Rapid Homemade Reverse Transcriptase PCR Detection and Phylogenetic Analysis for SARS-CoV-2 Based on E and M Genes

Abstract

Background & Objective: Coronavirus disease 2019 spreads worldwide and needs detection systems capable of rapid diagnostic of this virus (SARS-CoV-2). The aim of this study is to design the homemade RT-PCR method for the Detection and phylogenetic analysis of this virus Material & Methods: The genes selected for diagnosis were E and M genes for this virus. PCR product was cloned in pTZ57R/T plasmid for preparation of positive control. In order to determine the sensitivity of this molecular method, the genes mentioned in the clone pTZ57R/T vector and the Limit of detection (LOD) the genes were determined and phylogenetic analysis was performed using partial E and M gene sequences. Results: PCR product was observed for E and M genes 156 and 547 bp on the Agarose gel. The LOD of the E and M gene was 60 and 82 copies. There was also a positive response to the samples of patients who were positive by other methods. Conclusions: Since this virus is considered to be the cause of a pandemic in different countries all over the world, the present study is very important as a method of rapid and low-cost molecular diagnosis for monitoring this virus. Phylogenetic analysis is necessary for epidemiological studies for the control and prevention of the disease.

Keywords

• rt-pcr
• covid-19
• detection
• lod
• phylogenetic analysis