Where to trim/truncate reads in DADA2

[lizgehret]

Hi @joaomiranda,

@joaomiranda

So I observed that I have 4.169.369 sequences totally, both forward and reverse, that's a lot of reads. If I come to run a de-noising with both forward and reverse reads, and the filtering step exclude the half of my sequences, I will still have many sequences, what do you think?

Yes - I would just be mindful of the average quality score of your reads. Going back to your original question:

@joaomiranda

So I'm confused where I have to truncate and trim?

Since you've confirmed that there are no remaining barcodes in your sequences and that they have already been de-multiplexed, we can probably assume that you are just seeing poor quality forward reads from ~45-65nt. In that case, you could set your trim/truncate values to cut off that segment of poor quality forward reads, and just utilize something from, say 65nt to anywhere from 160-230nt (depending on the average quality score you'd like to use as your cutoff point).

@joaomiranda

In addition where to trim and truncate, I have doubts about use trunc-q and max-ee, if I should use those parameters in my particular case.

In response to this question, I'd like to direct you to this forum response which goes over trunc-q in detail, and whether you should utilize that in your analysis.

With regards to max-ee, here is a great forum response with additional reading on the functionality which should also help inform your decision on whether to utilize this parameter.

Cheers,
Liz