaccgc ctgaatgatg ccgatcccca ccccaccaag cagtgcgctg ctaagcagca 1680 gcgcactttg cgggtaaagc tcacgcatca atgcaccgac ggcaatcagc aacagactga 1740 tggcgacact gcgacgttcg ctgacatgct gatgaagcca gcttccggcc agcgccagcc 1800 cgcccatggt aaccaccggc agagcggtcg ac 1832 // |
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.
The output report file contains descriptions of the positions where there is a mismatch in the alignment and shows the alignment. Where there is a mismatch in the alignment, the chosen base is written in uppercase.
######################################## # Program: merger # Rundate: Sun 15 Jul 2007 12:00:00 # Commandline: merger # -asequence tembl:v00295 # -bsequence tembl:x51872 # Align_format: simple # Report_file: v00295.merger ######################################## #======================================= # # Aligned_sequences: 2 # 1: V00295 # 2: X51872 # Matrix: EDNAFULL # Gap_penalty: 50.0 # Extend_penalty: 5.0 # # Length: 3173 # Identity: 159/3173 ( 5.0%) # Similarity: 159/3173 ( 5.0%) # Gaps: 3014/3173 (95.0%) # Score: 795.0 # # #======================================= V00295 1 ttccagctgagcgccggtcgctaccattaccagttggtctggtgtcaaaa 50 X51872 1 -------------------------------------------------- 0 V00295 51 ataataataaccgggcaggccatgtctgcccgtatttcgcgtaaggaaat 100 X51872 1 -------------------------------------------------- 0 V00295 101 ccattatgtactatttaaaaaacacaaacttttggatgttcggtttattc 150 X51872 1 -------------------------------------------------- 0 V00295 151 tttttcttttacttttttatcatgggagcctacttcccgtttttcccgat 200 X51872 1 -------------------------------------------------- 0 V00295 201 ttggctacatgacatcaaccatatcagcaaaagtgatacgggtattattt 250 X51872 1 -------------------------------------------------- 0 V00295 251 ttgccgctatttctctgttctcgctattattccaaccgctgtttggtctg 300 [Part of this file has been deleted for brevity] X51872 1310 ctggcgtaaccgccgttaatcagaccgaagtaaacacccagcgtccacgc 1359 V00295 1501 -------------------------------------------------- 1500 X51872 1360 gcggggagtgaataccacgcgaaccggagtggttgttgtcttgtgggaag 1409 V00295 1501 -------------------------------------------------- 1500 X51872 1410 aggcgacctcgcgggcgctttgccaccaccaggcaaagagcgcaacaacg 1459 V00295 1501 -------------------------------------------------- 1500 X51872 1460 gcaggcagcgccaccaggcgagtgtttgataccaggtttcgctatgttga 1509 V00295 1501 -------------------------------------------------- 1500 X51872 1510 actaaccagggcgttatggcggcaccaagcccaccgccgcccatcagagc 1559 V00295 1501 -------------------------------------------------- 1500 X51872 1560 cgcggaccacagccccatcaccagtggcgtgcgctgctgaaaccgccgtt 1609 V00295 1501 -------------------------------------------------- 1500 X51872 1610 taatcaccgaagcatcaccgcctgaatgatgccgatccccaccccaccaa 1659 V00295 1501 -------------------------------------------------- 1500 X51872 1660 gcagtgcgctgctaagcagcagcgcactttgcgggtaaagctcacgcatc 1709 V00295 1501 -------------------------------------------------- 1500 X51872 1710 aatgcaccgacggcaatcagcaacagactgatggcgacactgcgacgttc 1759 V00295 1501 -------------------------------------------------- 1500 X51872 1760 gctgacatgctgatgaagccagcttccggccagcgccagcccgcccatgg 1809 V00295 1501 ----------------------- 1500 X51872 1810 taaccaccggcagagcggtcgac 1832 #--------------------------------------- # # Conflicts: V00295 X51872 # position base position base Using # # #--------------------------------------- |
