Linearly Amplified Single-Stranded RNA-Derived Transcriptome Sequencing (LAST-seq)

Abstract

Single-cell RNA sequencing (scRNA-seq) stands as a cutting-edge technology widely used in biological and biomedical research. Existing scRNA-seq methods rely on reverse transcription (RT) and second-strand synthesis (SSS) to convert RNA to cDNA before amplification. However, these methods often suffer from limited RT/SSS efficiency, which compromises the sensitivity of RNA detection. Here, we develop a new method, linearly amplified single-stranded RNA-derived transcriptome sequencing (LAST-seq), which directly amplifies the original single-stranded RNA without prior RT and SSS and offers high-sensitivity RNA detection and a low level of technical noise in single-cell transcriptome analysis. LAST-seq has been applied to quantify transcriptional bursting kinetics in human cells, advancing our understanding of chromatin organization’s role in regulating gene expression.