![]() ![]() (251-17 + 234-20 = 448) If your amplicon is 420 bp long, and you need 20 bp of overlap to complete the merge, that leaves you only 8 bp of wiggle room. Your sequence runs may be 2x300, but you’re only handing DADA2 448 base pairs to work with during the merge. I don’t think that the length of the amplicon is the problem, it has a length of around 420 bp. If you think this is not your situation, please share further sequencing details so we have a little more to work with. Let us know if you have trouble moving forward. Once you’ve determined whether this is your situation, there are quite a few posts here that discuss possible next steps. There’s a good discussion of this in these posts. If you’re working with a longer amplicon, it’s possible that many of your reads have insufficient overlap to merge. DADA2 requires ~20 base pairs of overlap between forward and reverse reads in order to join them. Your denoising-stats visualization suggests that sequences are dropping out because they are failing to merge. If so, please be more topic-specific in future. I’ll do my best to help out, but some housekeeping first - because this post was split from another post, I’m not sure whether you generated the title. Thanks so much for including screencaps and complete commands. ![]()
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