Polymorphisms of the prion protein gene (Prnp), especially the amino acid residue alterations at codons 136, 154, and 174, in sheep have been found to be associated with susceptibility to scrapie disease. We investigated Prnp polymorphisms in three local sheep breeds in Inner Mongolia, China. Blood samples were collected from 46 Ujumqin, 34 Sunite, and 22 Mongolian sheep. The genetic DNA of blood samples was extracted, amplified and sequenced, and amino acid alignment was determined. Polymorphisms were detected at 8 codons, among which M157I, Q220H, and R223K have not been previously reported. The frequency of the amino acid residues ARQ/ARQ at codons 136, 154, and 171, respectively, which is associated with medium-high susceptibility to scrapie, was 74.5%, and the frequency of scrapie-resistant genotype ARR/ARR was 7.9%. The highly susceptible genotype VRQ/VRQ at these codons as not detected from the tested sheep. Of the three sheep breeds, Ujumqin sheep had the highest frequency (15.2%) of scrapie-resistant amino acid sequence, ARR/ARR at codons 136, 154, and 171, respectively, accounting for 87.5% sheep that carry these polymorphisms. Our findings are of special importance for both live sheep export and sheep breeding.