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  • DrMTB
    Junior Member
    • Jun 2012
    • 3

    #1

    Fastx-Toolkit Newbie help IUB codes in fastq causing error

    I simply wanted to try out a few command line alignments of ABI sanger sequences. I converted the .phd.1 files to fastq using a BioPerl script (http://www.bioperl.org/wiki/HOWTO:SeqIO). Then I wanted to trim these files according to the quality scores and so have been using the fastx-toolkit. Problem was the fastq nucleotide sequence gave an error ('found invalid nucleotide sequence') using any of the tools.
    Using a test file as follows was OK:
    @test
    GTGCGGTGGTGGAGACGCACTTGATAGTCCTTCTCCGCAGGTACTTT
    +
    &$"'&&$&&&""+)(&(*,213+)*),,/4:=2-,&",.1.9-7884

    But replacing any base with an IUB code gave the error again.

    Eg using (see R at position 5):
    @test
    GTGCRGTGGTGGAGACGCACTTGATAGTCCTTCTCCGCAGGTACTTT
    +
    &$"'&&$&&&""+)(&(*,213+)*),,/4:=2-,&",.1.9-7884

    $ fastx_quality_stats -i test.fastq -o test.stats -Q33
    fastx_quality_stats: found invalid nucleotide sequence (GTGCRGTGGTGGAGACGCACTTGATAGTCCTTCTCCGCAGGTACTTT) on line 2

    This must be the cause of the error - but why? Fastq files can have IUB codes surely!?
    Thanks for any help on this apparently simple question.

    Using :
    Ubuntu
    FASTX Toolkit 0.0.13.1
  • maubp
    Peter (Biopython etc)
    • Jul 2009
    • 1544

    #2
    IUPAC ambiguity codes are valid in FASTQ, but typically you'll only see N (and ACGT). It seems that FASTX doesn't like this - you could report it.

    Comment

    • DrMTB
      Junior Member
      • Jun 2012
      • 3

      #3
      Thanks maubp
      Reply from Fastx-toolkit author says that these tools were designed to work with files from Illumina machines containing only ACGT & N and will therefore fail on other kinds of input.
      So that's that.

      Comment

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