Quick and Easy Assembly of a One-Step qRT-PCR Kit for COVID-19 Diagnostics Using In-House Enzymes

One-step reverse-transcription quantitative polymerase chain reaction (qRT-PCR) is the most widely applied method for COVID-19 diagnostics. Notwithstanding facts that one-step qRT-PCR well suited diagnosis of and there are many commercially available kits in market, their high cost unavailability due to airport closures shipment restriction became a major bottleneck had driven desire produce key components such locally. Here, we provide simple, economical, powerful kit based on patent-free, specifically tailored versions Moloney murine leukemia virus reverse transcriptase Thermus aquaticus DNA termed R3T (Rapid Research Response Team) qRT-PCR. We also demonstrate robustness our enzyme production strategies optimal conditions efficient augmentation approach. Our was routinely able reliably detect as low 10 copies synthetic RNAs SARS-CoV-2. More importantly, successfully detected clinical samples broad viral titers with similar reliability selectivity Invitrogen SuperScript III Platinum TaqPath RT-qPCR kits. Overall, has shown robust performance both laboratory settings Saudi Ministry Health-approved testing facility.