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  • yumiya
    Junior Member
    • Oct 2011
    • 3

    #1

    Mapped onto pseudogenes?

    I have a problem in TopHat-Cufflinks pipeline. I want to get splicing information of a gene family, but the annotation is bad so far, that is, nothing is annotated or huge exons are annotated.
    This family has a number of processed pseudogenes with almost same sequence. It seems that reads that are supposed to be mapped onto "real" genes are mapped onto these pseudogenes.
    Now, I have a gff file defining the location of those pseudogenes in the genome. Is there a way to mask these pseudogenes in TopHat mapping?
    I do not think -M option in Cufflinks works for me, because the reads are already mapped in .bam file from TopHat.
    I really need help. If someone knows, please tell me
  • yumiya
    Junior Member
    • Oct 2011
    • 3

    #2
    Masking

    I masked pseudogenes on the genome with "NNNNN....".
    That worked at least partly.

    Comment

    • Dario1984
      Senior Member
      • Jun 2011
      • 166

      #3
      Some pseduogenes are transcribed, though. So, this isn't a good solution, either. It's a problem for us, too. We get lots of poor transcript assemblies because of this.

      Comment

      • yumiya
        Junior Member
        • Oct 2011
        • 3

        #4
        Yes, that's true, but in our case we know that expression of pseudogenes is at a very low level compared to real genes. Moreover, our goal is qualitative rather than quantitative, that is, identification of alternative isoforms.
        If I find alternative isoforms in some genes, I can go back to the masked pseudogenes to see whether they might give the identified isoforms (I am talking about "processed" pseudogenes).
        However, masking is of course not a good solution for quantitative analyses...

        By the way, I found another key. If you are using TopHat and your target genes are highly expressed, --initial-read-mismatches should be 0. Minimize loss of reads derived from true genes by pseudogenes in initial mapping with this option.

        Comment

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