RNA-SeqEZPZ: A Point-and-Click Pipeline for Comprehensive Transcriptomics Analysis with Interactive Visualizations

Abstract

RNA-Seq analysis has become a routine task in numerous genomic research labs, driven by the reduced cost of bulk RNA sequencing experiments. These generate billions of reads that require accurate, efficient, effective, and reproducible analysis. But the time required for comprehensive analysis remains a bottleneck. Many labs rely on in-house scripts, making standardization and reproducibility challenging. To address this, we developed RNA-SeqEZPZ, an automated pipeline with a user-friendly point-and-click interface, enabling rigorous and reproducible RNA-Seq analysis without requiring programming or bioinformatics expertise. For advanced users, the pipeline can also be executed from the command line, allowing customization of steps to suit specific requirements. This pipeline includes multiple steps from quality control, alignment, filtering, read counting to differential expression and pathway analysis. We offer two different implementations of the pipeline using either (1) bash and SLURM or (2) Nextflow. The two implementation options allow for straightforward installation, making it easy for individuals familiar with either language to modify and/or run the pipeline across various computing environments. RNA-SeqEZPZ provides an interactive visualization tool using R shiny to easily select the FASTQ files for analysis and compare differentially expressed genes and their functions across experimental conditions. The tools required by the pipeline are packaged into a Singularity image for ease of installation and to ensure replicability. Finally, the pipeline performs a thorough statistical analysis and provides an option to perform batch adjustment to minimize effects of noise due to technical variations across replicates. RNA-SeqEZPZ is freely available and can be downloaded from https://github.com/cxtaslim/RNA-SeqEZPZ.