Latch Verified Bulk-RNA Seq toolkit: a cloud-based suite of workflows for bulk RNA-seq quality control, analysis, and functional enrichment

Abstract

Background Analysis of high-throughput bulk RNA-sequencing (RNA-seq) data reveals changes in gene expression between diverse conditions. Many tools have emerged to quality control RNA-seq reads, quantify expression levels, conduct functional enrichment among differentially expressed genes, or identify differential RNA splicing. However, unified toolkits for conducting these analyses are lacking. Moreover, existing software does not use cloud-based platforms that provide the necessary storage and computational resources to process RNA-seq data or intuitive graphical interfaces for easy use by experimental and computational scientists. Results To address these challenges, we introduce the Latch Verified Bulk RNA-Seq (LVBRS) toolkit, a flexible suite of programs packaged into a single workflow coupled with a graphical user interface for conducting quality control, transcript quantification, differential splicing, differential expression analysis, and functional enrichment analyses. For functional enrichment, the LVBRS toolkit supports three databases - Gene Ontology, KEGG Pathway, and Molecular Signatures database - capturing diverse functional information. We demonstrate the utility of the LVBRS toolkit by reanalyzing a publicly available dataset examining the impact of severe and mild models of hypoxia - induced by Cobalt (II) Chloride (CoCl2) and oxyquinoline treatment, respectively - on a human colon adenocarcinoma cell line. Our analyses reveal CoCl2 treatment results in more differentially expressed genes, recapitulating previously reported results that CoCl2 models more severe hypoxia. Moreover, including alternative splicing and functional enrichment analysis using a greater breadth of functional databases revealed additional biological insights - such as greater alternative splicing in the CoCl2 condition and differentially expressed DNA repair pathways. These results demonstrate the LVBRS toolkit's efficacy in facilitating biological insights from bulk RNA-seq data. Conclusions The LVBRS toolkit offers a robust unified framework for processing and analyzing Bulk RNA-Seq experiments. The easy-to-use graphical user interface will enable diverse scientists to conduct high-throughput bulk RNA-Seq analysis efficiently. Our aim is that the LVBRS toolkit will help streamline bulk RNA-seq workflows and facilitate deriving biologically meaningful insights from bulk RNA-seq data. The source code is freely available under the MIT license and hosted on the LatchBio Console (https://console.latch.bio/se/bulk-rnaseq), complete with documentation (https://latch.wiki/bulk-rna-seq-end-to-end).

Assessing RNA-Seq Workflow Methodologies Using Shannon Entropy

“…To address biological and methodological variations within a user-friendly framework, an expanding array of open-access semiautomated pipelines is emerging online. Examples include iDEP [18], LVBRS [19], RNAseqChef [20], and NormSeq [21]. It has been observed that achieving consensus among pipelines enhances the diagnostic accuracy of differentially expressed genes (DEGs), suggesting that combining diverse methodologies can yield more robust results [22].…”