Batch effect between biological replicates
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gtechbio ▴ 10
@gtechbio-13996
Last seen 17 months ago
Spain

I got an RNA-Seq project where the data looks like as in the PCA plot below - 4 biological replicates of 4 conditions, and there are 4 batches with each batch consisting of 1 sample from each condition. In essence the variation between biological replicates is larger than the variation in the tested conditions most likely due to batch effects. Likely the wet-lab has processed each replicate of all conditions at different days.

I was thinking to include a batch into the design formulae, but on the other hand (as far as I understand) it will "remove" the variation between biological replicates, so I was wondering if that analysis would make sense?

Any advice is much appreciated

DISCLOSURE: Cross-posted in Biostars a week ago https://www.biostars.org/p/9562841/#9563062 enter image description here

BatchEffect DESeq2 • 1.4k views
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@mikelove
Last seen 1 hour ago
United States

Your design is not confounded, you can use ~batch + condition. You will still have residual degrees of freedom because you have 16 samples and this design has 5 coefficients.

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Many thanks for clarifying Mike! Just one last doubt regarding the fact that this design will control for the difference between biological replicates, does it somehow affect the final interpretation of the results? Thank you once again

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