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MANGA, I., ŘÍHA, J., VRTKOVÁ, I.: Polymorphism of CSN3, Pit-1, LGB and its impact on milk performance traits at the Czech Fleckvieh and Holstein breed. Acta univ. agric. et silvic. Mendel. Brun., 2008, LVI, No. 1, pp. 131–136 The eff ect of known CSN3, Pit-1 and LGB genes single nucleotide polymorphism (SNP) on milk performance traits (milk yield, protein, fat and lactose content, count of somatic cells) at the Czech Fleckvieh and the Holstein cattle breed was estimated. For more exact results, tested cows of each breed came from one source, and were selected into two groups according to their lactation. We tested around 100 animals at the fi rst and 25 at the high (more than fi ve) lactation of both breeds. Our attention was aimed particularly on combined genotypes of favourable alleles. We confi rmed the positive tendention of the cheese yield valuable CSN3/BB with Pit-1/AA combined genotype for high milk production. By estimation of single gene eff ect, we found out signifi cant association (P ≤ 0.01) between milk yield (kg) and Pit-1 genotypes at Holstein cows with the high lactation. This dependence was observed at the Czech Fleckvieh breed too, even it didn’t get conclusive value. Among all tested individuals, CSN3 genotypes signifi cantly infl uenced milk yield (AA > BB, P ≤ 0.01), milk fat (BB > AA, P ≤ 0.01) and protein content (BB > AA, P ≤ 0.01). The genotypes AB and BB of the LGB gene at Holstein cows were joined with higher milk yield in selected cattle groups opposite to AA genotype (P ≤ 0.01, P ≤ 0.05). The infl uence of LGB on the somatic cell count in created cattle groups was not confi rmed. Estimation of allele and genotype frequency according to lactation stage didn’t reveal higher diff erences at non of tested genes in both breeds. CSN3, LGB, Pit-1, SNP polymorphism, milk performance traits The aim of breeding procedures of the dairycow breeds is getting improvement of milk features quali ty at current high cow’s milk production. In terms of longtime breeding proceedings, there have been excellent parameters of cow’s milk production accomplished. However, there exist considerable variability in milk performance traits in breeds among particular individuals. This fact could be explained by existing polymorphism of the milk proteins coding genes. Simultaneously, it off ers possibilities for other animals selection. Choice of optimal genetic markers in combination with classic breeding procedures represents the most eff ective tool of livestock breeding, saving a lot of time (Meuwissen, 2003). The set of markers in our work conform for eff ort of producing milk with high protein content and with optimal technological characteristics for cheese manufacturing (polymorphism CSN3 and LGB genes) at high total milk production (polymorphism Pit-1gene) (Renaville, 2005). The relation of CSN3 to milk performance traits was evaluated by many authors (Renaville, 2005; Kučerová, 2004). As the most valuable allele is mentioned allele B, which is binded with higher percentage of proteins and fat content and also with optimal features for cheese producing. Allele A is mostly associated with higher yield of milk, fat and protein. Tsiaras, (2005), explored relations between CSN3 genotypes and reproduction parameters (lenght of gravidity, age at the fi rst and the second lactation, 132 I. Manga, J. Říha, I.Vrtková calving interval) and he didn‘t fi nd any associations with negative infl uence. LGB type B could serve as a marker for high protein content in milk. Likewise as the type B of CSN3, LGB type B is joined with advantage cheese yield features. Milk with genotype BB of the LGB gene allows obtaining of higher cheese yield in relation to milk with genotype AA of the LGB (Van den Berg, 1990). Genotype BB is o en associated with higher percentual content of kaseins and fats in milk, whereas AA genotype enhances the level of whey proteins in milk (McLean, 1984; Hill, 1993). Tsiaras, (2005), recently tested these known associations of the LGB, where he detected signifi cant impact of the LGB genotypes on milk yield, fat yield, fat content as well as protein yield and content in milk. In addition, testing of LGB polymorphism revealed binding with cows resistance to mastitis. Genotypes BB and AB were in signifi cant relation with lower count of somatic cells (Walawski, 1997), which is an important parameter of milk quali ty and refers to good health state of dairy udder. Pit-1 presents specifi c transcription factor, regulated an expression of GH (growth hormone) in mammals, activates an expression of thyreotropine and prolactine too. Moreover it plays an important role by regulation of cell diff erentiation and hypophysis proliferation. Pit-1 together with GH