VPrimer: A Method of Designing and Updating Primer and Probe With High Variant Coverage for RNA Virus Detection

Abstract

Fatal infectious diseases caused by RNAviruses, such as COVID-19, have emerged around the world. RT-PCR is widely employed for virus detection, and its accuracy depends on the primers and probes since RT-PCR can detect a virus only when the primers and probes bind to the target gene of the virus. Most of primer design methods are for a single host and so require a great deal of effort to design for RNAvirus detection, including homology tests among the host and all the viruses for the host using BLAST-like tools. Furthermore, they do not consider variant sequences, which are very common in viruses. In this study, we describe VPrimer, a method of designing high-quality primer-probe sets for RNAviruses. VPrimer can find primer-probe sets that cover more than 95% of the variants of a target virus but do not cover any sequences of other viruses or the host. With VPrimer, we found 381,698,582 primer-probe sets for 3,104 RNAviruses. Multiplex PCR assays using the top 2 primer-probe sets suggested by VPrimer usually cover 100% of variants. To address the rapid changes in viral genomes, VPrimer finds the best and up-to-date primer-probe sets incrementally against the most recently reported variants.