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Simple Summary  MicroRNAs are essential regulators of gene expression and potential non-invasive biomarker candidates for various human cancers as they can be detected in bodily fluids. Several tools have been developed to analyze small RNA-sequencing data; however, they have limitations and restrictions such as lack of optimal configuration, parameterization, and interoperability with other tools and platforms. miRGalaxy is an open-source, Galaxy-based framework for analyzing NGS data focusing on microRNAs and their sequence variants—isomiRs. Galaxy is a web-based platform for data-intensive biomedical research, allowing user-friendly analysis and accessibility to hundreds of tools. miRGalaxy is designed specifically for identifying and classifying human microRNAs and isomiRs, as well as detecting deregulated microRNAs and isomiRs between two test groups, summarized by output visualization. By examining the differential expression of individual isomiR species across samples, miRGalaxy can help discover novel biomarkers.    Abstract  Tools for microRNA (miR) sequencing data analyses are broadly used in biomedical research. However, the complexity of computational approaches still remains a challenge for biologists with scarce experience in data analytics and bioinformatics. Here, we present miRGalaxy, a Galaxy-based framework for comprehensive analysis of miRs and their sequence variants—miR isoforms (isomiRs). Though isomiRs are commonly reported in deep-sequencing experiments, their detailed structure complexity and specific differential expression (DE) remain not fully examined by the majority of the available analysis tools. miRGalaxy encompasses biologist-user-friendly tools and workflows dedicated to the analysis of the isomiR-ome and its complex behavior in various biological samples. miRGalaxy is developed as a modular, accessible, redistributable, shareable, and user-friendly framework for scientists working with small RNA (sRNA)-seq data. Due to its modular workflow, advanced users can customize the steps and tools for their needs. In addition, the framework provides an analysis report where the significant output results are summarized in charts and visualizations. miRGalaxy can be accessed via preconfigured Docker image flavor and a Toolshed installation if the user already has a running Galaxy instance. Over the last decade, studies on the expression of miRs and isomiRs in normal and deregulated tissues have led to the discovery of their potential as diagnostic biomarkers. The detection of miRs in biofluids further expanded the exploration of the miR repertoire as a source of liquid biopsy biomarkers. Here we show the miRGalaxy framework application for in-depth analysis of the sRNA-seq data from two different biofluids, milk and plasma, to identify, annotate, and discover specific differentially expressed miRs and isomiRs.

miRGalaxy: Galaxy-Based Framework for Interactive Analysis of microRNA and isomiR Sequencing Data