Hi all,I have sequenced my samples using the qiaseq 16/ITS kit. Which primers should I specify for trimming while running the pipeline?

The primer sequences should be revealed in the kit description? If not, write the company. If still no answer, you could try to figute that out by yourself by looking at the read starts, but thats not recommended. Also not recommended, but as last resort, skip primer removal with cutadapt and rather trim read starts to make sure primer sequences are removed, that will lead to less accurate results however.

thank you @Daniel Straub,The kit uses 3 pools of primers as seen in the attachment, now there is where I get stuck

Interesting protocol, but this is not directly supported by the pipeline, I think. Perhaps you could run it once for every individual primer pair. In each run sequences are selected for further processing by identification (and removal) of the primer pair.

Yes, multi-region analysis might apply here. But mixing bacteria and fungi isnt supported out-of-the-box, because there isnt a multi-region reference database with bacteria, archaea, and ITS regions readily available.

I was able to run the pipeline but now it fail in the final 3 steps with this error ERROR ~ Error executing process > 'NFCORE_AMPLISEQAMPLISEQSUMMARY_REPORT (1)' Caused by: Process

NFCORE_AMPLISEQ:AMPLISEQ:SUMMARY_REPORT

input file name collision -- There are multiple input files for each of the following file names: Metadata.tsv Container: /tank/lmapunda/ARI/16s/work/singularity/depot.galaxyproject.org-singularity-mulled-v2-b2ec1fea5791d428eebb8c8ea7409c350d31dada-a447f6b7a6afde38352b24c30ae9cd6e39df95c4-1.img Tip: view the complete command output by changing to the process work dir and entering the command

cat .command.out

-- Check '.nextflow.log' file for details ERROR ~ Pipeline failed. Please refer to troubleshooting docs: https://nf-co.re/docs/usage/troubleshooting -- Check '.nextflow.log' file for details

Thats a new error, as far as I know. To make the pipeline succeed, you can skip the report generation with --skip_report. But the report is helpful, so its not desirable to skip it. If you provide more information, such as input parameters and the .nextflow.log of the run that failed, then I might be able to find the cause.

I meant to mention here, but my message was never sent, that if you run the pipeline several times you need to use different output directories for each run. Otherwise output files will overwrite each other. This probably has nothing to do with the above however.