I recently submitted a paper for publication, in which I describe the species of zooplankton identified in water samples using metabarcoding, as well as the percent composition. The percent composition estimated as the number of reads of a species over total reads. These results were compared for different markers A reviewer made the following comments below, but I do not understand exactly what they think I should have done.
Comment by reviewer:
"Are DNA metabarcoding data processing steps, i.e., standardization or rarefaction, data transformations, etc. performed? The methodology lacked this vital data analysis information which makes it difficult to comment on whether proper data processing was explored in the study to support a reliable interpretation of the results."
Maybe you don't understand this, because you did not perform any analysis on your data and worked on raw number of reads to calculate your frequences?
This is a possibility, but you should specify it I guess.
I do not really understand the comment neither, maybe the reviewer wanted to refer to any data cleaning process (https://benjjneb.github.io/decontam/vignettes/decontam_intro.html).
What surprises me is that DNA metabarcoding do not provide any quantitative data and you can't make any relation between the number of reads and the abundance of a species in the community sampled (because of PCR stochasticity). These analyses should only be done on presence/absence data, or at best on occurence frequencies (numbre of time a species is present over all samples), but not provide quantification based on reads.
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