Question: RNA-Seq analysis edgeR
0
gravatar for eb0906
4.8 years ago by
eb090620
United States
eb090620 wrote:

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?
ADD COMMENTlink modified 4.8 years ago by Devon Ryan92k • written 4.8 years ago by eb090620
3
gravatar for Devon Ryan
4.8 years ago by
Devon Ryan92k
Freiburg, Germany
Devon Ryan92k wrote:
  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.
ADD COMMENTlink written 4.8 years ago by Devon Ryan92k

Thank you for your help!

ADD REPLYlink written 4.8 years ago by eb090620
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