Supporting data for"Identification and quantification of SARS-CoV-2 leader subgenomic mRNA gene junctions in nasopharyngeal samples shows phasic transcription in animal models of COVID-19 and dysregulation at later time points that can also be identified in humans"

SARS-CoV-2 has a complex strategy for the transcription of viral subgenomic mRNAs (sgmRNAs), which are targets for nucleic acid diagnostics. Each of these sgmRNAs has a unique 5’ sequence, the leader-transcriptional regulatory sequence gene junction (leader-TRS-junction), that can be identified using sequencing. High resolution sequencing has been used to investigate the biology of SARS-CoV-2 and the host response in cell culture and animal models and from clinical samples. LeTRS, a bioinformatics tool, was developed to identify leader-TRS-junctions and be used as a proxy to quantify sgmRNAs for understanding virus biology. LeTRS is readily adaptable for other coronaviruses such as Middle East respiratory syndrome coronavirus (MERS-CoV) or a future newly discovered coronavirus. LeTRS accepts FASTQ files derived from Illumina paired-end or Oxford Nanopore sequencing (amplicon or direct RNA) and showed greater sensitivity and F-measure score than the other currently available tools for processing the Nanopore sequencing reads. LeTRS was tested on published datasets and novel clinical samples from patients and longitudinal samples from animal models with COVID-19. LeTRS identified known leader-TRS-junctions and identified putative novel sgmRNAs that were common across different mammalian species. This may be indicative of an evolutionary mechanism where plasticity in transcription generates novel open reading frames, that can then subject to selection pressure. The data indicated multi-phasic abundance of sgmRNAs in two different animal models. This recapitulates the relative sgmRNA abundance observed in cells at early points in infection, but not at late points. This pattern is reflected in some human nasopharyngeal samples, and therefore has implications for transmission models and nucleic acid-based diagnostics. LeTRS provides a quantitative measure of sgmRNA abundance from sequencing data. This can be used to assess the biology of SARS-CoV-2 (or other coronaviruses) in clinical and non-clinical samples, especially to evaluate different variants and medical countermeasures that may influence viral RNA synthesis.