Droplet-based Single-cell Total RNA-seq Reveals Differential Non-Coding Expression and Splicing Patterns during Mouse Development

Salmen, F.; De Jonghe, J.; Kaminski, T. S.; Alemany, A.; Parada, G.; Verity-Legg, J.; Yanagida, A.; Kohler, T. N.; Battich, N.; van den Brekel, F.; Ellermann, A. L.; Arias, A. M.; Nichols, J.; Hemberg, M.; Hollfelder, F.; van Oudenaarden, A.

doi:10.1101/2021.09.15.460240

bioRxiv · 10.1101/2021.09.15.460240

Droplet-based Single-cell Total RNA-seq Reveals Differential Non-Coding Expression and Splicing Patterns during Mouse Development

Salmen, F.·De Jonghe, J.·Kaminski, T. S.·Alemany, A.·Parada, G.·Verity-Legg, J.·Yanagida, A.·Kohler, T. N.·Battich, N.·van den Brekel, F.·Ellermann, A. L.·Arias, A. M.·Nichols, J.·Hemberg, M.·Hollfelder, F.·van Oudenaarden, A.

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Abstract

In recent years, single-cell transcriptome sequencing has revolutionized biology, allowing for the unbiased characterization of cellular subpopulations. However, most methods 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. Additionally, most workflows do not sequence the full transcript hindering the analysis of alternative splicing. We therefore developed VASA-seq to detect the total transcriptome in single cells. VASA-seq is compatible with both plate-based formats and droplet microfluidics. We applied VASA-seq to over 30,000 single cells in the developing mouse embryo during gastrulation and early organogenesis. The dynamics of the total single-cell transcriptome result in the discovery of novel cell type markers many based on non-coding RNA, an in vivo cell cycle analysis and an improved RNA velocity characterization. Moreover, it provides the first comprehensive analysis of alternative splicing during mammalian development.

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Salmen, F., De Jonghe, J., Kaminski, T. S., Alemany, A., Parada, G., Verity-Legg, J., Yanagida, A., Kohler, T. N., Battich, N., van den Brekel, F., Ellermann, A. L., Arias, A. M., Nichols, J., Hemberg, M., Hollfelder, F., van Oudenaarden, A.. 2021-09-15. Droplet-based Single-cell Total RNA-seq Reveals Differential Non-Coding Expression and Splicing Patterns during Mouse Development. https://doi.org/10.1101/2021.09.15.460240

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