Deep-RACE: Comprehensive search for novel ncRNAs associated to a specific locus

Abstract

Deep-RACE (or RACEseq) is a recently described method (Olivarius et al. BioTechniques 46(2):130–132, 2009) that applies next-generation sequencing to the Rapid Amplification of cDNA End (RACE) protocol to define the 5′ and 3′ ends of RNA transcripts. Conventional mapping of 5′ and 3′ ends is achieved by manually cloning the PCR product of RACE followed by Sanger sequencing; this process can become costly and time-consuming when investigating multiple transcripts. High-throughput sequencing of the RACE products streamlines this process by eliminating the need to manually cut bands from an agarose gel and to clone each product individually. Importantly, in addition to these advantages, next-generation sequencing can detect low abundance fragments that would be difficult to extract from gel and clone for Sanger sequencing. For these reasons, Deep-RACE is an ideal protocol for the comprehensive study of noncoding transcripts from both intergenic regions of the genome and from within the loci of protein coding gen s.