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How to handle Element AVITI data (16S amplicon sequencing)
I'm processing our first Element AVITI 16S amplicon sequencing data, using dada2 in QIIME2. I'm wondering how to do this optimally, as it appears that it behaves a bit differently from MiSeq data in dada2:
The phred quality stays much higher than in MiSeq usually, but to my surprise, I still need to truncate the reads just as short to get similar % accepted in DADA2. For more details & actual statistics, see this thread in the QIIME2 forums.
I was first wondering if the AVITI phred scores are a tad optimistic, but got a potentially interesting comment from AVITI bioinformatician. They think the scoring should be quite comparable vs MiSeq BUT: ”There are differences in the distribution of q scores within a read, however, which would be relevant to DADA2 filtering. AVITI data will have greater q score variance within a read--in AVITI data you would be more likely to find a single low q score in the middle of a high quality read.”
Therefore, they recommend we try tweaking maxEE. Does this sound like a good idea? How should we evaluate/validate the results?
We have a huge number of reads to begin with so that’s no problem, and by truncating close to minimum required overlap we get comparable % of reads through vs. MiSeq. But it feels stupid to throw away high quality data just because I don’t completely understand what’s going on.
The QIIME2 guys recommended me to open an issue here, because this calls for a deep level understanding of dada2.