High-throughput quantification of splicing isoforms

• DOI: 10.1261/rna.1877010
• Publication Date: 2009-12-29T01:54:20Z
• Authors: Jean-Philippe Bro...; Jean-François Lucier; Elvy Lapointe; Mathieu Durand; Daniel Gendron; Julien Gervais-Bird; Karine Tremblay; Jean-Pierre Perre...; Sherif Abou Elela
• Keywords: Protein isoform

Most human messenger RNAs (mRNAs) are alternatively spliced and many exhibit disease-specific splicing patterns. However, the contribution of most events to development maintenance diseases remains unclear. As alternative diagnosis prognosis is becoming increasingly recognized, it becomes important develop precise methods quantify abundance these isoforms in clinical samples. Here we present a pipeline for real-time PCR annotation (RASE) that allows accurate identification large number tissues. The RASE automatically designed specific primer pairs 81% all NCBI build 36 database. Experimentally, majority primers resulted isoform-specific amplification suitable quantification cell lines or formalin-fixed, paraffin-embedded (FFPE) RNA extract. Using this now possible rapidly identify isoform signatures different types tissues validate complete sets data generated by microarray expression profiling deep sequencing techniques.