Abstract
Accurate designing of polymerase chain reaction (PCR) primers targeting conserved segments in viral genomes is desirable for preventing false-negative results and decreasing the need for standardization across different PCR protocols. In this work, we designed and described a set of primers and probes targeting conserved regions identified from a multiple sequence alignment of 2341 Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) genomes from the Global Initiative on Sharing All Influenza Data (GISAID). We subsequently validated those primers and probes in 211,833 SARS-CoV-2 whole-genome sequences. We obtained nine systems (forward primer + reverse primer + probe) that potentially anneal to highly conserved regions of the virus genome from these analyses. In silico predictions also demonstrated that those primers do not bind to nonspecific targets for human, bacterial, fungal, apicomplexan, and other Betacoronaviruses and less pathogenic sub-strains of coronavirus. The availability of these primer and probe sequences will make it possible to validate more efficient protocols for identifying SARS-CoV-2.
Highlights
Accurate designing of polymerase chain reaction (PCR) primers targeting conserved segments in viral genomes is desirable for preventing false-negative results and decreasing the need for standardization across different PCR protocols
In silico predictions demonstrated that those primers do not bind to nonspecific targets for human, bacterial, fungal, apicomplexan, and other Betacoronaviruses and less pathogenic sub-strains of coronavirus
We identified 26 conserved segments (CS) in the SARS-CoV-2 genome based on an alignment of 2,341 full genome sequences and used these regions as a target for the design of universal primers and probes
Summary
Accurate designing of polymerase chain reaction (PCR) primers targeting conserved segments in viral genomes is desirable for preventing false-negative results and decreasing the need for standardization across different PCR protocols. In silico predictions demonstrated that those primers do not bind to nonspecific targets for human, bacterial, fungal, apicomplexan, and other Betacoronaviruses and less pathogenic sub-strains of coronavirus The availability of these primer and probe sequences will make it possible to validate more efficient protocols for identifying SARS-CoV-2. Organization (WHO) released some SARS-CoV-2 polymerase chain reaction (PCR) protocol assays produced by different reference institutions in the world[1] In addition to these initial protocols, an increasing number of works and commercial kits suggest new alternatives to identifying SARS-CoV-2 and its recent variants by molecular or immunological a pproaches[2,3,4]. We extended the analyses to include 211,833 SARS-CoV-2 sequences and the recent virus
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