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Help in understanding the Demultiplexing summary files

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  • Help in understanding the Demultiplexing summary files

    Hi all,
    I'm working on analysing my RNA-seq data from NextSeq 550 machine.

    Among all the things I'm trying to understand, there is a main concern that I haven't found any information on.
    When I open my fastq files (per sample-index), I can see that each read has assigned the appropriate index in most of the cases, but also some of them have similar but not exact the exact index assigned (not the index belonging to a different sample, but different), and some have completely random indexes that have no similarity to the index that should be there. Is this a problem? I have already checked that the indexes I introduced in the sample sheet are correct.

    Moreover, when looking into the demultiplexing summary, I can see that my indexes are top in the most popular indexes, but then there are a bunch of random indexes that do not match to the indexes available in my RNA library preparation kit.

    And finally, is this related anyhow to how many undetermined reads I got? Around 20-25% for each lane. I added 1% PhiX control library to my sequencing.

    Thanks a lot!!

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