The requirement to include replicates of samples when conducting experiments is rarely fulfilled in studies of microbial ecology (particularly those which utilise high-throughput sequencing techniques), although it is a basic requirement in most scientific disciplines including most fields in biology, and despite having been stressed out in the literature (e.g. Prosser 2010, EMI).
Nevertheless, with the use of tagged sequencing technique one can easily analyse replicated samples in parallel, yet this issue seems to be absent in the mothur tutorials.
I was wondering if people here have suggestions on how to incorporate the replicates in the different stages of sequence analysis in order to achieve greater statistical robustness.
Particularly I’m looking for suggestions that go beyond the trivial steps of merging the results or of manually examining whether the replicates have the same taxonomical patterns or diversity indices.
Rather I’m looking for ways to include replicates more explicitly, for instance when computing diversity metrics or testing hypotheses.
Suggestions are welcomed!
Roey