present essential genes for adjustment of mammary gland development and for milk secretion (Bauman, 1985; Peel, 1987). Substitution adenine (allele A) for guanine (allele B) at position 1178 of the Pit-1 gene (Dierkes et al., 1998) is joined with specifi c milk performance parameters too. Most o en, allele A infl uences milk production of dairycows positively. The aim of our work was to estimate actual status of screened genetic markers polymorphism at two, the most breeded cattle breeds for milk production in Czech Republic. Important part of our study constituted an evaluation of combined CSN3, Pit-1 genotypes infl uence on the observed milk performance traits. MATERIAL AND METHODS The tested groups of animals consisted of 125 cows of the Czech Fleckvieh breed and equal count of the Holstein breed, both breeds were represented almost entirely by pure breed animals. At the Fleckvieh breed, 100 cows were tested at the fi rst and 25 at the high lactation, at the Holstein breed, 105 cows were tested at the fi rst and 23 at the high lactation. DNA was isolated from milk using Jet quick blood and cell culture DNA spin kit (Genomed). To detect the genes polymorphism, we used PCR-RFLP method: for CSN3 analysis described by Soria et al., (2003), for Pit-1 described by Renaville et al., (2005) and for LGB described by Strzalkowska et. al., (2002). Results of testing genes polymorphism were evaluated in relation to follows milk performance parameters: milk yield (kg), percentage of fat, proteins and lactose, and count of somatic cells (thous/ml). Acquired results of analysis were processed in SAS 8.2 programme. ANOVA and GLM collections of methods were used for calculations of – 1) diff erences among groups caused by lactation (one-way Ftest); 2) diff erences among groups caused by genotype (Tukey post-hoc test); 3) diff erences among groups caused by genotype and lactation. Following model was used for GLM analysis: xijk = m + aj + bk + eijk, where: xijk is a value of milk parametr for individual i with genotype j in lactation k, m is a mean value of parametr, aj is an eff ect of j-th genotype, bk is an eff ect of k-th lactation, eijk is a random error. RESULTS AND DISCUSSION We estimated the relative genotypes and allele frequencies of tested markers at both breeds as shown in Table I, II and III. Acquired frequencies of the CSN3 gene (Table I.) are in agreement with fi ndings of other authors (Kučerová, 2004). Clear milk breed as Holstein featured with lower frequency of valuable allele B opposite to the combined meat-milk performance Czech Fleckvieh breed. Among particular lactations, there haven’t been signifi cant associations in allele frequency diff erences. We noted high signifi cant associations in milk yield, milk fat and protein content by analysis of all lactations according to CSN3 geno types regardless of the breed classifi cation. Here, the allele A positive infl uenced the milk yield (P ≤ 0.01), as well as the allele B increased the fat (P ≤ 0.01) and protein (P ≤ 0.01) content. These fi nding also confi rm fi ndings in other studies. The estimation of eff ect combined Pit-1 and CSN3 genotypes on milk yield lended outcomes, which are viewable in Table IV. The results indicate, that the impact of allele A in both genes on milk yield is viewable at the combined Pit-1/CSN3 genotypes too. By comparison detected evidential associations among combined genotypes, the genotype BB/BB featured always with lower milk production at the Holstein breed at the fi rst lactation. Even if the desirable AA/BB genotype, by reason of defi cient count of animals wearing this genotype, couldn’t be evaluated even at one breed, but the similar genotype AB/BB had equally positive eff ect on elevation milk production. The situation at the Fleckvieh breed was the same, though just the BB/AA combined genotypes at the fi rst lactation produced signifi cantly more milk opposite to BB/BB genotypes. Renaville, (2005), detected eff ect +273kg on milk yield/ per lactation/per dairycow of the AA/ BB combined genotypes in comparison to BB/AA genotypes at tested Holstein breed. This is in agreement with fi ndings of our work. Hence, selection of combined Pit-1/CSN3 genotypes AA/BB (AB/BB) at the dairycows breeds should aff ord a way, how to ensure production of milk with valuable BB/CSN3 geno types at simultaneously high milk production. Unfortunately, eff ect of combined genotypes on milk performance traits of cows tested at the high lactation can’t be evaluated considering their low count. Polymorphism of CSN3, Pit-1, LGB and its impact on milk performance traits 133 I: CSN3 gene: comparison of relative genotype and allelic frequency in breeds tested at the fi rst and the high lactation gene CSN3 lactation n genotype frequency allele frequency