pySeqRNA: a python-based package for RNASeq data analysis
Today, massive amounts of data are generated by Next-Generation Sequencing (NGS) technologies. In recent years, many algorithms, statistical methods, and software tools have been developed to perform the individual analysis steps of various NGS applications. However, streamlined analysis remains a significant barrier to effectively utilizing the technology. We have developed a Python package (pySeqRNA) that allows fast, efficient, manageable, and reproducible RNA-Seq analysis. It effectively uses current software and tools with newly written Python scripts without confining users to a collection of pre-defined methods and environments by combining many command-line tools and custom Python scripts.
PySeqRNA requires a input file containing information of samples and input read files. Input template and example files here:
| # Project title/Information lines should start with # | ||||
|---|---|---|---|---|
| SampleName | Replication | Identifier | File1 | File2 |
| AddFull Sample Name Here | Add Replication Here | Add sample Identifier Here | Add Sample File Name Here | Add Reverese File here if Paired END |
Example input file:
| #Arabidopsis transcriptome study under high light stress | ||||
|---|---|---|---|---|
| SampleName | Replication | Identifier | File1 | File2 |
| GL0.5h1 | GL0.5h1 | GL0.5 | SRR6767632_001.fastq.gz | SRR6767632_002.fastq.gz |
| GLO.5h2 | GLO.5h2 | GL0.5 | SRR6767633_001.fastq.gz | SRR6767633_002.fastq.gz |
| GL6h1 | GL6h1 | GL6 | SRR6767634_001.fastq.gz | SRR6767634_002.fastq.gz |
| GL6h2 | GL6h2 | GL6 | SRR6767635_001.fastq.gz | SRR6767635_002.fastq.gz |
| GL12h1 | GL12h1 | GL12 | SRR6767636_001.fastq.gz | SRR6767636_002.fastq.gz |
| GL12h2 | GL12h2 | GL12 | SRR6767637_001.fastq.gz | SRR6767637_002.fastq.gz |
| GL24h1 | GL24h1 | GL24 | SRR6767639_001.fastq.gz | SRR6767639_002.fastq.gz |
| GL24h2 | GL24h2 | GL24 | SRR6767640_001.fastq.gz | SRR6767640_002.fastq.gz |
| GL48h1 | GL48h1 | GL48 | SRR6767642_001.fastq.gz | SRR6767642_002.fastq.gz |
| GL48h2 | GL48h2 | GL48 | SRR6767643_001.fastq.gz | SRR6767643_002.fastq.gz |
| GL72h1 | GL72h1 | GL72 | SRR6767644_001.fastq.gz | SRR6767644_002.fastq.gz |
| GL72h2 | GL72h2 | GL72 | SRR6767645_001.fastq.gz | SRR6767645_002.fastq.gz |
The pySeqRNA perform RNA-Seq analysis in two steps:
- Uniquely mapped reads
- Multimapped reads
This source code was developed in Linux, and has been tested on Linux and OS X. The main prerequisite is Python > 3.7. Following are the external dependencies:
- Flexbar – flexible barcode and adapter removal https://github.com/seqan/flexbar
- Trimmomatic: A flexible read trimming tool for Illumina NGS data http://www.usadellab.org/cms/?page=trimmomatic
- Trim Galore https://github.com/FelixKrueger/TrimGalore
- SortMeRNA [https://github.com/sortmerna/sortmerna] (https://github.com/sortmerna/sortmerna)
- STAR Aligner https://github.com/alexdobin/STAR
- HISAT2 http://daehwankimlab.github.io/hisat2/
- Bowtie2 https://github.com/BenLangmead/bowtie2
- Subread https://subread.sourceforge.net/
- HTSeq https://github.com/simon-anders/htseq
- Samtools https://github.com/samtools/samtools
- Bamtools https://github.com/pezmaster31/bamtools
- R Language https://cran.r-project.org/bin/windows/base/
- DESeq2 https://bioconductor.org/packages/release/bioc/html/DESeq2.html
- edgeR https://bioconductor.org/packages/release/bioc/html/edgeR.html
- Python 3 https://www.python.org/downloads/
The installation of pySeqRNA can be done in two ways:
-
Create a dedicated miniconda3 environment
Download pySeqRNA 0.2 from:
https://bioinfo.usu.edu/pyseqrna/download/pySeqRNA-0.2.tar.gz
Download the Miniconda installer:
https://docs.conda.io/en/latest/miniconda.html#linux-installers
Extract the downloaded file:
tar -xvzf pySeqRNA-0.2.tar.gz
cd pySeqRNA-0.2
chmod 755 INSTALL
./INSTALL
-
Create a docker image from docker file for cross-platform
Download pySeqRNA 0.2 from:
https://bioinfo.usu.edu/pyseqrna/download/pySeqRNA-0.2.tar.gz
Extract the downloaded file:
tar -xvzf pySeqRNA-0.2.tar.gz
cd pySeqRNA-0.2
docker build -t pyseqrna .
pyseqrna -h
Duhan N and Kaundal R. pySeqRNA: an automated Python package for RNA sequencing data analysis [version 1; not peer reviewed]. F1000Research 2020, 9(ISCB Comm J):1128 (poster) (https://doi.org/10.7490/f1000research.1118314.1)
Written by Naveen Duhan ([email protected]),
Kaundal Bioinformatics Lab, Utah State University,
Released under the terms of GNU General Public Licence v3
In case of technical problems (bugs etc.) please contact Naveen Duhan ([email protected])
For any Questions on the scientific aspects of the pySeqRNA-0.2 method please contact:
Rakesh Kaundal, ([email protected])
Naveen Duhan, ([email protected])