primer input issue for feature-classifier

[Bovern_Arromrak (Bovern Suchart Arromrak)]

Dear QIIME2 wizards,

I'm encountering a plugin error from the feature-classifier. I'm trying to create an amplicon-specific classifier for COX1gene with the following primers (based on Leray et al. (2013)):

1. mlCOIintF - GGWACWGGWTGAACWGTWTAYCCYCC
2. jgHCO2198 - TAIACYTCIGGRTGICCRAARAAYCA

When I ran the command code, it returned with the following error:
Plugin error from feature-classifier:
Invalid character in sequence: b'I'.

I understand the message but was not sure what bp can I use to replace the "I". It could be biased though if I just choose any bp to replace. Super grateeful if anyone could provide any advice and support for the matter.

Here is the code for more details:
qiime feature-classifier extract-reads
--i-sequences NCBIdata_derep1_seqs.qza
--p-f-primer GGWACWGGWTGAACWGTWTAYCCYCC
--p-r-primer TAIACYTCIGGRTGICCRAARAAYCA
--p-n-jobs 20
--p-read-orientation 'forward'
--o-reads NCBIdata_miinF-jgHCO_seqs.qza

Thank you very much.
Cheers,
Bovern

[SoilRotifer (Mike Robeson)]

Hi @Bovern_Arromrak,

If you search the forum you'll see this helpful bit of information:

Handling ambiguous nucleotides in database construction

Hi @devonorourke & @Nicholas_Bokulich, I have had similar issues with I characters when using vsearch and other similar tools. Keep in mind that I stands for the chemical Inosine / deoxyInosine, which you can order as part of your oligos (e.g. from IDT). That is, it functions literally as an ambiguous N base. So, wherever you find and I replace with N. For example, take a look at the CO1 blocking primer we made in our swine diet paper. As for the U --> T bit… In the past, when I put that set o…

-Cheers!

[Bovern_Arromrak (Bovern Suchart Arromrak)]

This is a great discussion and super helpful! Thank you so much , Mike!

Cheers,

Bovern