>V00295 V00295.1 E. coli lacY gene (codes for lactose permease). ttccagctgagcgccggtcgctaccattaccagttggtctggtgtcaaaaataataataa ccgggcaggccatgtctgcccgtatttcgcgtaaggaaatccattatgtactatttaaaa aacacaaacttttggatgttcggtttattctttttcttttacttttttatcatgggagcc tacttcccgtttttcccgatttggctacatgacatcaaccatatcagcaaaagtgatacg ggtattatttttgccgctatttctctgttctcgctattattccaaccgctgtttggtctg ctttctgacaaactcgggctgcgcaaatacctgctgtggattattaccggcatgttagtg atgtttgcgccgttctttatttttatcttcgggccactgttacaatacaacattttagta ggatcgattgttggtggtatttatctaggcttttgttttaacgccggtgcgccagcagta gaggcatttattgagaaagtcagccgtcgcagtaatttcgaatttggtcgcgcgcggatg tttggctgtgttggctgggcgctgtgtgcctcgattgtcggcatcatgttcaccatcaat aatcagtttgttttctggctgggctctggctgtgcactcatcctcgccgttttactcttt ttcgccaaaacggatgcgccctcttctgccacggttgccaatgcggtaggtgccaaccat tcggcatttagccttaagctggcactggaactgttcagacagccaaaactgtggtttttg tcactgtatgttattggcgtttcctgcacctacgatgtttttgaccaacagtttgctaat ttctttacttcgttctttgctaccggtgaacagggtacgcgggtatttggctacgtaacg acaatgggcgaattacttaacgcctcgattatgttctttgcgccactgatcattaatcgc atcggtgggaaaaacgccctgctgctggctggcactattatgtctgtacgtattattggc tcatcgttcgccacctcagcgctggaagtggttattctgaaaacgctgcatatgtttgaa gtaccgttcctgctggtgggctgctttaaatatattaccagccagtttgaagtgcgtttt tcagcgacgatttatctggtctgtttctgcttctttaagcaactggcgatgatttttatg tctgtactggcgggcaatatgtatgaaagcatcggtttccagggcgcttatctggtgctg ggtctggtggcgctgggcttcaccttaatttccgtgttcacgcttagcggccccggcccg ctttccctgctgcgtcgtcaggtgaatgaagtcgcttaagcaatcaatgtcggatgcggc gcgacgcttatccgaccaacatatcataacggagtgatcgcattgaacatgccaatgacc gaaagaataagagcaggcaagctatttaccgatatgtgcgaaggcttaccggaaaaaaga cttcgtgggaaaacgttaatgtatgagtttaatcactcgcatccatcagaagttgaaaaa agagaaagcctgattaaagaaatgtttgccacggtaggggaaaacgcctgggtagaaccg cctgtctatttctcttacggttccaacatccatataggccgcaatttttatgcaaatttc aatttaaccattgtcgatgactacacggtaacaatcggtgataacgtactgattgcaccc aacgttactctttccgttacgggacaccctgtacaccatgaattgagaaaaaacggcgag atgtactcttttccgataacgattggcaataacgtctggatcggaagtcatgtggttatt aatccaggcgtcaccatcggggataattctgttattggcgcgggtagtatcgtcacaaaa gacattccaccaaacgtcgtggcggctggcgttccttgtcgggttattcgcgaaataaac gaccgggataagcactattatttcaaagattataaagttgaatcgtcagtttaaattata aaaattgcctgatacgctgcgcttatcaggcctacaagttcagcgatctacattagccgc atccggcatgaacaaagcgcaggaacaagcgtcgcatcatgcctctttgacccacagctg cggaaaacgtactggtgcaaaacgcagggttatgatcatcagcccaacgacgcacagcgc atgaaatgcccagtccatcaggtaattgccgctgatactacgcagcacgccagaaaacca cggggcaagcccggcgatgataaaaccgattccctgcataaacgccaccagcttgccagc aatagccggttgcacagagtgatcgagcgccagcagcaaacagagcggaaacgcgccgcc cagacctaacccacacaccatcgcccacaataccggcaattgcatcggcagccagataaa gccgcagaaccccaccagttgtaacaccagcgccagcattaacagtttgcgccgatcctg atggcgagccatagcaggcatcagcaaagctcctgcggcttgcccaagcgtcatcaatgc cagtaaggaaccgctgtactgcgcgctggcaccaatctcaatatagaaagcgggtaacca ggcaatcaggctggcgtaaccgccgttaatcagaccgaagtaaacacccagcgtccacgc gcggggagtgaataccacgcgaaccggagtggttgttgtcttgtgggaagaggcgacctc gcgggcgctttgccaccaccaggcaaagagcgcaacaacggcaggcagcgccaccaggcg agtgtttgataccaggtttcgctatgttgaactaaccagggcgttatggcggcaccaagc ccaccgccgcccatcagagccgcggaccacagccccatcaccagtggcgtgcgctgctga aaccgccgtttaatcaccgaagcatcaccgcctgaatgatgccgatccccaccccaccaa gcagtgcgctgctaagcagcagcgcactttgcgggtaaagctcacgcatcaatgcaccga cggcaatcagcaacagactgatggcgacactgcgacgttcgctgacatgctgatgaagcc agcttccggccagcgccagcccgcccatggtaaccaccggcagagcggtcgac |
Program name | Description |
---|---|
cons | Creates a consensus from multiple alignments |
megamerger | Merge two large overlapping nucleic acid sequences |