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We describe a method, microarray analysis of differential splicing (MADS), for discovery of differential alternative splicing from exon-tiling microarray data. MADS incorporates a series of low-level analysis algorithms motivated by the "probe-rich" design of exon arrays, including background correction, iterative probe selection, and removal of sequence-specific cross-hybridization to off-target transcripts. We used MADS to analyze Affymetrix Exon 1.0 array data on a mouse neuroblastoma cell line after shRNA-mediated knockdown of the splicing factor polypyrimidine tract binding protein (PTB). From a list of exons with predetermined inclusion/exclusion profiles in response to PTB depletion, MADS recognized all exons known to have large changes in transcript inclusion levels and offered improvement over Affymetrix's analysis procedure. We also identified numerous novel PTB-dependent splicing events. Thirty novel events were tested by RT-PCR and 27 were confirmed. This work demonstrates that the exon-tiling microarray design is an efficient and powerful approach for global, unbiased analysis of pre-mRNA splicing.

MADS: A new and improved method for analysis of differential alternative splicing by exon-tiling microarrays