The purpose of this work was to identify GHRH, GH and PRL gene polymorphisms in Anatolian water buffalo by means of the PCR-RFLP method. A total of 126 buffalo were included in this study. PCR amplification gave a 451 bp band for the GHRH gene, a 221 bp band for the GH gene and a 156 bp band for the PRL gene. The PCR products were digested by HaeIII for the GHRH gene, AluI for the GH gene and RsaI for the PRL gene. The GH/AluI and PRL/RsaI polymorphisms were found to be polymorphic, while the GHRH/HaeIII polymorphism was not found in Anatolian water buffalo. The frequencies of GH-L (0.87) and PRL-A (0.55) alleles were found to be high in the examined Anatolian water buffalo. The chi-square test showed that the Anatolian water buffalo were in Hardy-Weinberg (HW) equilibrium for the GH gene while significant deviation was observed from HW equilibrium for the PRL gene. The present study is the first to examine GHRH/HaeIII, GH/AluI and PRL/RsaI polymorphisms in Anatolian water buffalo.