Design and in silico validation of polymerase chain reaction primers to detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)
Covid-19; real time PCR; diagnosis; primer design;
The design of polymerase chain reaction (PCR) primers that target conserved segments of viral genomes is important to prevent false-negative results and reduce the need to standardize different PCR protocols for the same target. In this work, we designed and described a set of primers and probes that target conserved regions identified from multiple alignment of 2,341 SARS-CoV-2 genomes available in the GISAID (Global Initiative on Sharing All Influenza Data) database. Subsequently, the primers were validated together with the probes on 211,833 sequences from the entire genomes of SARS-CoV-2. Nine systems were obtained (primer forward+reverse+probes) that potentially anneal to the highly conserved regions of the virus genome identified in this analysis. In silico predictions also demonstrated that the primers do not interact with non-specific targets in sequences from humans, bacteria, fungi, Apicomplexa and other betacoronaviruses and less pathogenic coronavirus strains. The publication of these primer and probes sequences will make it possible to validate more efficient protocols for identifying SARS-CoV-2.