Influence of ethanol on the rennet-induced coagulation of milk

The influence of ethanol on the rennet-induced coagulation of milk was studied to investigate potential synergistic effects of these two mechanisms of destabilisation on the casein micelles. Addition of 5% (v/v) ethanol reduced the rennet coagulation time (RCT) of milk, whereas higher levels of ethanol (10-20%, v/v) progressively increased RCT. The temperature at which milk was coagulable by rennet decreased with increasing ethanol content of the milk. The primary stage of rennet coagulation, i.e., the enzymatic hydrolysis of kappa-casein, was progressively slowed with increasing ethanol content (5-20%, v/v), possibly due to ethanol-induced conformational changes in the enzyme molecule. The secondary stage of rennet coagulation, i.e., the aggregation of kappa-casein-depleted micelles, was enhanced in the presence of 5-15% ethanol, the effect being largest at 5% ethanol. Enhanced aggregation of micelles is probably due to an ethanol-induced decrease in inter-micellar steric repulsion. These results indicate an interrelationship between the effects of ethanol and chymosin on the casein micelles in milk, which may have interesting implications for properties of dairy products.     

Association between the bovine milk metabolome and rennet-induced coagulation properties of milk

The milk metabolomes of 407 individual Swedish Red dairy cows were analyzed by nuclear magnetic resonance spectroscopy as part of the Danish-Swedish Milk Genomics Initiative. By relating these metabolite profiles to total milk protein concentration and rheological measurements of rennet-induced milk coagulation together using multivariate data analysis techniques, we were able to identify several different associations of the milk metabolome to technological properties of milk. Several novel correlations of milk metabolites to protein content and rennet-induced coagulation properties were demonstrated. Metabolites associated with the prediction of total protein content included choline, N-acetyl hexosamines, creatinine, glycerophosphocholine, glutamate, glucose 1-phosphate, galactose 1-phosphate, and orotate. In addition, levels of lactate, acetate, glutamate, creatinine, choline, carnitine, galactose 1-phosphate, and glycerophosphocholine were significantly different when comparing noncoagulating and well-coagulating milks. These findings suggest that the mentioned metabolites are associated with milk protein content and rennet-induced coagulation properties and may act as quality markers for cheese milk.     

Characterisation of non-coagulating milk and effects of milk composition and physical properties on rennet-induced coagulation in Swedish Red Dairy Cattle

Non-coagulating milk is a serious problem in the cheese industry, since it decreases cheese yield, resulting in decreased economic output. This study evaluated rennet-induced coagulation properties and composition of milk from individual Swedish Red Dairy Cattle. Milk samples from 679 individual cows were rheologically evaluated, of which 18.1% of the cows produced non-coagulating milk and 18.9% produced poor-coagulating milk. This resulted in 37% of the milk samples being non-optimal in cheese production, which is an alarmingly high figure. A comparison between non-coagulating and coagulating milk showed a significantly lower calcium content and less free Ca²⁺ in non-coagulating milk. The results provide more information about non- and poor-coagulating milk and will be further used to understand the genetic background of non-coagulating milk and breed against this undesired milk property.     

Investigation of an in-line prototype fluorescence and infrared backscatter sensor to monitor rennet-induced coagulation of skim milk at different protein concentrations

Coagulation of milk is one of the most important steps in cheese manufacture. Cutting the coagulum at optimum firmness is important to optimise the yield and quality of the cheese produced. The aim of this study was to investigate a prototype sensor to monitor rennet-induced coagulation of skim milk at different protein concentrations (3.3%, 4.0% and 4.7%) and to develop a model to predict the coagulum cutting time at a desired storage modulus (G′). Fluorescence and infrared backscatter profiles were recorded at wavelengths of 350 and 880 nm, respectively. Rheological measurements were used as a reference method to determine the times required for the coagulum to reach G′ values of 0.5, 5 and 20 Pa. Time parameters extracted from the optical profiles generated during the coagulation process were used to develop a model to predict the cutting time at which the coagulum reaches selected G′ values. This study demonstrated that the investigated prototype sensor, combined with the developed prediction model, can be used as an in-line PAT tool for real-time monitoring of milk coagulation and prediction of cutting time in cheese manufacturing.     

