HTG EdgeSeq miRNA on plasma samples: DESeq2 normalization concerns

Question

[myguestp]

I have counts data from a HTG Edge-Seq miRNA Whole Transcriptome experiment perfomed on plasma samples from patients.

I wonder if the mean normalization that DESEq2 does assumes that the global expression of miRNAs is identical between the conditions. For example, if my study condition causes a drop in global miRNA levels, could I see it after normalizing with DESeq2? Wouldn't this create an artificial scaling to match the global expression between samples?

In these samples no spike-in was added, is there any normalization method that does not start from the premise that the global expression is identical between samples?

Thanks in advance

Answer 1

[swbarnes2]

Without spike-ins I don't think it's possible to distinguish between your having fewer miRNA molecules, and the people loading the instrument simply loading a little less sample.

[myguestp]

Thanks. I have no evidence that this type of phenomenon occurs in my study condition, but I wanted to be sure of the implications of the normalization method.