Single-cell RNA sequencing with molecular spikes to determine RNA counting accuracy

Molecule counting is central to single-cell sequencing, yet no experimental strategy to evaluate counting performance exist. Here, we introduce RNA spike-ins containing inbuilt unique molecular identifiers (molecular spikes) that we use to monitor single-cell RNA counting performance across methods and to identify experimental steps essential for accurate counting. In this dataset, we add molecular spikes to popular single-cell RNA-seq protocols: SCRB-seq, Smart-seq3 and 10x Genomics (v2). For SCRB-seq and Smart-seq3, we also include variations of the library preparation procedure that are suspected to lead to changes in the UMI counting accuracy.