Effect of high hydrostatic pressure processing on the rennet coagulation kinetics and physicochemical properties of sheep milk rennet-induced gels

The effects of high hydrostatic pressure on the constituents and coagulation ability and their effect on cheese production of sheep milk have not been studied in detail. The objective of this work was to evaluate the effect of high hydrostatic pressure processing on the coagulation kinetics and physicochemical properties of sheep milk and to explore how such treatment could improve the cheesemaking process. Five batches of milk were tested: 1 untreated control batch and 4 batches each subjected to a different pressure (150, 300, 450, or 600 MPa) for 5 min at 10°C. As treatment pressure increased, values of electrical conductivity and oxidation-reduction potential were found to decrease. However, no significant reduction in pH was recorded. Treatment pressures >300 MPa produced milk with lower lightness (luminosity) and a more yellow and green hue. Pressures >150 MPa resulted in micellar fragmentation, as well as significant increases in particle size, viscosity, and water-holding capacity as a consequence of the denaturing of soluble proteins. High-pressure treatments increased the solubility of colloidal calcium phosphate, leading to a considerable increase in the concentration of minerals in the serum phase. The highest concentrations of calcium and phosphorus in the rennet whey of milk were reached at 300 MPa. Curd coagulation time was reduced by 28% at pressures >300 MPa, and an increase in the curd firming rate was observed. As treatment pressure increased to 450 MPa, the firmness, elasticity, and the percentage creep recovery of gels increased, whereas values of compliance and fracture strain were reduced. Thus, we can conclude that 300 MPa is the optimum treatment pressure for milk intended for cheesemaking by enzymatic coagulation. This pressure produced milk with optimal coagulation kinetics and water-holding properties with the least loss of fat and protein to the whey.     

Addition of sodium caseinate to skim milk inhibits rennet-induced aggregation of casein micelles

The objective of this paper was to observe the rennet-induced aggregation behaviour of casein micelles in milk in the presence of additional sodium caseinate. Analysis of the centrifugal supernatants by size exclusion chromatography confirmed an increase in the soluble protein in the milk serum phase after addition of sodium caseinate. Although the total amount of κ-casein hydrolyzed over time was not affected, there was a significant effect of soluble casein on milk gelation, with a dose-dependent decrease of the gelation time as measured by rheology. Light scattering experiments also confirmed that the addition of soluble caseins inhibited the aggregation of casein micelles. Addition of 1 mM CaCl₂ prior to renneting increased the extent of rennet aggregation in samples containing additional sodium caseinate, but the inhibiting effect was still evident. The amount of soluble casein (as measured by chroma tography) significantly decreased after renneting, suggesting its association with the micellar fraction. Supporting experiments carried out with purified fractions of soluble caseins demonstrated that both α_s-casein and β-casein played a role as protective colloids (increasing steric repulsion) during renneting. It was concluded that the inhibiting effect observed during gelation was caused by the adsorption of soluble casein molecules on the surface of rennet-altered casein micelles.     

Role of CSN2, CSN3, and BLG genes and the polygenic background in the cattle milk protein profile

