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RNA-Seq analysis edgeR

We have a time course infection experiment with 12 samples (2 technical replicates and 2 biological replicates per sample). The samples were sequenced on a HiSeq over 2 lanes, 1 x 50 bp reads.

  1. 24hr_infection1_lane3
  2. 24hr_infection1_ lane4
  3. 24hr_infection2_lane3
  4. 24hr_infection2_lane4
  5. 24hr_control1_lane3
  6. 24hr_control1_lane4
  7. 24hr_control2_lane3
  8. 24hr_control2_lane4
  9. 48hr_infection1_lane3
  10. 48hr_infection1_lane4
  11. 48hr_infection2_lane3
  12. 48hr_infection2_lane4

I aligned the samples using Tophat and then converted the .bam files to .sam files and created a counts file using HTSeq for each sample. My questions now are:

  1. Should the technical replicates be collapsed (add gene counts for technical replicates)?
  2. How should the design matrix be setup in edgeR?
edger rna-seq technicalvsbiological_replicates
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updated by Devon Ryan Devon Ryan • written by eb0906 • eb0906

1 answer

  1. Yes, add the technical replicates together.
  2. ~time+treatment, where time is a factor of 24hr and 48hr and treatment is control and infection. You obviously can't distinguish between an effect of time and an interaction between time and treatment.
• 0 views • link
written by Devon Ryan Devon Ryan

Thank you for your help!

• 1 views • link
written by eb0906 • eb0906

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