PWT: Pilon With iTerations

Introduction

PWT (pronounced 'pout') is a wrapper for the genome polishing tool pilon. It uses a pair of Illumina reads in fastq format as well as a file of unpaired reads to improve the quality of a genome assembly. PWT will run pilon multiple times until pilon doesn't make changes to the assembly.

Method:

1. align reads to the assembly
2. run pilon
3. classify the changes
4. if there are changes, return to step 1 using the assembly produced by pilon

Installation

git clone https://github.com/rehrlich/pwt.git

Install dependencies:

conda install -c bioconda bwa

conda install -c bioconda samtools

conda install -c bioconda pilon

conda install pandas

By default, pilon doesn't use enough memory for many projects. Run 'which pilon' to find the pilon file, then change default_jvm_mem_opts to values that are appropriate for your system

Developed using:

bwa 0.7.17-r1188

samtools 1.9

pilon 1.23

pandas 0.25.3

Usage:

loop_pilon.py [-h] [-m MAX_ITERATIONS] [-t THREADS] --r1 R1 --r2 R2 -u UNPAIRED -f FASTA_PATH -o OUTDIR -s STRAIN_NAME

optional arguments:

-h, --help show this help message and exit

-m MAX_ITERATIONS, --max_iterations MAX_ITERATIONS Max number of times to run pilon (default: 10)

-t THREADS, --threads THREADS Number of threads to use (default: 16)

Required arguments:

--r1 R1 Path to r1 paired reads in fastq format

--r2 R2 Path to r2 paired reads in fastq format

-u UNPAIRED, --unpaired UNPAIRED Path to unpaired reads in fastq format

-f FASTA_PATH, --fasta_path FASTA_PATH Path to reference fasta

-o OUTDIR, --outdir OUTDIR Directory for output, must not exist

-s STRAIN_NAME, --strain_name STRAIN_NAME Nickname for the reference fasta

Format of outdir/changes.csv:

Each row corresponds to one change made by pilon

pilon_num - the iteration number

variant - the type of change made by pilon. Each change is explained below under 'Variant Types'

size_change - the change in the length of the sequence. A positive number means that Pilon increased the size of the sequence

source / destination - the source or destination from the pilon output. Source refers to the reference fasta; destination refers to the fasta produced by pilon

*_seq - the sequence. A period means the sequence is empty

*_ctg - the name of the contig

*_start - the start position of the variant

*_end - the end position of the variant

Variant Types:

Insertion - Pilon added one or more consecutive bases to the reference. Size change is positive

Deletion - Pilon removed one or more consecutive bases from the reference. Size change is negative

SNP - Pilon changed one base in the reference to a different base

Complex variant - Any change not listed above