High-throughput total RNA sequencing in single cells using VASA-seq

Fredrik Salmen, Joachim De Jonghe, Tomasz S Kaminski, Anna Alemany, Guillermo E Parada, Joe Verity-Legg, Ayaka Yanagida, Timo N Kohler, Nicholas Battich, Floris van den Brekel, Anna L Ellermann, Alfonso Martinez Arias, Jennifer Nichols, Martin Hemberg, Florian Hollfelder, Alexander van Oudenaarden

Onderzoeksoutput: Bijdrage aan wetenschappelijk tijdschrift/periodieke uitgaveArtikelWetenschappelijkpeer review

35 Citaten (Scopus)


Most methods for single-cell transcriptome sequencing amplify the termini of polyadenylated transcripts, capturing only a small fraction of the total cellular transcriptome. This precludes the detection of many long non-coding, short non-coding and non-polyadenylated protein-coding transcripts and hinders alternative splicing analysis. We, therefore, developed VASA-seq to detect the total transcriptome in single cells, which is enabled by fragmenting and tailing all RNA molecules subsequent to cell lysis. The method is compatible with both plate-based formats and droplet microfluidics. We applied VASA-seq to more than 30,000 single cells in the developing mouse embryo during gastrulation and early organogenesis. Analyzing the dynamics of the total single-cell transcriptome, we discovered cell type markers, many based on non-coding RNA, and performed in vivo cell cycle analysis via detection of non-polyadenylated histone genes. RNA velocity characterization was improved, accurately retracing blood maturation trajectories. Moreover, our VASA-seq data provide a comprehensive analysis of alternative splicing during mammalian development, which highlighted substantial rearrangements during blood development and heart morphogenesis.

Originele taal-2Engels
TijdschriftNature Biotechnology
StatusE-pub ahead of print - 27 jun. 2022


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