The aim of this study was to investigate the nucleotide diversity and the phylogenetic analysis of pancreatic ribonuclease gene in two different species, Baluchi and Urial sheep. Baluchi sheep blood samples were collected from Abbas Abad Animal Breeding Center, Mashhad and Urial sheep blood samples were collected from Khorasan Razavi Environmental Department. The 4347 bp pancreatic ribonuclease gene was amplified and sequenced using seven primer pairs that were designed to overlap each other. Comparison of 4347 bp pancreatic ribonuclease gene sequences obtained from both Baluchi and Urial sheep strains showed 10 nucleotide differences with 99.78% similarity. Also results of comparison of pancreatic ribonuclease gene from the sheep reference genome (Accession number of XM_004010384) with Urial sheep and Baluchi showed differences in 18 nucleotides with 99.61% similarity and 10 nucleotides with 99.78% similarity, respectively. The results of the present study confirmed that although the pancreatic ribonuclease coding enzyme sequences of the two species, Baluchi and Urial and the reference gene sequences from the NCBI database are completely similar, but the upstream and downstream nucleotide sequences of this gene have 10 nucleotide differences with each other. There are 18 nucleotide differences with the sheep reference sequence. Results of genetic interval matrix and phylogenetic tree sequences of Baluchi sheep and Urial sheep obtained in this study with other gene sequences registered from pancreatic ribonuclease in the NCBI database, it confirmed that both Baluchi and Urial sheep species with the sheep reference sequence for pancreatic ribonuclease gene sequence were assigned to a goat species belonged to the same clade.