RoCK and ROI: single-cell transcriptomics with multiplexed enrichment of selected transcripts and region-specific sequencing

Abstract

Abstract Single-cell profiling technologies allow exploring molecular mechanisms that drive development, health, and disease. However, current methods still fall short of profiling single cell transcriptomes comprehensively, with one major challenge being high non-detection rates of specific transcripts and transcript regions. Such information is often crucial to understanding the biology of cells. Here, we introduce RoCK and ROI (Robust Capture of Key transcripts and Regions Of Interest), a scRNA-seq workflow encompassing two techniques. RoCKseq uses targeted capture to enrich for key transcripts, thereby supporting the detection and identification of cell types and complex phenotypes in scRNA-seq experiments. ROIseq directs a subset of reads to a specific region of interest via selective priming. Importantly, RoCK and ROI enables retrieval of specific sequence information without compromising overall single cell transcriptome information. We validate RoCK and ROI across diverse biological systems highlighting the versatility and showing the power of the method to retrieve critical transcriptomic features.