Melting Curve‑based Assay as an Alternative Technique for the Accurate Detection of SARS‑CoV‑2

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.

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.

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.

Further studies to improvement of the assay are needed to make it an inexpensive and reliable tool for the diagnosis of COVID‑19.