Single cell
RNA sequencing (
scRNA-Seq) studies have provided critical insight into the pathogenesis of Severe Acute Respiratory Syndrome CoronaVirus 2 (SARS-CoV-2), the causative agent of
COronaVIrus Disease 2019 (COVID-19).
scRNA-Seq workflows are generally designed for the detection and quantification of eukaryotic host mRNAs and not viral RNAs. Here, we compare different
scRNA-Seq methods for their ability to quantify and detect SARS-CoV-2 RNAs with a focus on subgenomic mRNAs (sgmRNAs). We present a data processing strategy, single cell CoronaVirus sequencing (scCoVseq), which quantifies reads unambiguously assigned to sgmRNAs or genomic
RNA (gRNA). Compared to standard 10X Genomics
Chromium Next GEM Single Cell 3' (10X 3') and
Chromium Next GEM Single Cell V(D)J (10X 5') sequencing, we find that 10X 5' with an extended read 1 (R1) sequencing strategy maximizes the detection of sgmRNAs by increasing the number of unambiguous reads spanning leader-
sgmRNA junction sites. Using this method, we show that viral gene expression is highly correlated across cells suggesting a relatively consistent proportion of viral
sgmRNA production throughout
infection. Our method allows for quantification of coronavirus
sgmRNA expression at single-cell resolution, and thereby supports high resolution studies of the dynamics of coronavirus
RNA synthesis.