This repository contains the workflow used to analyze ITSxpress in the paper

Software Authors

• Adam R. Rivers
• Kyle Weber

Paper

Rivers, A.R., Weber. K.C., Gardner T. G., Liu, S., and Armstrong, S.D. 2018. ITSxpress: software to rapidly trim internally transcribed spacer sequences with quality scores for marker gene analysis. F1000 Research. In prep.

Setup

• Create an Conda environment. For instructions on installing Anaconda or Miniconda see this page: https://conda.io/miniconda.html
• The workflow is designed to be run on a HPC with a SLURM scheduler

# Create a new conda test environment
conda create --name itsxpresstestenv python=3.6 pandas

# activate the new environment
source activate itsxpresstestenv

# Install itsxpress and ITSx into the new environment
conda install -c bioconda itsxpress=1.6.1 itsx=1.1b

Scripts

The workflow is broken into two parts. The first script, run1.sh distributes tasks with the SLURM scheduler and creates the trimmed fastq's and timing data. Specifically it:

• merges reads and creates fasta files for ITSx
• De-replicates fasta files to estimate the proportion of unique reads
• Creates a CSV file summarizing the number of unique reads with dedup.py
• Submits the script its_samples.sh for the ITS1 and ITS2 regions to calculate the speedup from using ITSxpress
• Submits its_threads.sh sbatch jobs for the its1 and its2 regions to understand how the program scales with more cores
• Submits the script its_100_samples.sh to compare alignment similarity at 100% identity clustering

The second script, run2.sh, summarizes the data and creates graphs. specifically it:

• Creates CSV's summarizing the timing data for the sample and thread experiments
• Runs pairwise alignment on the ITSx and ITSxpress results summarizing the differences
• Creates the plots in the paper using itsxpress_plots.R

Directory structure

.
|-- data
|   |-- its1
|   |   |-- r1
|	  |	  |-- r2
|	  |  	`-- readme.txt
|   |-- its1_derep
|   |-- its2_merged
|	  |-- its1_fasta
|	  |-- its2
|   |   |-- r1
|  	|	  |-- r2
|  	|	  `-- readme.txt
| 	|-- its2_derep
|   |-- its2_merged
|	  `-- its2_fasta
|--scripts
|   |-- run1.sh
|   |-- run2.sh
|   |-- derep.py
|   |-- test_samples.sh
|   |-- test_100_samples.sh
|   |-- test_threads.sh
|   |-- compare_alignments.py
|   |-- parsetiming.py
|   `-- plots.R
|-- output
|-- dedup_out
|-- analysis
`-- README.md