Quick start

Paste in or upload DNA sequences and hit "Submit query". The RevTrans server will then virtually translate the DNA sequences and align the resulting peptide sequences using MAFFT with default settings (other alignment program can be selected). Finally RevTrans constructs a multiple DNA alignment using the peptide alignment as a scaffold.

If you want more control over the alignment process RevTrans also accepts user provided peptide alignments. This will give you the opportunity to use your preferred alignment software and to optimize the parameters. If you need to translate your DNA sequences prior to alignment this can be done by using the "Translate only" button (or by following the link to the "Virtual Ribosome" server if you want more fine.grained control over the translation process). The translation has full support for degenerate nucleotides and alternative translation tables can be selected.

When providing your own peptide alignment, RevTrans will accept arbitrarily large input files.

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Detailed instructions

Supply DNA sequences

The DNA sequences must either be pasted into the webpage or uploaded via the "Choose file" button. The input must be in FASTA, MSF, or ALN (Clustal) format.

The full IUPAC degenerated DNA alphabet (not case sensitive) is supported:

A C G T R Y M K W S B D H V N

Please note that gaps and unknown symbolse.g. - and X will be discarded before processing.

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Optional: Supply aligned peptide sequences

For greater control of the alignment process you have the option of also supplying a pre-computed peptide alignment. RevTrans will then use this as the scaffold for the DNA alignment. The peptide alignment must be in FASTA, MSF, or ALN (Clustal) format.

By default "-", "." and "~" will be interpreted as gap symbols.

If a peptide alignment is not supplied the RevTrans web-server will automatically construct one using the selected multiple alignment program (deafult: MAFFT). In all case the alignment program will be run with default parameters.

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Submit query

Click on the "Submit query" button. If the processing of the query takes more than a few seconds you'll will get the option of supplying your email address and be notified when the job is done.

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Advanced options

RevTrans has support for a number of advanced options. Typically it is not necessary to set these manually and most users can safely skip this section and proceed to submitting the query.

• Data format, DNA sequences:
  By default the DNA file format is automatically detected. Alternatively you may specify the format as being FASTA, MSF, or ALN (Clustal).

• Data format, aligned peptide sequences:
  By default the peptide alignment file format is automatically detected. Alternatively you may specify the format as being FASTA, MSF, or ALN (Clustal).

• Output format:
  By default the final multiple DNA alignment will be in ALN (Clustal) format. Alternatively you may specify FASTA or MSF.

• Gap-In:
  Here you can specify which symbol(s) denote(s) a gap in user provided peptide alignemnt. The default should be correct for virtually all standard alignment files.

• Gap-out:
  Here you can specify which gap symbol to use in the output.

• Match DNA and peptide sequences by:
  This option gives the user control over how DNA sequences paired to their peptide counterpart.

  • Translation:
    (Default) The DNA sequences are translated using the standard genetic code (or an alternative translation table if selected below) with full IUPAC support and compared to the peptide sequences. The DNA sequence is paired with the first matching peptide sequence found.

  • Name:
    DNA sequences are paired with peptide counterparts based on sequence entry names. Entry names must be unique within files and identical across files. If you experience trouble when using name based matching, please make sure that sequences names do match across files as some alignment software may truncate or otherwise alter sequence names.

  • Position:
    DNA sequences are paired with peptide counterparts simply based on their order of appearance in the files.

• Translation table:
  Select an alternative translation table - used for "matching-by-translation" and with the "Translate Only" functionality.

  The numbering of the translation table is the one defined by the NCBI Taxonomy Group. For a detailed description of each genetic code, please consult the following web page at NCBI: The Genetic Codes . [Main site: Taxonomy]

• Alignment method
  (New in RevTrans 1.4) RevTrans offers a selection of programs for performing the peptide alignment step:

  • Dialign 2.2:
    
    Reference:
    B. Morgenstern (1999).
    DIALIGN 2: improvement of the segment-to-segment approach to multiple sequence alignment.
    Bioinformatics 15, 211 - 218.

