Water buffalo breeding is practiced in various provinces of Turkey, and is also included in the National Breeding Project. It was aimed in this study to investigate polymorphisms on exon 4-intron 5 and exon 4 only of the kappa-casein (CSN3) gene for milk yields of water buffalo bred in Sivas Province. Blood samples were taken from 135 water buffaloes. The phenol/chloroform method for DNA extraction was conducted on collected blood. Polymerase chain reaction (PCR) was used to amplify CSN3-specific regions, and amplified products were run using 2% agarose gel electrophoresis (AGE). Various restriction endonucleases (RE) were utilized in respect to their protocols to investigate polymorphisms in amplified products. Following the RE digestion, mixtures were run in 2.5% AGE. It was observed that only the BB genotype was obtained in terms of Hinf I enzyme; however, the E allele (Mae III) was not obtained for the CSN3 gene. For the Hind III enzyme, only the BB genotype was obtained from exon 4 of the CSN3 gene, while both AB (27.41%) and BB (72.59%) genotypes were determined from exon 4-intron 5 of the CSN3 gene. No statistical difference was detected for genotypes of the restriction enzymes used in Anatolian water buffaloes (P > 0.05). In conclusion, the Hinf I/BB genotype, associated with higher milk protein amount and milk fat, was observed in Anatolian water buffaloes.