Demultiplexing dual-barcoded paired-end sequences from freshwater metabarcoding Illumina sequencing

Hi everyone!
I never used Qiime2 before and I was trying to understand how I can demultiplex my reads.
I have two fasq.gz file of paired-end reads from Illumina Miseq 2x150 sequencing. My reads have a sample 8 bases sample tag and 2/4 leading Ns. Both forward and reverse primers are tagged, so I used a combinatorial indexing approach. I can create a tsv with each sample-tag combination.

I have found various q/a regarding this topic so I am so sorry if this post is redundant but I can not figure out which plugin or external tool I can use.

(I work on freshwater eDNA, I have amplyfied the 12s for fishes and CO1 for invertebrates so I have already analyzed my reads using Obitools retriving the MOTUs, now I want to retrives the ASVs)

For importing, check out this portion of our importing tutorial. From the information that you have provided, that looks like the correct approach.

Since it sounds like you are already an experienced researcher, I think you will find this page very helpful for understanding the QIIME 2 workflow and relevant tools. The chart show here may also help with choosing which tool to use.

I think specifically you should look at using q2-cutadapt (demultiplexing + removal of non-biological sequences) and dada2 (denoising and ASV generation) to go from reads to ASVs with your data. An important thing with dada2 is that all non-biological sequences must be removed before denoising!

An off-topic reply has been split into a new topic: Now I am in the taxonomy assignment step. Do you have any tips or link to tutorials in which classify-consensus-vsearch is used?

Please keep replies on-topic in the future.

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