To devise better selection strategies in dairy cattle breeding programs, a deeper knowledge of the role of the major genes encoding for milk protein fractions is required. The aim of the present study was to assess the effect of the CSN2, CSN3, and BLG genotypes on individual protein fractions (α_S1-CN, α_S2-CN, β-CN, κ-CN, β-LG, α-LA) expressed qualitatively as percentages of total nitrogen content (% N), quantitatively as contents in milk (g/L), and as daily production levels (g/d). Individual milk samples were collected from 1,264 Brown Swiss cows reared in 85 commercial herds in Trento Province (northeast Italy). A total of 989 cows were successfully genotyped using the Illumina Bovine SNP50 v.2 BeadChip (Illumina Inc.), and a genomic relationship matrix was constructed using the 37,519 SNP markers obtained. Milk protein fractions were quantified and the β-CN, κ-CN, and β-LG genetic variants were identified by reversed-phase HPLC (RP-HPLC). All protein fractions were analyzed through a Bayesian multitrait animal model implemented via Gibbs sampling. The effects of days in milk, parity order, and the CSN2, CSN3, and BLG genotypes were assigned flat priors in this model, whereas the effects of herd and animal additive genetic were assigned Gaussian prior distributions, and inverse Wishart distributions were assumed for the respective co-variance matrices. Marginal posterior distributions of the parameters of interest were compared before and after the inclusion of the effects of the 3 major genes in the model. The results showed that a high portion of the genetic variance was controlled by the major genes. This was particularly apparent in the qualitative protein profile, which was found to have a higher heritability than the protein fraction contents in milk and their daily yields. When the genes were included individually in the model, CSN2 was the major gene controlling all the casein fractions except for κ-CN, which was controlled directly by the CSN3 gene. The BLG gene had the most influence on the 2 whey proteins. The genetic correlations showed the major genes had only a small effect on the relationships between the protein fractions, but through comparison of the correlation coefficients of the proteins expressed in different ways they revealed potential mechanisms of regulation and competitive synthesis in the mammary gland. The estimates for the effects of the CSN2 and CSN3 genes on protein profiles showed overexpression of protein synthesis in the presence of the B allele in the genotype. Conversely, the β-LG B variant was associated with a lower concentration of β-LG compared with the β-LG A variant, independently of how the protein fractions were expressed, and it was followed by downregulation (or upregulation in the case of the β-LG B) of all other protein fractions. These results should be borne in mind when seeking to design more efficient selection programs aimed at improving milk quality for the efficiency of dairy industry and the effect of dairy products on human health.     

The mineral profile affects the coagulation pattern and cheese-making efficiency of bovine milk

Natural variations in milk minerals, their relationships, and their associations with the coagulation process and cheese-making traits present an opportunity for the differentiation of milk destined for high-quality natural products, such as traditional specialties or Protected Designation of Origin (PDO) cheeses. The aim of this study was to quantify the effects of the native contents of Ca, P, Na, K, and Mg on 18 traits describing traditional milk coagulation properties (MCP), curd firming over time (CF_t) equation parameters, cheese yield (CY) measures, and nutrient recoveries in the curd (REC) using models that either included or omitted the simultaneous effects of milk fat and casein contents. The results showed that, by including milk fat and casein and the minerals in the statistical model, we were able to determine the specific effects of each mineral on coagulation and cheese-making efficiency. In general, about two-thirds of the apparent effects of the minerals on MCP and the CF_t equation parameters are actually mediated by their association with milk composition, especially casein content, whereas only one-third of the effects are direct and independent of milk composition. In the case of cheese-making traits, the effects of the minerals were mediated only negligibly by their association with milk composition. High Ca content had a positive effect on the coagulation pattern and cheese-making traits, favoring water retention in the curd in particular. Phosphorus positively affected the cheese-making traits in that it was associated with an increase in CY in terms of curd solids, and in all the nutrient recovery traits. However, a very high P content in milk was associated with lower fat recovery in the curd. The variation in the Na content in milk only mildly affected coagulation, whereas with regard to cheese-making, protein recovery was negatively associated with high concentrations of this mineral. Potassium seemed not to be actively involved in coagulation and the cheese-making process. Magnesium content tended to slow coagulation and reduce CY measures. Further studies on the relationships of minerals with casein and protein fractions could deepen our knowledge of the role of all minerals in coagulation and the cheese-making process.     

