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  • Tophat2 normal alignment rate

    Hi there, I am a complete RNA seq newbie, so let me excuse myself preventively for any stupid questions.

    Recently I started working with mouse RNA seq data, after running the data though Tophat2 I got a 91.0% overall read alignment rate and a 83.1% concordant pair alignment rate.

    Could somebody with more experience in the field tell me if this are good alignment values, and if not which are the normal values I should expect ?

    Thanks in advanced.

  • #2
    what read length, not bad if 100mers

    Comment


    • #3
      Sorry I knew I had forgotten to mention something, the reads have a length of 101.
      Last edited by JJP; 07-18-2013, 04:56 AM.

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      • #4
        Dear Jon
        I have posted the same problem here;
        Discussion of next-gen sequencing related bioinformatics: resources, algorithms, open source efforts, etc

        any solution ?

        Originally posted by Jon_Keats View Post
        what read length, not bad if 100mers

        Comment


        • #5
          I just finished running tophat on about 20 cancer samples and my concordant pair alignment rate ranged between 50-60% for 50 bp reads. Should I be worried? Will this have an impact on the downstream cufflinks/cuffdiff analysis?

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          • #6
            I presume you are talking about *human* samples. The 50-60% rate is low. I suspect that since the samples are more-or-less uniform in their mapping rate then you won't have too many problems with cufflinks/cuffdiff aside from not having as much data to work with.

            Did you run non-cancer controls? Or are those part of the "20" you mention above. How about replicates -- technical and biological?

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            • #7
              Yes, human samples. We did run non-cancer controls but they did not have this problem. What do you think this means? Is this more a reflection of biology or an indication of a technical issue? We do not have any technical replicates, only biological.

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              • #8
                The lack of technical replicates is not a big concern -- although they can help troubleshooting usually the technical process is flawless. Although when problems do occur you really wish that you had technical reps. :-)

                As for the question "is a 50-60% concordant rate normal for cancer samples when the non-cancer samples have a higher concordant rate?" ... I am going to have to skip on answering this. I simply do not have enough experience with human cancer samples to say one way or another. Cancer genome rearrangements can be widespread so may 50-60% if expected. Can anyone with more human experience chime in?

                BTW: You may wish to make a new thread on this topic with the keywords "human"
                and "cancer" in it. That might attract more knowledgeable people.

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