Purpose of this study was to examine leptin (LEP) and pituitary-specific transcription factor-1 (Pit-I) gene polymorphisms in Holstein cattle (n= 352) in Turkey. In order to determine the Pit-I-HinfI and LEP-Sau3AI polymorphisms, polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP) were performed. A 600 bp length fragment of Pit-I and a 422 bp length fragment of LEP were amplified. In this study, two types of alleles, A and B, for both of the Pit-I and LEP genes were observed. The highest frequencies of the alleles were estimated, and were the B allele (0.68) for the Pit-I gene and the A allele (0.87) for the LEP gene. According to the results of the chi-square test, a significant deviation from the Hardy-Weinberg equilibrium was determined for both of these genes in the investigated breed (in the same order; X-2: 40.75, p<0.001and X-2: 8.4, p< 0.01).