Merging, Quality control and overlapping

Hello Armin,

Thank you for posting this really great question along with the detailed screenshots. I think you are on the right track.

denoised: 99,046, merged: 57

:scream_cat:
Yep, that's your problem. Let's fix it!


Sure. The pair of Illumina reads sequence your amplicons from both ends, with some overlap in the middle.

250 bp amplicon  |-------------------------|
150 bp read      |--------------->
150 bp read                <---------------|
50 bp overlap (correct!)    ^^^^^

Here's how I work it out:

(forward read) + (reverse read) - (length of amplicon)  = overlap
150 forward + 150 reverse - 250 amplicon = 50 overlap

This matches the diagram I showed above.

For your data, here are the numbers I know so far:

216 forward + (218 - 2) reverse - (16S-341F and 16S-805R) amplicon = overlap
216 forward + 216 reverse - (805 - 341) amplicon = overlap
432 - 464 = overlap
-32 = overlap

EDIT: So your overlap is negative, which means there is a 32 bp gap between your paired ends. :scream_cat:

I agree! Let's me know if this helps answer your questions, then we can see how we can fix it!

Colin

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