There were observed milk production parameters in daughters of 37 Czech Pied cattle sires in our study. Sires were genotyped for markers CSN3 and ETH10. The relation between CSN3 genotype and milk production parameters was found to be significant. The effect of CSN3 genotype AA on milk, protein and fat yield and effect of genotype BB on protein and fat percentage was proved. Frequency of allele B and genotype BB in Czech Pied cattle was similar to other Simmental breeds and was also higher than allele frequencies found in Holsteins. These findings after verification on larger populations using granddaughter design population structure can be used in breeding program of Czech Pied cattle. Significant relation was also found between microsatellite markerETH10 and milk fat content. Further study is needed to prove this finding in larger populations of sires and their grandsires with higher genetic variability of their breeding values before considering implementing markers into breeding