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  • fastq_quality_trimmer error

    I'm having problems in running commands of fastx tool. For example, when I ran fastx_trimmer,
    fastx_trimmer -i in.txt -o out.fastq

    I received this error
    fastx_trimmer: Invalid quality score value (char '#' ord 35 quality value -29) on line 4

    Even I tried to add "-Q 64", still the same error.
    fastx_trimmer -i in.txt -Q 64 -o out.fastq


    Why is that??

  • #2
    If your data is in sanger format then -Q 33 would be the option to add.

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    • #3
      Sorry, I forgot to mention my sample is Illumina. The sample length is 17kb.

      Comment


      • #4
        Originally posted by shuang View Post
        I'm having problems in running commands of fastx tool. For example, when I ran fastx_trimmer,
        fastx_trimmer -i in.txt -o out.fastq

        I received this error
        fastx_trimmer: Invalid quality score value (char '#' ord 35 quality value -29) on line 4

        Even I tried to add "-Q 64", still the same error.
        fastx_trimmer -i in.txt -Q 64 -o out.fastq


        Why is that??
        Illumina stopped using the Phred+64 offset sometime ago; they now use Phred+33. The fastx toolkit assumes Phred+64 by default so your two examples were exactly the same. As GenoMax said you need to use -Q33.

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        • #5
          When I use -Q 33, it didn't go much further, neither. Below is the command and the error message.
          fastx_trimmer -i in.txt -Q 33 -o all.fastq


          fastx_trimmer: Error: invalid quality score data on line 448 (quality_tok = "??????B?D?DDDBB?FFFF;FFBB>FFHHDD?EGH>EFEAECEDHHHHHGHFE:FDGA-CEDHHDBF>AAACF=AAEBGFGF?CDDEHH+=CCCCBDAFF?DF.6BFF66...=B<*645,,5A,5,,53>?;>A,A5==*..8A?*:C"

          Comment


          • #6
            shuang: Can you tell us what OS you are using to run the fastx_trimmer?

            Did you open/edit the sequence file ("in.txt") in a non-unix OS (e.g. windows/OS X)?

            What do you mean by sample length is 17kb?

            Comment


            • #7
              I use Ubuntu. The in.txt is a fastq file, which was downloaded directly from the sequencing facility website.

              17kb is the expected fully assembled length.

              Comment


              • #8
                same problem

                I am using fastq_quality_ trimmer for the first time and I am having this same problem also on Illumina generated data. Does anyone know what the solution to this is?

                Comment


                • #9
                  Just use TRIMMOMATIC.
                  Works better with more options

                  Comment


                  • #10
                    There should be no space between the Q and the 33.

                    You also need to specify how you want to trim by setting variables for -f and -l e.g.

                    fastx_trimmer -Q33 -f 1 -l 50 -i input.fastq -o output.fastq

                    AFAIK Fastq's also need to be uncompressed for input but can be compressed on output using the -z flag.

                    Use fastx_trimmer -h for help

                    Comment


                    • #11
                      I had the exactly same problem! It seems the problem is more of -Q33 option as I tried it.
                      Two things here:

                      1) which is the wrong quality score, i.e. ASCII char when you have the -Q33 on?

                      2) our sequence machine was run with the MS-Windows platform and the data was deposited on Linux storage disk. There might be a issue with this cross platform situation, but how to solve it?

                      Thanks a lot!

                      Comment

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