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SARS-CoV-2 spike gene Sanger sequencing methodology to identify variants of concern

Fatimah S Alhamlan^1,2,3, Dana M Bakheet^2,4, Marie F Bohol¹

¹Department of Infection & Immunity, King Faisal Specialist Hospital & Research Center, Riyadh, 11211, Saudi Arabia.

|February 16, 2023

Abstract:

The global demand for rapid identification of circulating SARS-CoV-2 variants of concern has led to a shortage of commercial kits. Therefore, this study aimed to develop and validate a rapid, cost-efficient genome sequencing protocol to identify circulating SARS-CoV-2 (variants of concern). Sets of primers flanking the SARS-CoV-2 spike gene were designed, verified and then validated using 282 nasopharyngeal positive samples for SARS-CoV-2. Protocol specificity was confirmed by comparing these results with SARS-CoV-2 whole-genome sequencing of the same samples. Out of 282 samples, 123 contained the alpha variant, 78 beta and 13 delta, which were indicted using in-house primers and next-generation sequencing; the numbers of variants found were 100% identical to the reference genome. This protocol is easily adaptable for detection of emerging variants during the pandemic.

Keywords:

PCR SARS-CoV-2 Sanger sequencing VOCs alpha beta delta genomic surveillance oligonucleotide synthesis

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