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  • Help with RSEM: how to change default phred33 to phred64

    Hi all,

    When I was using the RSEM to align my reads.fastq to my transcripts.fasta (trinityrnaseq_r2013-02-25/util/RSEM_util/run_RSEM_align_n_estimate.pl), I happened to see this:

    bowtie -q --phred33-quals -n 2 -e 99999999 -l 25 -I 1 -X 1000 --chunkmbs 200 -p 4 -a -m 200 -S TRANS -1

    But my fastq used phred64, so is there a way to change the default setting (phred33)? Does it affect much if I used the default setting?

    Thanks,
    He

  • #2
    It shouldn't matter much. But if you want to change the quality, you could download BBTools and do this:

    reformat.sh in=reads.fastq out=fixed.fastq qin=64 qout=33

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    • #3
      From the trinity page--


      # To pass additional parameters to rsem-calculate-expression,
      # type ' -- ' followed by additional pass-through params
      You could try <original command> --phred64-quals
      Other parameters can be passed in the same way.

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      • #4
        Hi Yueluo,
        I tried typing --phred64-quals behind my command (../run_RSEM_align_n_estimate.pl/ --transcripts) but it said: Unknown option: phred64-quals.
        He

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        • #5
          Brian,
          I will try it and see if it works.
          Thanks,
          He

          Comment


          • #6
            Well I guess you could follow what Brian suggested. But it is strange you can't use the '--phred64-quals' option as it is mentioned on both trinity's and RSEM's websites.

            Converting the phred-qual isn't too difficult, but it is time-consuming when you process large data sets and adds pressure to the system's I/O and storage -- which is a concern to myself and probably some others.

            Comment


            • #7
              Originally posted by yueluo View Post
              Converting the phred-qual isn't too difficult, but it is time-consuming when you process large data sets and adds pressure to the system's I/O and storage -- which is a concern to myself and probably some others.
              So you would think! But no, reformat.sh is extremely fast, clocked at over 500MB/s (note that a single HDD won't be able to deliver this). And you can pipe it, like this:

              reformat.sh in=reads.fq out=stdout.fq qin=64 qout=33 | someOtherProcess

              ...which will add no load to the system's I/O or storage, as long as the other process accepts stdin. For, you see, I am also concerned with these things!

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              • #8
                Good to hear Brian ! I'll give it a go soon and see how well that goes

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