Goat farm variability affects milk Fourier-transform infrared spectra used for predicting coagulation properties

Driven by the large amount of goat milk destined for cheese production, and to pioneer the goat cheese industry, the objective of this study was to assess the effect of farm in predicting goat milk-coagulation and curd-firmness traits via Fourier-transform infrared spectroscopy. Spectra from 452 Sarda goats belonging to 14 farms in central and southeast Sardinia (Italy) were collected. A Bayesian linear regression model was used, estimating all spectral wavelengths' effects simultaneously. Three traditional milk-coagulation properties [rennet coagulation time (min), time to curd firmness of 20 mm (min), and curd firmness 30 min after rennet addition (mm)] and 3 curd-firmness measures modeled over time [rennet coagulation time estimated according to curd firmness change over time (RCT_eq), instant curd-firming rate constant, and asymptotical curd firmness] were considered. A stratified cross validation (SCV) was assigned, evaluating each farm separately (validation set; VAL) and keeping the remaining farms to train (calibration set) the statistical model. Moreover, a SCV, where 20% of the goats randomly taken (10 replicates per farm) from the VAL farm entered the calibration set, was also considered (SCV₈₀). To assess model performance, coefficient of determination (R²_VAL) and the root mean squared error of validation were recorded. The R²_VAL varied between 0.14 and 0.45 (instant curd-firming rate constant and RCT_eq, respectively), albeit the standard deviation was approximating half of the mean for all the traits. Although average results of the 2 SCV procedures were similar, in SCV₈₀, the maximum R²_VAL increased at about 15% across traits, with the highest observed for time to curd firmness of 20 mm (20%) and the lowest for RCT_eq (6%). Further investigation evidenced important variability among farms, with R²_VAL for some of them being close to 0. Our work outlined the importance of considering the effect of farm when developing Fourier-transform infrared spectroscopy prediction equations for coagulation and curd-firmness traits in goats.     

Novel functional mutation of the PDIA3 gene affects milk composition traits in Chinese Holstein cattle

Protein disulfide isomerase family A member 3 (PDIA3) is a multifunctional protein, and it plays a vital role in modulating various cell biological functions under physiological and pathological conditions. Our previous study on Mediterranean buffalo demonstrated that PDIA3 is a potential candidate gene associated with milk yield based on genome-wide association study analysis. However, the genetic effects of the PDIA3 gene on milk performance in dairy cattle and the corresponding mechanism have not been documented. This study aims to explore the genetic effects of PDIA3 polymorphisms on milk production traits in 362 Chinese Holstein cattle. The results showed that 4 SNPs were identi?ed from the 5′ untranslated region of the PDIA3 gene in the studied population, of which 2 SNPs (g.-1713 C>T and g.-934 G>A) were confirmed to be significantly associated with milk protein percentage, whereas g.-434 C>T was significantly associated with milk fat percentage. Notably, linkage disequilibrium analysis indicated that 3 SNPs (g.-1713 C>T, g.-934 G>A, and g.-695 A>C) formed one haplotype block, which was found to be significantly associated with milk protein percentage. The luciferase assay demonstrated that allele C of g.-434 C>T exhibited a higher promotor activity compared with allele T, suggesting that g.-434 C>T might be a potential functional mutation affecting PDIA3 expression. Furthermore, overexpression of the PDIA3 gene was found to induce higher levels of triglyceride and BODIPY fluorescence intensity. In addition, PDIA3 overexpression was also found to positively regulate the synthesis and secretion of α-casein, β-casein, and κ-casein, whereas knockdown of this gene showed the opposite effects. In summary, our findings revealed significant genetic effects of PDIA3 on milk composition traits, and the identified SNP and the haplotype block might be used as genetic markers for dairy cow selected breeding.     

