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Extensive rate of variations in the S1 gene (spike glycoprotein subunit gene) of infectious bronchitis virus (IBV) causes challenges for clinicians in counting variants for differentiation of infected from vaccinated birds and addressing the variants of unknown significance. This study investigated the possibility of using an RNA‐dependent RNA polymerase gene (RdRp) as a target for molecular characterization of IBV strains in Iran. Trachea samples were collected from commercial broiler flocks (n = 52) showing respiratory syndrome. Specific PCR primers were designed for a variable region located in the RdRp gene flanked by highly conserved regions. Reverse transcriptase PCR followed by sequence analysis identified eight IBV variants, with an overall prevalence of 44.2%. Deduced nucleotide and amino acid sequences were compared with published sequences for IBV strains. Because of the long‐distance similarities, the field samples could be discriminated from vaccine strains. Phylogenetic analysis of RdRp gene sequences resulted in clustering of the IBV strains related to each area. Using RdRp as a genetic marker eliminates the challenges arising from the enormous variations that make it difficult to discriminate between field and vaccine strains as well as affiliate certain variants to various geographical areas. [ABSTRACT FROM AUTHOR]