Bio-Tradis

A set of tools to analyse the output from TraDIS analyses

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License: GPL v3
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Contents

Introduction

The Bio::TraDIS pipeline provides software utilities for the processing, mapping, and analysis of transposon insertion sequencing data. The pipeline was designed with the data from the TraDIS sequencing protocol in mind, but should work with a variety of transposon insertion sequencing protocols as long as they produce data in the expected format.

For more information on the TraDIS method, see http://bioinformatics.oxfordjournals.org/content/32/7/1109 and http://genome.cshlp.org/content/19/12/2308.

Installation

Bio-Tradis has the following dependencies:

Required dependencies

There are a number of ways to install Bio-Tradis and details are provided below. If you encounter an issue when installing Bio-Tradis please contact your local system administrator. If you encounter a bug please log it here or email us at path-help@sanger.ac.uk.

Bioconda

Install conda and enable the bioconda channel.

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conda install-c bioconda biotradis=1.4.5=1

Docker

Bio-Tradis can be run in a Docker container. First install Docker, then install Bio-Tradis:

docker pull sangerpathogens/bio-tradis

To use Bio-Tradis use a command like this (substituting in your directories), where your files are assumed to be stored in /home/ubuntu/data:

docker run --rm -it -v /home/ubuntu/data:/data sangerpathogens/bio-tradis bacteria_tradis -h

Running the tests

The test can be run with dzil from the top level directory:

dzil test

Usage

For command-line usage instructions, please see the tutorial in the file “BioTraDISTutorial.pdf”. Note that default parameters are for comparative experiments, and will need to be modified for gene essentiality studies.

Bio-Tradis provides functionality to:

The functions are avalable as standalone scripts or as perl modules.

Scripts

Executable scripts to carry out most of the listed functions are available in the bin:

A help menu for each script can be accessed by running the script with no parameters.

Analysis Scripts

Three scripts are provided to perform basic analysis of TraDIS results in bin:

Internal Objects and Methods

Bio::Tradis::DetectTags

Bio::Tradis::AddTagsToSeq

Bio::Tradis::FilterTags

Bio::Tradis::RemoveTags

Bio::Tradis::Map

Bio::Tradis::TradisPlot

Bio::Tradis::RunTradis

Perl Programming Examples

You can reuse the Perl modules as part of other Perl scripts. This section provides example Perl code. Check whether file.bam contains TraDIS tag fields and, if so, adds the tags to the reads’ sequence and quality strings.

my $detector = Bio::Tradis::DetectTags(bamfile => 'file.bam');
if($detector->tags_present){
	Bio::Tradis::AddTagsToSeq(bamfile => 'file.bam', outfile => 'tradis.bam')->add_tags_to_seq;
}

Filter a FastQ file with TraDIS tags attached for those matching the given tag. Then, remove the same tag from the start of all sequences in preparation for mapping.

Bio::Tradis::FilterTags(
	fastqfile => 'tradis.fastq',
	tag => 'TAAGAGTGAC', 
	outfile => 'filtered.fastq'
)->filter_tags;
Bio::Tradis::RemoveTags(
	fastqfile => 'filtered.fastq',
	tag => 'TAAGAGTGAC', 
	outfile => 'notags.fastq'
)->remove_tags;

Create mapping object, index the given reference file and then map the fastq file to the reference. This will produce index files for the reference and a mapped SAM file named tradis_mapped.sam.

my $mapping = Bio::Tradis::Map(
	fastqfile => 'notags.fastq', 
	reference => 'path/to/reference.fa', 
	outfile => 'tradis_mapped.sam'
);
$mapping->index_ref;
$mapping->do_mapping;

Generate insertion site plot for only reads with a mapping score >= 50

Bio::Tradis::TradisPlot(mappedfile => 'mapped.bam', mapping_score => 50)->plot;

Run complete analysis on fastq files listed in file.list. This includes filtering and removing the tags allowing one mismatch to the given tag, mapping, BAM sorting and creation of an insertion site plot and stats file for each file listed in file.list.

Bio::Tradis::RunTradis(
	fastqfile => 'file.list', 
	tag => 'GTTGAGGCCA', 
	reference => 'path/to/reference.fa', 
	mismatch => 1
)->run_tradis;

License

Bio-Tradis is free software, licensed under GPLv3.

Feedback/Issues

Please report any issues to the issues page or email path-help@sanger.ac.uk

Citation

If you use this software please cite:

“The TraDIS toolkit: sequencing and analysis for dense transposon mutant libraries”, Barquist L, Mayho M, Cummins C, Cain AK, Boinett CJ, Page AJ, Langridge G, Quail MA, Keane JA, Parkhill J. Bioinformatics. 2016 Apr 1;32(7):1109-11. doi: 10.1093/bioinformatics/btw022. Epub 2016 Jan 21.