The aim of this study was to identify the prion protein (PrP) gene polymorphism in a total of 1,110 healthy sheep from 18 Turkish native sheep breeds. There were nine alleles and 22 genotypes observed based on codons 136, 154, and 171 of the PrP gene. The ARQ allele was predominant for all breeds. The most resistant allele to scrapie, ARR, was present in all breeds. The VRQ allele, associated with the highest susceptibility to scrapie, was detected at low frequencies in İvesi (0.06), Kıvırcık (0.021), Sakız (0.010), Karayaka (0.011), Çine Çaparı (0.012), and Güneykaraman (0.017). In general, the ARQ/ARQ genotype was predominant in all breeds. The most resistant genotype to scrapie, ARR/ARR, was found with the frequency lower than 0.180. The most susceptible genotype, VRQ/VRQ, was found in only Kıvırcık. The TRR and TRH alleles and the genotypes of ARR/TRR, ARR/ARK, and ARH/TRH have been found for the first time in Turkish native sheep breeds. According to these results, all breeds belong to risk group R3 followed by R2. It is propounded that the susceptibility to scrapie increased from eastern to western part of Turkey. Our findings of Turkish native sheep breeds with PrP gene polymorphisms will assist the sheep breeding program for selection of scrapie resistance genotypes to reduce the risk of scrapie.