RNA-seq has become the gold standard for mapping transcriptomes, profiling changes in splicing and measuring gene expression levels. The most widely used method for RNA-seq library construction is the dUTP approach. Although this approach provides high-quality strand-specific RNA-seq profiles, it involves generation of a single library for a single sample. As such, this method is time consuming and expensive to perform on many samples, limiting its utility for applications that require profiling hundreds or thousands of individual samples, such as whole-transcriptome profiling of cancer samples or screening the effects of genetic perturbations on gene expression.
Here researchers from the California Institute of Technology report RNAtag-Seq, a method for generating a single RNA-seq library containing large numbers of RNA samples that are barcoded and pooled before library construction. Barcoding in RNAtag-Seq is achieved through direct ligation of adaptors to RNA, enabling strand-specific, quantitative sequencing of full-length transcripts in diverse prokaryotic and eukaryotic species at a quality highly comparable to that of the well-established dUTP method for single-sample library construction.