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Old 12-16-2017, 05:27 AM   #1
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Location: Oxford, Ohio

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Question MiSeq Sequencing Short Amplicons


If I am sequencing a heterogeneous mixture of amplicons using a MiSeq (2x250bp) to ensure overlap, and there are some short amplicons (i.e. ~ 150 bp), what happens to the run? Will the rxns simply be wasted on the short amplicons, or will there be adverse affect on the data?

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Old 12-16-2017, 08:25 PM   #2
Jafar Jabbari
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Two effects:

1- As noted wasting sequencing
2- Low overall Q30 score for the run without affecting inserts Q scores
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Old 12-18-2017, 04:09 PM   #3
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if you have a lot of the shorter amplicons, there's a chance that the software will just kill the run because of difficulty focusing when a lot of the clusters are dark. I've run up to 25% or so short amplicons without killing a run, but never tried higher than that
Microbial ecologist, running a sequencing core. I have lots of strong opinions on how to survey communities, pretty sure some are even correct.
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amplicon sequencing

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