Pooling samples for de novo chimera detection
UCHIME order dependency
I now consider de novo
UCHIME obsolete for OTU clustering pipelines because
UPARSE is a superior approach..
I recommend that you combine samples for de novo
chimera detection. (Previously, I have recommended that PCR runs should be
processed separately, but I now realize that it is probably better to combine
runs). The reasoning is as follows: the main concern is false negatives (FNs),
because undetected chimeras are usually more harmful in a biological analysis
than false positives (FPs).
FNs occur if a chimera has has read abundance that is greater than or equal one
of its parents. Combining runs is likely to increase the abundance of a parent
(because the same species occurs in multiple samples), but is unlikely to
increase the abundance of a chimera. If the chimera is reproduced in a second
sample, the parents must also be present and are likely to be present in higher
abundances. So pooling should tend to reduce FNs, and there is no reason I can
think of why FPs would tend to increase.