Breed of goat affects the prediction accuracy of milk coagulation properties using Fourier-transform infrared spectroscopy

The prediction of traditional goat milk coagulation properties (MCP) and curd firmness over time (CF_t) parameters via Fourier-transform infrared (FTIR) spectroscopy can be of significant economic interest to the dairy industry and can contribute to the breeding objectives for the genetic improvement of dairy goat breeds. Therefore, the aims of this study were to (1) explore the variability of milk FTIR spectra from 4 goat breeds (Camosciata delle Alpi, Murciano-Granadina, Maltese, and Sarda), and to assess the possible discriminant power of milk FTIR spectra among breeds, (2) assess the viability to predict coagulation traits by using milk FTIR spectra, and (3) quantify the effect of the breed on the prediction accuracy of MCP and CF_t parameters. In total, 611 individual goat milk samples were used. Analysis of variance of measured MCP and CF_t parameters was carried out using a mixed model including the farm and pendulum as random factors, and breed, parity, and days in milk as fixed factors. Milk spectra for each goat were collected over the spectral range from wavenumber 5,011 to 925 × cm^?1. Discriminant analysis of principal components was used to assess the ability of FTIR spectra to identify breed of origin. A Bayesian model was used to calibrate equations for each coagulation trait. The accuracy of the model and the prediction equation was assessed by cross-validation (CRV; 80% training and 20% testing set) and stratified CRV (SCV; 3 breeds in the training set, one breed in the testing set) procedures. Prediction accuracy was assessed by using coefficient of determination of validation (R²_VAL), the root mean square error of validation (RMSE_VAL), and the ratio performance deviation. Moreover, measured and FTIR predicted traits were compared in the SCV procedure by assessing their least squares means for the breed effect, Pearson correlations, and variance heteroscedasticity. Results showed the feasibility of using FTIR spectra and multivariate analyses to correctly assign milk samples to their breeds of origin. The R²_VAL values obtained with the CRV procedure were moderate to high for the majority of coagulation traits, with RMSE_VAL and ratio performance deviation values increasing as the coagulation process progresses from rennet addition. Prediction accuracy obtained with the SCV were strongly influenced by the breed, presenting general low values restricting a practical application. In addition, the low Pearson correlation coefficients of Sarda breed for all the traits analyzed, and the heteroscedastic variances of Camosciata delle Alpi, Murciano-Granadina, and Maltese breeds, further indicated that it is fundamental to consider the differences existing among breeds for the prediction of milk coagulation traits.     

Genetic variation in the kappa-casein gene (CSN3) of Chinese yak (Bos grunniens) and phylogenetic analysis of CSN3 sequences in the genus Bos

Variants of κ-casein (CSN3) have been extensively studied in cattle and 13 alleles have been identified at the protein and DNA levels to date. Evolution of some of these alleles and a possible common ancestor remain unclear. Polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) analysis of CSN3 exon IV in domesticated yak revealed a 2-allele polymorphism showing migration patterns different from known cattle variants. The PCR products of both yak CSN3 SSCP alleles were sequenced. All yak had nucleotide sequences coding for Thr in AA position 136 (identical to bovine CSN3*A) and Ala in position 148 (identical to bovine CSN3*B). The sequencing results were confirmed by PCR-RFLP analysis using HindIII and TaqI. A 12-bp insertion in the coding region, representing a repeated nucleotide and AA motif, was found in 1 yak allele. The duplication corresponds to the codons for AA 147 to 150 (Glu-Ala-Ser-Pro) or 148 to 151 (Ala-Ser-Pro-Glu), which are repeated identically. In 21 yak samples genotyped by PCR-SSCP analysis, frequencies for the insertion variant and the short variant were about 68 and 32%, respectively. The loss of the insertion may have led to the ancestral CSN3 allele from which all currently known variants of CSN3 in the genus Bos evolved. This is the first report of polymorphisms in the yak CSN3 gene and may be helpful for future studies on genetic variation within and between yak populations or on associated traits.     

