Which sequence files should be used for demultiplexing and DADA2 in QIIME 2?

User Support
1

Hi everyone,

I’m working with multi-sample 16S/ITS sequencing data in QIIME 2 and have a question about the qiime tools import step.

Specifically, I’m unsure which type of sequence files should be used as input:

1.raw FASTQ files;

2.Per-sample raw FASTQ files after primer trimming;

3.A single FASTQ file combining all samples;

4.Per-sample quality-filtered FASTA files;

My main questions is:

When importing for demultiplexing, which file should I use?

qiime tools import \

--type 'SampleData[PairedEndSequencesWithQuality]' \

--input-path

--input-format PairedEndFastqManifestPhred33V2 \

--output-path

qiime demux summarize

--i-data

--o-visualization

qiime dada2 denoise-paired \

--i-demultiplexed-seqs demux-paired-end.qza \\

--p-trunc-len-f  

--p-trunc-len-r  \\

--p-trim-left-f  \\

--p-trim-left-r  \\

--p-max-ee 5  \\

--p-n-threads  \\

--o-representative-sequences representative-sequences.qza \\

--o-table table.qza \\

--o-denoising-stats denoising-stats.qza \\

--verbose > dada2_log 2>&1
2

Hey @CWZ,

From the two commands you've included, it looks like you did already import your data and got your demux summary as well as your feature table?

Disregarding the other commands you included, to address your main question - if you are planning to demultiplex your data within QIIME 2 you would use your raw FASTQ files. The import format you use will depend on what data you received from your sequencing center. I'd recommend reviewing our guide on importing data into QIIME 2, this will provide some helpful examples on importing depending on the type of raw data you have:

Cheers :lizard:

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