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  • "Stranded" data may actually be unstranded?

    Hello!

    I have some paired-end RNA-seq data that I was told were "stranded", but nothing about whether it's RF or FR.

    I aligned some randomly sampled pairs of reads to the reference genome of the organism. I see that 50% of the read 1s map to the "+" strand, the other 50% to the "-" strand. Same as the read 2s.

    Does this mean that the data is actually unstranded?

    P.S. I can't get QoRTs to work on my data, so I can't use it to tell me whether the data is stranded or not.

    Thank you!

  • #2
    Hi DMNO,

    did you take the transcripts' strand-orientation into account? Try using RSeQC's infer-experiment tool. It gives you a good way to assess if your library was stranded and in which orientation.

    Cheers,

    Michael

    Comment


    • #3
      Thank you very much, Michael!

      The output of infer_experiment.py is:


      Fraction of reads failed to determine: 0.0098
      Fraction of reads explained by "1++,1--,2+-,2-+": 0.0089
      Fraction of reads explained by "1+-,1-+,2++,2--": 0.9813


      From what I understand, this means that the library is RF?

      Comment

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