Multi - marker analysis 16S, 18S, ITS, tufA

I am analysing a dataset where multiple markers have been amplified: 16S, 18S, ITS, tufA.

Can I run qiime2 pipeline on all of those or only for 16S and ITS?


Absolutely! QIIME 2 is in no ways specific to 16S and ITS. You just need to use the appropriate reference databases for those markers at relevant steps, e.g., during taxonomy classification, and I expect you will have slightly different processing pipelines for each of these.

Check around this forum — lots of people using 18S have posted discussion here, you may get some ideas about what others are doing.

Yes, I found something but it’s not clear if nowadays this issue has been resolved the post is dated middle 2017.

Then, is there any way to demultiplex my samples (parse them amplicon by amplicon within QIIME2) which have same indexs for different molecular markers?

Many thanks

I was meaning this topic: Multi-gene amplicon sequences with dada2

Hi @Fra,
Sorry for the delay. The issue regarding multiple markers in that post is not whether QIIME 2 can analyze these markers, but that all 4 markers share the same barcodes, so cannot be demultiplexed from one another. Sounds like that is the same setup you have.

That’s a fairly peculiar setup so the challenges are not unique to QIIME 2. You would need to figure out a way to demultiplex these — probably with a custom script — prior to importing to QIIME 2.

If primers and barcodes are present in the reads, you can use the primers as a part of the barcode, hence distinguishing individual samples AND individual marker genes.

You could also process in QIIME and then use something like qiime quality-control exclude-seqs to separate out sequences later on, but that could be tricky.

The best thing to do is to NOT multiplex multiple marker genes with the same barcodes to begin with!

Once you have those marker genes separated out from each other, analysis of any marker genes in QIIME 2 should be easy.

I hope that helps!

Thanks for the reply Nicholas.

I worked it out using a script able to sort each molecular marker in different .fastq files.


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