The occurrence of noncoagulating milk and the association of bovine milk coagulation properties with genetic variants of the caseins in 3 Scandinavian dairy breeds

Substantial variation in milk coagulation properties has been observed among dairy cows. Consequently, raw milk from individual cows and breeds exhibits distinct coagulation capacities that potentially affect the technological properties and milk processing into cheese. This variation is largely influenced by protein composition, which is in turn affected by underlying genetic polymorphisms in the major milk proteins. In this study, we conducted a large screening on 3 major Scandinavian breeds to resolve the variation in milk coagulation traits and the frequency of milk with impaired coagulation properties (noncoagulation). In total, individual coagulation properties were measured on morning milk collected from 1,299 Danish Holstein (DH), Danish Jersey (DJ), and Swedish Red (SR) cows. The 3 breeds demonstrated notable interbreed differences in coagulation properties, with DJ cows exhibiting superior coagulation compared with the other 2 breeds. In addition, milk samples from 2% of DH and 16% of SR cows were classified as noncoagulating. Furthermore, the cows were genotyped for major genetic variants in the α_S1- (CSN1S1), β- (CSN2), and κ-casein (CSN3) genes, revealing distinct differences in variant frequencies among breeds. Allele I of CSN2, which had not formerly been screened in such a high number of cows in these Scandinavian breeds, showed a frequency around 7% in DH and DJ, but was not detected in SR. Genetic polymorphisms were significantly associated with curd firming rate and rennet coagulation time. Thus, CSN1S1 C, CSN2 B, and CSN3 B positively affected milk coagulation, whereas CSN2 A², in particular, had a negative effect. In addition to the influence of individual casein genes, the effects of CSN1S1-CSN2-CSN3 composite genotypes were also examined, and revealed strong associations in all breeds, which more or less reflected the single gene results. Overall, milk coagulation is under the influence of additive genetic variation. Optimal milk for future cheese production can be ensured by monitoring the frequency of unfavorable variants and thus preventing an increase in the number of cows producing milk with impaired coagulation. Selective breeding for variants associated with superior milk coagulation can potentially increase raw milk quality and cheese yield in all 3 Scandinavian breeds.     

Short communication: Influence of composite casein genotypes on additive genetic variation of milk production traits and coagulation properties in Holstein-Friesian cows

The aim of the study was to quantify the effects of composite β- and κ-casein (CN) genotypes on genetic variation of milk coagulation properties (MCP); milk yield; fat, protein, and CN contents; somatic cell score; pH; and titratable acidity (TA) in 1,042 Italian Holstein-Friesian cows. Milk coagulation properties were defined as rennet coagulation time (RCT) and curd firmness (a₃₀). Variance components were estimated using 2 animal models: model 1 included herd, days in milk, and parity as fixed effects and animal and residual as random effects, and model 2 was model 1 with the addition of composite β- and κ-CN genotype as a fixed effect. Genetic correlations between RCT and a₃₀ and between these traits and milk production traits were obtained with bivariate analyses, based on the same models. The inclusion of casein genotypes led to a decrease of 47, 68, 18, and 23% in the genetic variance for RCT, a₃₀, pH, and TA, respectively, and less than 6% for other traits. Heritability of RCT and a₃₀ decreased from 0.248 to 0.143 and from 0.123 to 0.043, respectively. A moderate reduction was found for pH and TA, whereas negligible changes were detected for other milk traits. Estimates of genetic correlations were comparable between the 2 models. Results show that composite β- and κ-CN genotypes are important for RCT and a₃₀ but cannot replace the recording of MCP themselves.     

