IsoSplitter

IsoSplitter identifies and validates alternative splicing (AS) sites de novo from long-read transcriptome sequencing data without requiring a reference genome.

Key Features:

• Reverse-Tracing and Validation: Employs a modified version of SIM4 to identify transcript split sites and enable reverse-tracing of AS events from long-read data.
• Quantification and Clustering: Quantifies each identified split site to reveal transcript diversity and groups putative isoforms into gene clusters.
• Optional Short-Read Alignment: Optionally aligns short reads to validate split sites by detecting unique junction reads and to reveal and quantify tissue-specific alternative splice isoforms.
• Assembly-free Long-Read Processing: Operates assembly-free from simple sequence files and leverages long-read transcriptome data to capture full-length RNA molecules and diverse splicing patterns.

Scientific Applications:

• Plant transcriptome AS discovery: Applied to multiple model and non-model plant species to identify alternative splicing events in diverse transcriptomes.
• Comparative evaluation in Arabidopsis thaliana: In evaluations using Arabidopsis thaliana data, identified more than twice as many AS events as AStrap.
• Reference concordance assessment: Reported 94.13% of IsoSplitter-predicted AS events were also identified by PASA, which requires a reference genome.

Methodology:

Begins with long-read transcriptome data, uses a modified SIM4 to identify transcript split sites and reverse-trace AS events, quantifies split sites, groups putative isoforms into gene clusters, and optionally aligns short reads to validate split sites via unique junction reads and quantify tissue-specific isoforms.