  • Dialign-T 0.1.3:
    
    Reference:
    Amarendran R. Subramanian, Jan Weyer-Menkhoff, Michael Kaufmann, Burkhard Morgenstern:
    DIALIGN-T: An improved algorithm for segment-based multiple sequence alignment
    Bioinformatics 2005, 6:66.

  • ClustalW 1.83:
    
    Reference:
    Higgins D., Thompson J., Gibson T. Thompson J. D., Higgins D. G., Gibson T. J.(1994).
    CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting,position-specific gap penalties and weight matrix choice.
    Nucleic Acids Res. 22:4673-4680.

  Example data

  Sample DNA dataset

  The following is a set of unaligned Alpha-globin genes from a range for organisms.

  >Sheep
  ATGGCCCTGTGGACACGCCTGGTGCCCCTGCTGGCCCTGCTGGCACTCTGGGCCCCCGCC
  CCGGCCCACGCCTTCGTCAACCAGCACCTGTGCGGCTCCCACCTGGTGGAGGCGCTGTAC
  CTGGTGTGCGGAGAGCGCGGCTTCTTCTACACGCCCAAGGCCCGCCGGGAGGTGGAGGGC
  CCCCAGGTGGGGGCGCTGGAGCTGGCCGGAGGCCCCGGCGCGGGTGGCCTGGAGGGGCCC
  CCGCAGAAGCGTGGCATCGTGGAGCAGTGCTGCGCCGGCGTCTGCTCTCTCTACCAGCTG
  GAGAACTACTGTAACTAG
  >Pig
  ATGGCCCTGTGGACGCGCCTCCTGCCCCTGCTGGCCCTGCTGGCCCTCTGGGCGCCCGCC
  CCGGCCCAGGCCTTCGTGAACCAGCACCTGTGCGGCTCCCACCTGGTGGAGGCGCTGTAC
  CTGGTGTGCGGGGAGCGCGGCTTCTTCTACACGCCCAAGGCCCGTCGGGAGGCGGAGAAC
  CCTCAGGCAGGTGCCGTGGAGCTGGGCGGAGGCCTGGGCGGCCTGCAGGCCCTGGCGCTG
  GAGGGGCCCCCGCAGAAGCGTGGCATCGTGGAGCAGTGCTGCACCAGCATCTGTTCCCTC
  TACCAGCTGGAGAACTACTGCAACTAG
  >Dog
  ATGGCCCTCTGGATGCGCCTCCTGCCCCTGCTGGCCCTGCTGGCCCTCTGGGCGCCCGCG
  CCCACCCGAGCCTTCGTTAACCAGCACCTGTGTGGCTCCCACCTGGTAGAGGCTCTGTAC
  CTGGTGTGCGGGGAGCGCGGCTTCTTCTACACGCCTAAGGCCCGCAGGGAGGTGGAGGAC
  CTGCAGGTGAGGGACGTGGAGCTGGCCGGGGCGCCTGGCGAGGGCGGCCTGCAGCCCCTG
  GCCCTGGAGGGGGCCCTGCAGAAGCGAGGCATCGTGGAGCAGTGCTGCACCAGCATCTGC
  TCCCTCTACCAGCTGGAGAATTACTGCAACTAG
  >OwlMonkey
  ATGGCCCTGTGGATGCACCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCCGAG
  CCAGCCCCGGCCTTTGTGAACCAGCACCTGTGCGGCCCCCACCTGGTGGAAGCCCTCTAC
  CTGGTGTGCGGGGAGCGAGGTTTCTTCTACGCACCCAAGACCCGCCGGGAGGCGGAGGAC
  CTGCAGGTGGGGCAGGTGGAGCTGGGTGGGGGCTCTATCACGGGCAGCCTGCCACCCTTG
  GAGGGTCCCATGCAGAAGCGTGGCGTCGTGGATCAGTGCTGCACCAGCATCTGCTCCCTC
  TACCAGCTGCAGAACTACTGCAACTAG
  >Human
  ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGAC
  CCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTAC
  CTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGGAGGCAGAGGAC
  CTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCTGGTGCAGGCAGCCTGCAGCCCTTG
  GCCCTGGAGGGGTCCCTGCAGAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGC
  TCCCTCTACCAGCTGGAGAACTACTGCAACTAG
  >GreenMonkey
  ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGAC
  CCGGTCCCGGCCTTTGTGAACCAGCACCTGTGCGGCTCCCACCTGGTGGAAGCCCTCTAC
  CTGGTGTGCGGGGAGCGAGGCTTCTTCTACACGCCCAAGACCCGCCGGGAGGCAGAGGAC
  CCGCAGGTGGGGCAGGTAGAGCTGGGCGGGGGCCCTGGCGCAGGCAGCCTGCAGCCCTTG
  GCGCTGGAGGGGTCCCTGCAGAAGCGCGGCATCGTGGAGCAGTGCTGTACCAGCATCTGC
  TCCCTCTACCAGCTGGAGAACTACTGCAACTAG
  >Chimp
  ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGTGCTGCTGGCCCTCTGGGGACCTGAC
  CCAGCCTCGGCCTTTGTGAACCAACACCTGTGCGGCTCCCACCTGGTGGAAGCTCTCTAC
  CTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGGAGGCAGAGGAC
  CTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCTGGTGCAGGCAGCCTGCAGCCCTTG
  GCCCTGGAGGGGTCCCTGCAGAAGCGTGGTATCGTGGAACAATGCTGTACCAGCATCTGC
  TCCCTCTACCAGCTGGAGAACTACTGCAACTAG
  >GuineaPig
  ATGGCTCTGTGGATGCATCTCCTCACCGTGCTGGCCCTGCTGGCCCTCTGGGGGCCCAAC
  ACTAATCAGGCCTTTGTCAGCCGGCATCTGTGCGGCTCCAACTTAGTGGAGACATTGTAT
  TCAGTGTGTCAGGATGATGGCTTCTTCTATATACCCAAGGACCGTCGGGAGCTAGAGGAC
  CCACAGGTGGAGCAGACAGAACTGGGCATGGGCCTGGGGGCAGGTGGACTACAGCCCTTG
  GCACTGGAGATGGCACTACAGAAGCGTGGCATTGTGGATCAGTGCTGTACTGGCACCTGC
  ACACGCCACCAGCTGCAGAGCTACTGCAACTAG
  >Mouse
  ATGGCCCTGTTGGTGCACTTCCTACCCCTGCTGGCCCTGCTTGCCCTCTGGGAGCCCAAA
  CCCACCCAGGCTTTTGTCAAACAGCATCTTTGTGGTCCCCACCTGGTAGAGGCTCTCTAC
  CTGGTGTGTGGGGAGCGTGGCTTCTTCTACACACCCAAGTCCCGCCGTGAAGTGGAGGAC
  CCACAAGTGGAACAACTGGAGCTGGGAGGAAGCCCCGGGGACCTTCAGACCTTGGCGTTG
  GAGGTGGCCCGGCAGAAGCGTGGCATTGTGGATCAGTGCTGCACCAGCATCTGCTCCCTC
  TACCAGCTGGAGAACTACTGCAACTAA
  >Chicken
  ATGGCTCTCTGGATCCGATCACTGCCTCTTCTGGCTCTCCTTGTCTTTTCTGGCCCTGGA
  ACCAGCTATGCAGCTGCCAACCAGCACCTCTGTGGCTCCCACTTGGTGGAGGCTCTCTAC
  CTGGTGTGTGGAGAGCGTGGCTTCTTCTACTCCCCCAAAGCCCGACGGGATGTCGAGCAG
  CCCCTAGTGAGCAGTCCCTTGCGTGGCGAGGCAGGAGTGCTGCCTTTCCAGCAGGAGGAA
  TACGAGAAAGTCAAGCGAGGGATTGTTGAGCAATGCTGCCATAACACGTGTTCCCTCTAC
  CAACTGGAGAACTACTGCAACTAG
  

  Sample peptide dataset

  The following is a the Alpha-globin genes from the dataset above: translated using the standard genetic code, and aligned using MAFFT.

