Metabolomics and Systems Biology

Through alternative splicing, most human genes express multiple isoforms that often differ in function. To infer isoform regulation from high-throughput sequencing of cDNA fragments (RNA-seq), a pipeline is developed to detect differential alternative splicing events between RNA-Seq samples of treatment and control conditions. It is based on a metric defined as splicing ratio (SR) which was used in some splicing-related studies (SR Splicing is flexible to handle different types of study design. Either control group or treatment group can have single or replicated/pooled samples. It can analyze all major types

of alternative splicing patterns and use RNA-Seq reads and use RNA-Seq reads mapped to splice junctions. By comparing with other commonly used software, the performance of SR Splicing is evaluated from three aspects: sensitivity, accuracy, and validation. SR Splicing has comparable sensitivity and higher accuracy. Experimental validation using qRT-PCR (quantitative real time polymerase chain reaction) confirmed a selected set of splicing events that are significantly changed in Pten knock out data of the mouse MEF cell line, demonstrating the utility of the approach applied on experimental biological data sets.SR Splicing used different statistics and less filtering to return a list of significantly changed splicing events, with associated p values and false discovery corrections. It includes detailed information on the detected splicing differences such as which exon/ junctions are involved, alternative splice type (skipped exon, mutually exclusive exons, retained intron, alternative 5' splice site, and alternative 3' splice site), magnitude of difference, and coverage.