Linking promoters to functional transcripts in small samples with nanoCAGE and CAGEscan

Large-scale sequencing projects have revealed an unexpected complexity in the origins, structures and functions of mammalian transcripts. Many loci are known to produce overlapping coding and noncoding RNAs with capped 5\u2032 ends that vary in size. Methods to identify the 5\u2032 ends of transcripts will facilitate the discovery of new promoters and 5\u2032 ends derived from secondary capping events. Such methods often require high input amounts of RNA not obtainable from highly refined samples such as tissue microdissections and subcellular fractions. Therefore, we developed nano\u2013cap analysis of gene expression (nanoCAGE), a method that captures the 5\u2032 ends of transcripts from as little as 10 ng of total RNA, and CAGEscan, a mate-pair adaptation of nanoCAGE that captures the transcript 5\u2032 ends linked to a downstream region. Both of these methods allow further annotation-agnostic studies of the complex human transcriptome