2-4- Phylogenetic assessment
PCR products underwent purification using High Pure PCR Product Purification Kit (Roche Diagnostic GmbH, Mannheim, Germany) corresponding to the manufacturer’s instructions. Also, an ABI 3730 XL sequencer was used for bidirectional sequencing. Raw data were trimmed and analyzed using popular bioinformatics software CLC workbench 5, and Basic Local Alignment Search Tool (BLAST). All sequences were submitted into GeneBank database (https://www.ncbi.nlm.nih.gov/genbank/) and released accession numbers were used to draw phylogenetic tree. SARS-CoV-2 reference sequence (NC_045512.2) and other coronavirus sequences were obtained from NCBI (https://blast.ncbi.nlm.nih.gov) database. Moreover, MEGA X software was used for multiple sequence alignment (MSA) and the phylogenetic trees were drown using 1000 replicate bootstrap method. In addition, the neighbor joining and maximum likelihood methods were carried out for similarity assessment and evolution analysis.