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  • How to trim dual index adapters with Trim Galore!

    Hi everybody,
    I am trying to trim some adapters with Trim Galore from my bisulfite sequencing data, but I don't know what command to use.
    It has been really difficult to take the information of library construction and I only know that it has been done using IDT unique dual indexing adapters in paired end data, detailed as follows:

    1 - P5 ATATGCGC, P7 CTGATCGT
    30 - P5 AACGTCTG, P7 GCGTCATT
    31 - P5 TATCGGTC, P7 ATCCGGTA
    32 - P5 CGCTCTAT, P7 CGTTGCAA

    When doing the FASTQC analysis I have found some overrepresented sequences such as:
    - In one of the sequence of the pair
    AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA
    - In the other sequence of the pair
    GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG
    TTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTT

    An also in the adapter content section the Illumina Universal adapters are identified

    Does all this make sense?
    So I need to add something more to those P5 and P7 adapter sequences when running Trim Galore?

    Will this command be correct for the first example of unique dual index adapters?

    > trim_galore --paired -a ATATGCGC -a2 CTGATCGT -q 15 --stringency 5 -e 0.05 --length 50 --fastqc --gzip file_1.fastq.gz file_2.fastq.gz

    Thanks in advance!

  • #2
    Index sequences are never part of an actual read in Illumina sequencing so you are never going to find index sequences you noted above in R1/R2 reads.

    As for the poly-A and ploy-G (no signal, two color chemistry) you just need to remove them during trimming.

    Comment


    • #3
      Thanks for your answer!
      As the only information of the sequencing service was that about the dual index adapters I though that those were the sequences that I need to trim.
      So for the command how do I need to add this poly A, T and Gs?
      One by one in each sequence file? or combining the paired end sequences somehow?

      Thanks again!

      Comment


      • #4
        I suggest you try "bbduk.sh" from BBMap suite instead. A guide is available. You can trim out all these things in one pass.

        I am not a trim galore user but you may be able to additional specify poly-A and poly-G there as well.

        Comment

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