Melting Curve-Based Assay As an Alternative Technique for the Accurate Detection of Sars-Cov-2 Publisher



Aboutalebian S¹ ; Mirzaaghaei S¹ ; Fakhim H² ; Faramarzi S¹ ; Mousavi S¹ ; Ghafel S³ ; Gholipour S⁴ ; Farhang A⁵ ; Mirhendi H¹ ; Nikaeen M⁶ Show Affiliations
Source: Advanced Biomedical Research Published:2022

Abstract

Background: Early and cost-effective diagnosis and monitoring of the infection caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are critically important to anticipate and control the disease. We aimed to set up a SYBR Green-based one-step real-time polymerase chain reaction (PCR) as a lower-cost alternative method to detect the virus. Materials and Methods: An in-house SYBR Green-based PCR assay targeting the envelope (E) and RNA-dependent RNA polymerase (RdRp) genes, was set up to diagnose the infection, and was compared with the reference probe-based PCR method. Results: When the commercial probe-based assay was considered as the reference method, SYBR Green-based PCR had a slightly lower sensitivity (81.98% and 86.25% for E and RdRp targets, respectively) and a good specificity (100% and 94.44% for E and RdRp targets, respectively). For both gene targets, three different melting temperature (Tm) patterns were found in the PCRs of the nasopharyngeal/oropharyngeal swab samples, but no size polymorphism was seen in agarose gel electrophoresis. Conclusion: Further studies to improvement of the assay are needed to make it an inexpensive and reliable tool for the diagnosis of COVID-19. © 2022 Advanced Biomedical Research | Published by Wolters Kluwer-Medknow.