Fig. 1: Amplicon-seq analysis characterize sgRNA relative abundance in clinical swabs in COVID-19 positive patients.
From: Reduced subgenomic RNA expression is a molecular indicator of asymptomatic SARS-CoV-2 infection
a SARS-CoV-2 RNAs existed in the upper respiratory tracts from both symptomatic and asymptomatic COVID-19 positive patients were analyzed by Amplicon-seq. Amplicons tilted across full-length SARS-CoV-2 genomes were amplified by specific primers in the multiplex PCRs in two separate pools. Subgenomic RNAs (sgRNAs) specific amplicons can be identified by specific amplification with 5′ forward primer closest to the transcription regulatory sequence leader (TRS-L) site and 3′ reverse primers closest to the TRS-Body (TRS-B) sites followed by sequencing and alignment. Distribution of sgRNAs normalized counts (b) and sgRNA to genomic RNA (gRNA) ratio (p = 4.9  × 10−12) (c) between symptomatic and asymptomatic cases (p = 5.6 × 10−12). d Sequencing coverage across the 5′ 400 nucleotides of SARS-CoV-2 genome shows the contribution from sgRNAs and gRNA, respectively. e Amplicon-seq coverages across 5′ 400 nucleotides from representative symptomatic and asymptomatic cases. f Distribution of the normalized counts of individual sgRNA species measured in symptomatic and asymptomatic cases (P values of pairwise comparison for S, M, ORF6, ORF7b, and N are 6 × 10−11, 9 × 10−12, 2 × 10−11, 2 × 10−7, and 9 × 10−10). All statistical tests are two-sided Wilcoxon rank-sum test. Center line, median; boxes, first and third quartiles; whiskers, 1.5Ă— the interquartile range.
