Ab initio reconstruction of cell type–specific transcriptomes in mouse reveals the conserved multi-exonic structure of lincRNAs

Abstract

High-throughput sequencing of total cellular RNA by RNA-Seq promises rapid reconstruction of spliced transcripts in a cell population. Guttman et al. accomplish this using only paired-end RNA-seq data and an unannotated genome sequence, and apply the method to better define many new, conserved long intergenic noncoding RNAs (lincRNAs). Massively parallel cDNA sequencing (RNA-Seq) provides an unbiased way to study a transcriptome, including both coding and noncoding genes. Until now, most RNA-Seq studies have depended crucially on existing annotations and thus focused on expression levels and variation in known transcripts. Here, we present Scripture, a method to reconstruct the transcriptome of a mammalian cell using only RNA-Seq reads and the genome sequence. We applied it to mouse embryonic stem cells, neuronal precursor cells and lung fibroblasts to accurately reconstruct the full-length gene structures for most known expressed genes. We identified substantial variation in protein coding genes, including thousands of novel 5′ start sites, 3′ ends and internal coding exons. We then determined the gene structures of more than a thousand large intergenic noncoding RNA (lincRNA) and antisense loci. Our results open the way to direct experimental manipulation of thousands of noncoding RNAs and demonstrate the power of ab initio reconstruction to render a comprehensive picture of mammalian transcriptomes.