# An integrative R package for analysing alternative splicing using RNAseq ## Authors Estefania Mancini, Javier Iserte, Marcelo Yanovsky, Ariel Chernomoretz ## Introduction Alternative splicing (AS) is a common mechanism of post-transcriptional gene regulation in eukaryotic organisms that expands the functional and regulatory diversity of a single gene by generating multiple mRNA isoforms that encode structurally and functionally distinct proteins. The development of novel high-throughput sequencing methods for RNA (RNAseq) has provided a powerful means of studying AS under multiple conditions and in a genome-wide manner. However, using RNAseq to study changes in AS under different experimental conditions is not trivial. In this vignette, we describe how to use ASpli, an integrative and user-friendly R package that facilitates the analysis of changes in both annotated and novel AS events. This package combines statistical information from exon, intron, and splice junction differential usage (p-value, FDR), with information from splice junction reads to calculate differences in the percentage of exon inclusion Delta-PSI and intron retention Delta-PIR). The proposed methodology reliably reflect the magnitude of changes in the relative abundance of different annotated and novel AS events. This method can be used to analyze both simple and complex experimental designs involving multiple experimental conditions. ## Getting started ### Installation ASpli is available at Bioconductor site and can be downloaded using **BiocManager::install()**: if (!requireNamespace("BiocManager", quietly=TRUE)) install.packages("BiocManager") BiocManager::install("ASpli") library(ASpli) **BiocManager::install()** will take care of installing all the packages that ASpli depends on e.g. edgeR, GenomicFeatures, GenomicRanges, GenomicAlignments, Gviz, and other R package required. Some packages depend on operating system packages, like **curl**, that are not installed automatically, and should be installed by the user.