Brief comparative evaluation of six open one-step RT-qPCR mastermixes for the detection of SARS-CoV-2 RNA using a Taqman probe

Abstract

Background: Facing the emergence of a new RNA virus, clinical laboratories are often helpless in the case of a shortage of reagents recommended by Reference Centres.

Objectives: To compare five open one step RT-qPCR reagents to the SuperScript™ III Platinum™ One-Step qRT-PCR kit (Invitrogen) considered as the reference one in France at the beginning of the pandemic for detection of the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) in respiratory specimens by using a laboratory-developed assay targeting the viral RNA dependant RNA polymerase (RdRp) gene.

Study design: A total of 51 NUCLISENS easyMAG extracts from respiratory specimens was tested on ABI 7500 thermocycler with TaqMan Fast Virus 1-Step Master Mix (Applied Biosystems), Luna® Universal Probe One-Step RT-qPCR Kit (New England Biolabs), GoTaq® Probe 1- Step RT-qPCR System (Promega), LightCycler® Multiplex RNA Virus Master (Roche) and One-step PrimeScript RT-PCR kit (Takara). The CT values obtained using the 5 challenged reagents were compared to those obtained using the reference assay.

Results: The percentages of concordance were all above 95 %. When comparing the CT values of the 48 extracts exhibiting CT values < 35 obtained with the reference reagent, the results were similar between the reagents although the differences of CT values were quite dispersed.

Conclusions: All five reagents can be considered as alternative reagents to the reference for detecting SARS-CoV-2 RNA.

Keywords: Coronavirus; Emerging virus; Laboratory-developed assay; RNA virus; RT-qPCR; SARS-CoV-2.