Seqanswers Leaderboard Ad

Collapse

Announcement

Collapse
No announcement yet.
X
 
  • Filter
  • Time
  • Show
Clear All
new posts

  • Sliding window methylation analysis suggestions

    I want to do sliding window analysis for differential methylation calculations for WGBS and target bisulfite sequencing libraries. Obviously I am not comparing the 2 library types against each other. I have found the software to make the sliding windows with specific step sizes but my question is how do you know how big your windows should be and then what step to use. I've read papers that use 3Kb windows all the way down to 500bp windows. Is choosing the window and step just arbitrary? I would like to hear others suggestions on what they have done to determine how big their windows and steps are. I am thinking for low coverage WGBS the larger window is probably best to get enough coverage but I am not sure what "step" to use. For my targeted libraries, I have very high coverage so I should probably use a smaller window. I would just like to hear what others would do. Thanks.

  • #2
    If you have a look at the Differential Methylation Leture document on our methylation training course material and have a look at slides 11 and 12 you can see some information about this.

    The short answer is that the best window size is a balance between statistical power and biological effect size. You need the window to be large enough that you get enough data in it to be able to achieve significance, but you need it to be small enough that it doesn't end up larger than the region whose methylation is changing so you don't get averaging against the surrounding regions.

    We also don't generally recommend using fixed size windows (eg 500bp), since the uneven distribution of CpGs within the genome means that different windows will have very different numbers of CpGs within them, and you therefore get a statistical power bias which can mean getting biases in the regions of the genome where you find hits. We tend to prefer defining windows based on the number of CpGs then contain, which then means you get a variation in resolution, but a more even statistical power.

    Comment


    • #3
      If I use fixed windows versus number of CpG windows as you are suggesting, is the concern that I will get no significance or that I will be flooded with regions of significance that are really false positives?

      Comment

      Latest Articles

      Collapse

      • seqadmin
        Genetic Variation in Immunogenetics and Antibody Diversity
        by seqadmin



        The field of immunogenetics explores how genetic variations influence immune responses and susceptibility to disease. In a recent SEQanswers webinar, Oscar Rodriguez, Ph.D., Postdoctoral Researcher at the University of Louisville, and Ruben Martínez Barricarte, Ph.D., Assistant Professor of Medicine at Vanderbilt University, shared recent advancements in immunogenetics. This article discusses their research on genetic variation in antibody loci, antibody production processes,...
        11-06-2024, 07:24 PM
      • seqadmin
        Choosing Between NGS and qPCR
        by seqadmin



        Next-generation sequencing (NGS) and quantitative polymerase chain reaction (qPCR) are essential techniques for investigating the genome, transcriptome, and epigenome. In many cases, choosing the appropriate technique is straightforward, but in others, it can be more challenging to determine the most effective option. A simple distinction is that smaller, more focused projects are typically better suited for qPCR, while larger, more complex datasets benefit from NGS. However,...
        10-18-2024, 07:11 AM

      ad_right_rmr

      Collapse

      News

      Collapse

      Topics Statistics Last Post
      Started by seqadmin, 11-01-2024, 06:09 AM
      0 responses
      29 views
      0 likes
      Last Post seqadmin  
      Started by seqadmin, 10-30-2024, 05:31 AM
      0 responses
      21 views
      0 likes
      Last Post seqadmin  
      Started by seqadmin, 10-24-2024, 06:58 AM
      0 responses
      26 views
      0 likes
      Last Post seqadmin  
      Started by seqadmin, 10-23-2024, 08:43 AM
      0 responses
      57 views
      0 likes
      Last Post seqadmin  
      Working...
      X