Milk protein genetic variants and isoforms identified in bovine milk representing extremes in coagulation properties

A gel-based proteomic approach consisting of 2-dimensional gel electrophoresis coupled with mass spectrometry was applied for detailed protein characterization of a subset of individual milk samples with extreme rennet coagulation properties. A milk subset with either good or poor coagulation abilities was selected from 892 Danish Holstein-Friesian and Jersey cows. Screening of genetic variants of the major milk proteins resulted in the identification of common genetic variants of β-casein (CN; A(1), A(2), B), κ-CN (A, B), and β-lactoglobulin (LG; A, B), as well as a low frequency variant, κ-CN variant E, and variants not previously reported in Danish breeds (i.e., β-CN variant I and β-LG variant C). Clear differences in the frequencies of the identified genetic variants were evident between breeds and, to some extent, between coagulation groups within breeds, indicating that an underlying genetic variation of the major milk proteins affects the overall milk coagulation ability. In milk with good coagulation ability, a high prevalence of the B variants of all 3 analyzed proteins were identified, whereas poorly coagulating milk was associated with the β-CN variant A(2), κ-CN variant A or E, and β-LG variant A or C. The β-CN variant I was identified in milk with both good and poor coagulation ability, a variant that has not usually been discriminated from β-CN variant A(2) in other studied cow populations. Additionally, a detailed characterization of κ-CN isoforms was conducted. Six κ-CN isoforms varying in phosphorylation and glycosylation levels from each of the genetic variants of κ-CN were separated and identified, along with an unmodified κ-CN form at low abundance. Relative quantification showed that around 95% of total κ-CN was phosphorylated with 1 or 2 phosphates attached, whereas approximately 35% of the identified κ-CN was glycosylated with 1 to 3 tetrasaccharides. Comparing isoforms from individual samples, we found a very consistent κ-CN isoform pattern, with only minor differences in relation to breed, κ-CN genetic variant, and milk coagulation ability.     

Use of the Foodtexture Puff Device to monitor milk coagulation

The further automation of cheese-making on an industrial level requires the development of sensor devices to monitor the gelation process and especially the firming phase. In this paper, the Foodtexture Puff Device (FPD) is tested for its ability to monitor the gelation process by comparing it with classical rheometry (G′ and G″) in a series of coagulations at different initial milk pH (6.01 to 6.61). The FPD measures the deformation of the surface of the milk during coagulation after applying an air puff directed on this surface. The maximal and minimal deformation values and the deformation range were calculated. A nonlinear model of the registered characteristics with the time point from adding rennet until the end of the gelation process was fitted on the FPD data and also on the classic rheology parameters. It was concluded that the FPD monitored the coagulation process in the same way as the rheology. Moreover, the start point of the coagulation process as well as the strength of the coagulum could be estimated nondestructively. Therefore, the presented technology together with the nonlinear model may be a basis for the development of an industrial monitoring device.     

Variation in milk protein concentrations associated with genetic polymorphism and environmental factors

Concentrations of alpha s-casein, beta-casein, kappa-casein, beta-lactoglobulin, alpha-lactalbumin, serum albumin, and immunoglobulin in milk from 1888 Holstein cows were determined monthly over the lactation period. Cows were phenotyped for genetic variants of alpha s1-casein, beta-casein, kappa-casein, and beta-lactoglobulin. Least squares analyses showed variations in individual proteins due to parity number, month of test, stage of lactation, somatic cell count, fat content, milk yield, and phenotypes of cows for milk proteins. beta-Casein declined and serum proteins increased with advancing age of cows. Concentration of individual proteins decreased during the first 2 to 3 mo in lactation and then increased as lactation progressed. alpha s1-Casein variants significantly affected concentrations of alpha s-casein (BC greater than BB greater than AB) and beta-lactoglobulin (AB greater than BB greater than BC). Variant B for beta-casein is associated with lower alpha s-casein, beta-lactoglobulin, immunoglobulins, and higher beta-casein and alpha-lactalbumin concentrations than variant A1, A2, or A3. Milk from BB kappa-casein, and BB beta-lactoglobulin cows contained more alpha s-casein, kappa-casein, and less beta-lactoglobulin than milk from AA cows for the two proteins. Concentrations of all proteins were negatively correlated with milk production. Increased somatic cell counts were associated with lower beta-casein and higher concentrations of other proteins. Fat content of milk was positively correlated with the three casein fractions and beta-lactoglobulin.     

