Alignment Problem

[benjjneb (Ben Callahan)]

@akknight216

Also, just out of curiosity since this is so strange, I recently reran dada2 and the only difference was whether I trimmed 10 nucleotides off the front of the forward read, and suddenly I went from 30,000 features with the trimming to >300,000 features. Is that to be expected? And potentially related to this problem? Though the feature table does write out normally.

That is not expected behavior unless... do you have primers/linker/non-bio-nts in your reads? If so, you need to remove them (whether by trimming off the primer with trim-left or with a dedicated tool).

DADA2 (and any method that infers exact sequences) will interpret the ambiguous nucleotides that are in most primers as biological variation, which can artefactually multiply the number of features you get out at the end. For normal data, w/o primer sequence, the number of features should change relatively little when chopping off 10 nts.