Hi @Shinthotrang,
Just a few things I'd like to add to @timanix' great comments...
You should be aware of these confounding issues:
- PCR reaction biases
- PCR primers will incur primer amplification biases that will not necessarily be fixed by trimming to the same length. Often the composition and diversity of samples can be primarily associated with the PCR primers used over the biological signal you are investigating.
- truncation / trimming parameters
- should be identical for the same primer pairs. That is, if you are using the same amplicon across different runs, and truncate the reverse read by just a few base pairs for one run and not the other, you'll generate many non-overlapping ASVs between the runs. This is why it is often recommended to use the same parameters for all runs that use the same primers.
- In your case, when trying to combine sequencing runs generated from different primers sets, even if these primers target the same region and only differ by a few bases, you'll observe even more exaggerated problems from #1 above. With the additional conflated problem of requiring different trimming / truncation values.
The TL;DR is, using different trimming/ truncation parameters along with different primer pairs, across different runs, can lead to lots of spurious interpretations.
Check out this post.