DADA2- Samples from Multiple MiSeq Sequencing Runs

I am working on implementing a QIIME2 pipeline using the DADA2 plugin. Since ours is a high-throughput sequencing facility, we have 1000s of samples in one project and these samples come from multiple MiSeq Runs.
Usually for our Mothur-based Pipeline, we put the FastQs for these samples from separate sequencing runs together using the Manifest File provided and then run them through the pipeline.
I am not certain if I can utilize the same process with QIIME2/DADA2-plugin as well? (Means if it’s possible to analyze samples together that are from multiple MiSeq runs and push them through the QIIME2/DADA2 or would we need to analyze samples from one run at a time and then put the OTU/feature-table results together in the end?
Tl;dr-Whats the best strategy to do analysis for large sample sizes that are from multiple MiSeq sequencing runs?

Hello @Shalabh_Suman!

In QIIME 2, the pipeline requires you to import all of your files and condense them into a demux.qza file. For questions on how to import data into QIIME 2, please refer to the importation tutorial.
Once this has been done, one can run DADA 2 once on that demux.qza file to prepare a single table.qza file, and representative-sequences.qza file. For help on how to do this, please refer to this tutorial.

NOTE: If you are running paired data, the DADA2 step will be slightly different, but information for that can be found here.

Hoe that helps!
Sirtaj Singh

P.S. The DADA2 step is known to take some time on QIIME 2. If it doesn’t spit out an error and hasn’t finished completing, just give it some time. Just be sure that there is enough disc space for the output files, and that you have given it enough RAM to run smoothly.


Hi @Shalabh_Suman,

To expand on @Sirtaj-Singh’s answer a bit, you probably shouldn’t mix multiple runs.
DADA2 is meant to operate on a single run at a time (it builds an error model for each run, so mixing runs will just confuse the model and the way it picks out the ASVs).

Once you are familiar with the moving pictures tutorial, you should check out this one, which specifically deals with managing multiple runs.

Let me know if you have any other questions about this!


An off-topic reply has been split into a new topic: Multiple runs and splitting up an existing demux artifact

Please keep replies on-topic in the future.

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