Seqanswers Leaderboard Ad

Collapse
X
 
  • Filter
  • Time
  • Show
Clear All
new posts
  • bioliyezhang
    Member
    • Mar 2011
    • 19

    Question: problem with data containing non-finite values in cummeRund processing

    Hi,all:

    Recently, I am trying to identify the differentially expressed genes between two conditions. Each condition has three replicates(paired end). I got a problem when I am trying to analyze the data using Tophat v2.0.4, Cufflinks v2.0.2 and cummeRbund v2.0.0.

    The problem is that when I try to generate the density plot using cummeRbund. It showed warning message like this:


    Warning messages:

    1: Removed 24226 rows containing non-finite values (stat_density).

    2: Removed 22713 rows containing non-finite values (stat_density).

    And the volcano plot (http://www.freeimagehosting.net/ot994) also did not seem to be right. I am not clear the cause of non-finite values in my dataset and why there are so many of them have this problem.

    I suspect that this is because I use "--compatible-hits-norm" normalization parameter in cufflinks. I just wonder whether you have some experience on this and any suggestions will be greatly appreciated.

    Best, Liye
    Last edited by bioliyezhang; 10-09-2012, 01:03 PM.
  • blanco
    Member
    • Apr 2012
    • 28

    #2
    Hi Liye,
    I believe this is caused by fpkm values of zero. When the program takes the log of zero it becomes minus infinite (-Inf) and these values are excluded from the analysis.

    I think there is an option called "pseudocount=0.0001" which adds 0.0001 to all zero values to work around this and keep all values in the analysis. If you do that the density should pile up close to the y-axis. Not sure if that is what you would want but it is an option.

    I tend to think it is ok to discard the zero fpkm values.

    Comment

    • bioliyezhang
      Member
      • Mar 2011
      • 19

      #3
      Hi, Blanco:

      thanks a lot. Your explanation for the non-finite values is very clear. And I tried to set the pseudocount, and it worked as you described.
      However, it seem to me that the problem in Volcano plot is another problem. Because, even if I set pseudocount to an non-zero value, same error occurred again.

      Removed 4573 rows containing missing values (geom_point).

      I am pretty new to Cufflinks and CummeRbund. Because previously there are 24226 and 22713 transcripts have zero value in FPKM. My guess is that there are 4573 genes have zero FPKM value in both sample and control case, which lead to the missing values (geom_point) in Volcano plot? I am still trying to check this by looking into the data itself.

      I wonder whether you have any clue on the cause of this missing values?
      Thanks a lot.

      Comment

      • lgoff
        Member
        • Feb 2008
        • 82

        #4
        This is a similar situation in the volcano plot, with the exception that this warning is not arising from zero-values, but rather we restrict the axes to make the plot a bit more visually interpretable due to very high and very low log-fold change values. You should be able to adjust the x and y axes to be more inclusive of the missing values if you like, but be forewarned that the log-fold change values can be very high in these data and this will compress the rest of the image.

        -Loyal

        Comment

        • bioliyezhang
          Member
          • Mar 2011
          • 19

          #5
          After checking into the data, I see what you are trying to say.
          Basically, the Removed 4573 rows is due to their have higher test_stat value (one column of .diff output) the default cutoff, which seems to be 40.
          So basically the data with too high test-stat value is removed for visualization purpose. So there is no problem with the data it self.

          Thanks.

          Comment

          • rosielee
            Junior Member
            • Dec 2013
            • 1

            #6
            cummeRbund (ref the Jan 2014 Bioconductor package manual) describes how one can add pseudocount=eg. 0.0001 or 1.0 (for reasons discussed above)

            Is there any specific reason why a value of 0.0001 is used in the csBoxplot function and it is set to 1.0 in other functions (eg. csDistHeat)?

            Or have these values been shown just as examples of the type of values that one could use?

            If carrying out an analysis with one set of data, would you keep the pseudocount value the same across all the functions?

            (sorry if these questions are daft, I am very new to RNA-seq and cummeRbund)

            Thanks in advance
            Rosielee

            Comment

            Latest Articles

            Collapse

            • seqadmin
              Pathogen Surveillance with Advanced Genomic Tools
              by seqadmin




              The COVID-19 pandemic highlighted the need for proactive pathogen surveillance systems. As ongoing threats like avian influenza and newly emerging infections continue to pose risks, researchers are working to improve how quickly and accurately pathogens can be identified and tracked. In a recent SEQanswers webinar, two experts discussed how next-generation sequencing (NGS) and machine learning are shaping efforts to monitor viral variation and trace the origins of infectious...
              03-24-2025, 11:48 AM
            • seqadmin
              New Genomics Tools and Methods Shared at AGBT 2025
              by seqadmin


              This year’s Advances in Genome Biology and Technology (AGBT) General Meeting commemorated the 25th anniversary of the event at its original venue on Marco Island, Florida. While this year’s event didn’t include high-profile musical performances, the industry announcements and cutting-edge research still drew the attention of leading scientists.

              The Headliner
              The biggest announcement was Roche stepping back into the sequencing platform market. In the years since...
              03-03-2025, 01:39 PM

            ad_right_rmr

            Collapse

            News

            Collapse

            Topics Statistics Last Post
            Started by seqadmin, 03-20-2025, 05:03 AM
            0 responses
            49 views
            0 reactions
            Last Post seqadmin  
            Started by seqadmin, 03-19-2025, 07:27 AM
            0 responses
            57 views
            0 reactions
            Last Post seqadmin  
            Started by seqadmin, 03-18-2025, 12:50 PM
            0 responses
            50 views
            0 reactions
            Last Post seqadmin  
            Started by seqadmin, 03-03-2025, 01:15 PM
            0 responses
            201 views
            0 reactions
            Last Post seqadmin  
            Working...