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HMP tutorial 454 reads, Human Microbiome Project samples |
Part 2. Implementing the pipeline script
Part 1. UPARSE pipeline from reads to OTU table
This tutorial uses reads of 11 samples deposited in archive SRR058098 by the Human Microbiome Project. I took a random subset of 5% of the reads, giving 19,735 FASTQ records. See Part 2 for more information about the reads and how the UPARSE pipeline was implemented.
Download this archive:
Tutorial files: hmptut.tar.gz
I'll assume your downloaded file is in ~/Downloads, if you downloaded to a different path then replace as needed below.
Make a top-level directory for the tutorials, change to that directory and extract the data files using tar for the tutorial files. See tutorial directories for description of subdirectories.
Unzip for the data files:
mkdir -p ~/tutorials
cd ~/tutorials
tar -zxvf ~/Downloads/hmptut.tar.gz
Set the $usearch environment variable to the path name of your usearch binary file.
Create the utax database by running the setup_utax.bash script, like this.
cd ~/tutorials/hmptut/scripts
./setup_utax.bash
Notice the dot and slash (./) before setup_utax.bash. This tells the shell to look for the command file (script or binary) in your current directory (dot means current directory). This is needed if the current directory is not in your PATH. Tutorial scripts always assume that they are being run like this, i.e. from inside the scripts/ subdirectory.
The setup_utax.bash script uses curl to fetch the data. Some systems don't have curl in which case you can use wget. There is a wget command in the script which is commented out so it's a simple edit of the script to comment out curl instead.
The UPARSE pipeline is implemented in the run_uparse.bash script. Run it like this:
cd ~/tutorials/hmptut/scripts
./run_uparse.bash
This should reproduce the pre-computed files in the hmptut/out/ directory.
The most important output files are the OTU tables, which are named outtab.txt (QIIME classic format), otutab.json (BIOM format) and otutab.mothur (mothur "shared" file format) . You can use these to perform further analysis in QIIME or mothur.