DESEQ2 design with unequal matching and replicates?
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divyak • 0
@divyak-23015
Last seen 4.8 years ago

Hi

I have an experiment with two different tissue samples taken from 8 patients. But not all 8 of them have both tissue samples. 5 have both tissue samples, 2 have only the first kind of sample and 1 has the second kind but not first. Additionally, one of the samples also has two technical replicates. How do I factor all this into DESEQ2 design?

  1. Can I do design = ~patient + sample, despite not all of my patients having both kinds of samples?
  2. Can I pick one out the two technical replicates alone for the analysis?
deseq2 RNASeq • 1.1k views
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@mikelove
Last seen 6 days ago
United States

We typically add technical replicates (same library, multiple sequencing runs).

In DESeq2, you can only include full pairing, but in limma you could use duplicateCorrelation to incidate partial pairing.
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Thanks for your reply! What generally happens if DESeq2 is run without replicates, though?

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Can you be more specific? I don't know what situation you are describing.

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Sorry! In my experiment, I have 5 samples in each of the two groups (A and B) I am comparing. The 5 samples are from 5 patients. So they aren't technical replicates in that they aren't from the same library. We've taken a sample of A from a patient and then a sample of B from another patient and repeated this with 5 patients. Will DESeq2 still work then to do a Group A vs B comparison?

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Yes you’d do patient + condition as the design.

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Awesome! Thanks. But what if I have 8 on each group but only 5 match with patients? The other 3 are from different patients and do not have a match in group B. Should I discard those three or will patient + condition still work?

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Yeah we can only make use of paired or not, whereas limma can strike a balance between these.

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Thanks for your reply! What generally happens if DESeq2 is run without replicates, though?

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