tourmaline | Amplicon sequence processing workflow using QIIME | Genomics library

Tourmaline provides Snakemake rules for DADA2 (single-end and paired-end) and Deblur (single-end). For each type of processing, there are four steps:. Steps 2–4 have unfiltered and filtered modes, the difference being that in the taxonomy step of filtered mode, undesired taxonomic groups or individual sequences from the representative sequences and feature table are removed. The diversity and report rules are the same for unfiltered and filtered modes, except the output goes into separate subdirectories.
the denoise rule imports FASTQ data and runs denoising, generating a feature table and representative sequences;
the taxonomy rule assigns taxonomy to representative sequences;
the diversity rule does representative sequence curation, core diversity analyses, and alpha and beta group significance; and
the report rule generates an HTML report of the outputs plus metadata, inputs, and parameters. Also, the report rule can be run immediately to run the entire workflow.
Before you download the Tourmaline commands and directory structure from GitHub, you first need to install QIIME 2, Snakemake, and the other dependencies of Tourmaline. Two options are provided: a native installation on a Mac or Linux system and a Docker image/container. See the Install page for more details.
Install Docker Desktop (Mac or Windows) or Docker (Linux) from Docker.com.
Increase the memory to 8 GB or more (Preferences -> Resources -> Advanced -> Memory).
Download the Docker image from DockerHub (command below).
Run the Docker image (command below).
copy metadata and manifest files to your container;
create symbolic links from your container to your FASTQ files and reference database;
copy your whole Tourmaline directory out of the container when the run is completed (alternatively, you can clone the Tourmaline directory inside the mounted volume).
If this is your first time running Tourmaline, you'll need to set up your directory. Simplified instructions are below, but see the Wiki's Setup page for complete instructions.
Prepare reference database: Option 1: Put taxonomy and FASTA files in 00-data. Check that filenames match those in config.yaml. Option 2: Put imported QIIME 2 archives in 01-imported. Check that filenames match those in config.yaml.
Prepare sequence data: Option 1: Edit FASTQ manifests in 00-data so file paths point to your .fastq.gz files (they can be anywhere on your computer) and sample names match the metadata file. You can use a text editor such as Sublime Text, nano, vim, etc. Check that manifest filenames match those in config.yaml. Option 2: Put your pre-imported FASTQ .qza files in 01-imported. Check that .qza filenames match those in config.yaml.
Edit metadata file metadata.tsv to contain your sample names and any relevant metadata for your samples. You can use a spreadsheet editor like Microsoft Excel or LibreOffice, but make sure to export the output in tab-delimited format.
Edit configuration file config.yaml to change input file names (or preferably, rename your input files to match the defaults), DADA2/Deblur parameters (sequence truncation/trimming, sample pooling, chimera removal, etc.), rarefaction depth, taxonomic classification method, and other parameters. This YAML (yet another markup language) file is a regular text file that can be edited in Sublime Text, nano, vim, etc.
Go to Run Snakemake.