Overview

stepRNA is a RNA-seq read processor, based on bowtie2, that will align small RNA-seq query reads (passenger sequences) to reference sequences (siRNAs) and output information about the type and length of overhangs uncovered. It was originally developed for uncovering Dicer processing signatures but is not limited to just this.

Table of Contents:

• Reporting Issues
• Installation
• Use
• Example
• News
• Licence
• Additional Information

Reporting Issues

Please report any issues to the stepRNA GitHub page or via email:

• https://github.com/Vicky-Hunt-Lab/stepRNA/issues
• benmurcott96@gmail.com or bmm41@bath.ac.uk (Ben Murcott)
• v.l.hunt@bath.ac.uk (Vicky Hunt)

Installation

In order for stepRNA to run you must have:

• Bowtie2 >= v2.3.4 (see BOWTIE2 website for more details)
• biopython >= v0.3.0 (automatically installed with pip)
• numpy >= v1.19.0 (autoamtically installed with pip)
• pysam >= v0.16.0.1 (autoamtically installed with pip)

To install with pip (recommended):

pip install stepRNA

To upgrade to v1.0.6

pip install --upgrade stepRNA

If this doesn't work due to non-root access issues see Installing into a Virtual Environment

To install from source:

1. Download stepRNA-1.0.6.tar.gz
2. Unzip it
3. Move into stepRNA-1.0.6 repository
4. Run the installation

tar -xvzf stepRNA-1.0.6.tar.gz

cd stepRNA-1.0.6

python3 setup.py install

Installing into a Virtual Environment

This can be useful if root privalages are not available to the user.

1. Create a virutal environment (recommend using virtualenv):

pip install virtualenv

2. Create and activate the environment:
3. Run the installation

virtualenv stepRNA_env

source /stepRNA_env/bin/activate

pip install stepRNA

Use:

See the documentation for a detailed description on how to use stepRNA (MANUAL)

The quickest way to use stepRNA:

stepRNA --reference REFERENCE --reads QUERY

This will align the reads to the reference sequences and output into the current diretory using the READS filename as the prefix. REFERENCE and QUERY must have unique FASTA headers (if not use -u)

Helpful options:

• --name can be used to customise the prefix name
• --directory can be used to specify an output directory

Example:

Using the reads from stepRNA/example_data/experimental_data:

stepRNA --reference stepRNA/example_data/experimental_data/26G_embryo.fa --reads stepRNA/example_data/experimental_data/LF_embryo.fa --directory stepRNA_example

This will create a new direcotry called stepRNA_example that contains:

• LF_embryo_AligmentFile/; a directory containing BAM files for each of the overhang lengths
• CSVs; containing count information

See the MANUAL for more information

News

Latest release notes:

Version 1.0.6 - 28 Feb, 2023

• Update to setup.py to include alive_progress

See NEWS for historical updates of release notes

Licence

stepRNA is licensed under the MIT license. See LICENSE file for details.

Additional Information

For more information:

• Go to FAQs to see commonly asked quesitons
• Look at the USER MANUAL to see detailed instructions and all of the available options

If you use stepRNA in your work please cite:

stepRNA: Identification of Dicer cleavage signatures and passenger strand lengths in small RNA sequences (Murcott et al. 2022)