This tutorial was developed at the Microbiome Bioinformatics With QIIME 2 workshop in Las Vegas, NV in June 2017. This has been tested with QIIME 2 2017.6, but is not guaranteed to work outside of this workshop. If you're looking for the QIIME 2 documentation, see https://docs.qiime2.org. If you're interested in attending a QIIME 2 workshop, see https://workshops.qiime2.org.
Fit a classifier for the UNITE Database
wget https://unite.ut.ee/sh_files/sh_qiime_release_20.11.2016.zip
wget https://dl.dropboxusercontent.com/u/2868868/temp/sh_refs_qiime_ver7_99_20.11.2016.fasta
wget https://dl.dropboxusercontent.com/u/2868868/temp/sh_taxonomy_qiime_ver7_99_20.11.2016.txt
unzip sh_qiime_release_20.11.2016.zip
cd sh_qiime_release_20.11.2016
qiime tools import \
--type FeatureData[Sequence] \
--input-path sh_refs_qiime_ver7_99_20.11.2016.fasta \
--output-path unite-ver7-99-seqs-20.11.2016.qza
qiime tools import \
--type FeatureData[Taxonomy] \
--input-path sh_taxonomy_qiime_ver7_99_20.11.2016.txt \
--output-path unite-ver7-99-tax-20.11.2016.qza \
--source-format HeaderlessTSVTaxonomyFormat
qiime feature-classifier fit-classifier-naive-bayes \
--i-reference-reads unite-ver7-99-seqs-20.11.2016.qza \
--i-reference-taxonomy unite-ver7-99-tax-20.11.2016.qza \
--o-classifier unite-ver7-99-classifier-20.11.2016.qza
Download an ITS mock community from mockrobiota
wget -O "mock-25-sample-metadata.tsv" https://raw.githubusercontent.com/caporaso-lab/mockrobiota/master/data/mock-25/sample-metadata.tsv
wget https://s3-us-west-2.amazonaws.com/mockrobiota/latest/mock-25/mock-forward-read.fastq.gz
Create a file, fastqmanifest.csv, with the following two lines:
sample-id,absolute-filepath,direction
Mock.1,$PWD/mock-forward-read.fastq.gz,forward
qiime tools import \
--type SampleData[SequencesWithQuality] \
--input-path fastqmanifest.csv \
--output-path demux.qza \
--source-format SingleEndFastqManifestPhred33
qiime demux summarize \
--i-data demux.qza \
--o-visualization demux.qzv
In the following denoise-single command, we want to provide qiime dada2 denoise-single, but a bug was discovered that currently prevents that.
qiime dada2 denoise-single \
--i-demultiplexed-seqs demux.qza \
--p-trim-left 10 \
--p-trunc-len 140 \
--o-representative-sequences rep-seqs.qza \
--o-table table.qza
qiime feature-classifier classify-sklearn \
--i-classifier unite-ver7-99-classifier-20.11.2016.qza \
--i-reads rep-seqs.qza \
--o-classification taxonomy.qza
qiime taxa tabulate \
--i-data taxonomy.qza \
--o-visualization taxonomy.qzv
qiime taxa barplot \
--i-table table.qza \
--i-taxonomy taxonomy.qza \
--m-metadata-file mock-25-sample-metadata.tsv \
--o-visualization taxa-bar-plots.qzv
The expected composition of this community can be found here.