  >Sheep
  MALWTRLVPLLALLALWAPAPAHAFVNQHLCGSHLVEALYLVCGERGFFYTPKARREVEG
  PQVGALELAGGPGAGG-----LEGPPQKRGIVEQCCAGVCSLYQLENYCN
  >Pig
  MALWTRLLPLLALLALWAPAPAQAFVNQHLCGSHLVEALYLVCGERGFFYTPKARREAEN
  PQAGAVELGGGLG--GLQALALEGPPQKRGIVEQCCTSICSLYQLENYCN
  >Dog
  MALWMRLLPLLALLALWAPAPTRAFVNQHLCGSHLVEALYLVCGERGFFYTPKARREVED
  LQVRDVELAGAPGEGGLQPLALEGALQKRGIVEQCCTSICSLYQLENYCN
  >OwlMonkey
  MALWMHLLPLLALLALWGPEPAPAFVNQHLCGPHLVEALYLVCGERGFFYAPKTRREAED
  LQVGQVELGGGSITGSLPP--LEGPMQKRGVVDQCCTSICSLYQLQNYCN
  >Human
  MALWMRLLPLLALLALWGPDPAAAFVNQHLCGSHLVEALYLVCGERGFFYTPKTRREAED
  LQVGQVELGGGPGAGSLQPLALEGSLQKRGIVEQCCTSICSLYQLENYCN
  >GreenMonkey
  MALWMRLLPLLALLALWGPDPVPAFVNQHLCGSHLVEALYLVCGERGFFYTPKTRREAED
  PQVGQVELGGGPGAGSLQPLALEGSLQKRGIVEQCCTSICSLYQLENYCN
  >Chimp
  MALWMRLLPLLVLLALWGPDPASAFVNQHLCGSHLVEALYLVCGERGFFYTPKTRREAED
  LQVGQVELGGGPGAGSLQPLALEGSLQKRGIVEQCCTSICSLYQLENYCN
  >GuineaPig
  MALWMHLLTVLALLALWGPNTNQAFVSRHLCGSNLVETLYSVCQDDGFFYIPKDRRELED
  PQVEQTELGMGLGAGGLQPLALEMALQKRGIVDQCCTGTCTRHQLQSYCN
  >Mouse
  MALLVHFLPLLALLALWEPKPTQAFVKQHLCGPHLVEALYLVCGERGFFYTPKSRREVED
  PQVEQLELGGSPG--DLQTLALEVARQKRGIVDQCCTSICSLYQLENYCN
  >Chicken
  MALWIRSLPLLALLVFSGPGTSYAAANQHLCGSHLVEALYLVCGERGFFYSPKARRDVEQ
  PLVSS-PLRGEAG--VLPFQQEEYEKVKRGIVEQCCHNTCSLYQLENYCN
  

By default the DNA alignment will be in ALN (Clustal) format. The user may select FASTA or MSF instead. The full alignment is shown in the browser window and is also available for download via a link at the top of the page (click on the floppy icon).

In the event of errors and warnings these will be shown at the top of the result page.

New in RevTrans 1.3: the case of the individual letters in the amino acid alignments is now carried over to the DNA alignment. Dialign2 marks amino acids considered to be fully aligned with UPPERCASE and uses lowercase to mark less well aligned regions.

The simple fact that proteins are built from 20 amino acids while DNA only contains four different bases, means that the 'signal-to-noise ratio' in protein sequence alignments is much better than in alignments of DNA. Besides this information-theoretical advantage, protein alignments also benefit from the information that is implicit in empirical substitution matrices such as BLOSUM-62.

Taken together with the generally higher rate of synonymous mutations over non-synonymous ones, this means that the phylogenetic signal disappears much more rapidly from DNA sequences than from the encoded proteins. It is therefore preferable to align coding DNA at the amino acid level and it is for this purpose we have constructed the program RevTrans.

RevTrans constructs a multiple DNA alignment by: (i) translating the DNA; (ii) aligning the resulting peptide sequences; and (iii) building a multiple DNA alignment by 'reverse translation' of the aligned protein sequences. In the resulting DNA alignment, gaps occur in groups of three corresponding to entire codons, and analogous codon positions are therefore always lined up. These features are useful when constructing multiple DNA alignments for phylogenetic analysis. RevTrans also accepts user-provided protein alignments for greater control of the alignment process.