Polymorphic AP-1 binding site in bovine CSN1S1 shows quantitative differences in protein binding associated with milk protein expression

Polymorphisms in 5′-flanking regions of milk protein encoding genes can influence the binding activity of the affected response elements and thus have an impact on the expression of the gene products. However, precise quantitative data concerning the binding properties of such variable response elements have so far not been described. In this study we present the results of a quantitative fluorescent electromobility shift assay comparing the allelic variants of a polymorphic activator protein-1 binding site in the promoter region of the bovine α_s1-casein encoding gene (CSN1S1), which is affected by an A→G exchange at −175 bp (CSN1S1^−175bp). A supershift assay using a commercial c-jun antibody was carried out to verify the specificity of protein binding. The gel shift analysis revealed specific and significantly reduced protein binding of oligonucleotides containing the G variant of the CSN1S1^−175bp binding site. Further investigations comprised genotyping of the variable CSN1S1^−175bp activator protein-1 element by an NmuCl restriction fragment length polymorphism in 62 cows of the breed Simmental and 80 cows of the breed German Holstein. Single milk proteins from at least 4 milk samples per cow were quantified by alkaline urea polyacrylamide gel electrophoresis. Homozygotes for CSN1S1^−175bp*G were not observed, and the allele frequencies were 0.19 in Simmental and 0.05 in German Holstein. Carriers of CSN1S1^−175bp*G showed higher content (%) as well as quantity (g/d) of α_s1-casein than CSN1S1^−175bp*A homozygotes, independent of breed. We assume that the positive association of the CSN1S1^−175bp*G variant with CSN1S1 expression is likely to be caused by a reduced affinity of the affected response element to a c-jun-containing CSN1S1 dimer with repressor properties.     

Effect of milk protein genetic polymorphisms on rennet and acid coagulation properties after standardisation of protein content

The effects of milk protein genetic polymorphisms on the rennet and acid coagulation properties of milk after protein standardisation were investigated. Skim milk samples were adjusted to a protein concentration of 6.07 ± 0.06% by ultrafiltration (UF) before evaluating rennet coagulation and acid coagulation properties. Only the β-lactoglobulin (β-LG) genotypes influenced the rennet-clotting time before standardisation for the total protein concentration by UF; however, this effect was confounded with the β-LG concentration. After UF-concentration, a similar protein concentration between the samples was achieved in the retentate, then the rennet clotting time and rennet curd firmness at 30 min were significantly influenced by both the κ-casein (κ-CN) and β-LG genotypes. κ-CN genotypes significantly influenced the acid coagulation properties of both skim milk and retentate. Variations in the concentration of milk proteins (mostly α_S2-CN-12P) explained most of the differences in the rennet and acid coagulation properties of milk after protein standardisation by UF.     

Short communication: estimates of genetic variation of milk fatty acids in US Holstein cows

Interest in changing the milk fatty acid profile is growing. However, little is known about the genetic variability of milk fatty acids in the US Holstein population. Therefore, genetic parameters for milk fatty acids were estimated using a single-trait, mixed, linear animal model on 592 individual milk samples from 233 daughters of 53 sires in a cow herd genetically representative of the US Holstein population. Heritability (h²) and repeatability (r) estimates ± standard errors for yields of individual fatty acids ranged from 0.00 ± 0.08 (C4:0) to 0.43 ± 0.13 (C12:0) for heritabilities and from 0.21 ± 0.05 (C18:1) to 0.43 ± 0.05 (C12:0) for repeatabilities. Saturated (h² = 0.23 ± 0.12; r = 0.36 ± 0.05) and de novo synthesized fatty acids (C6:0 to C14:0; h² = 0.30 ± 0.13; r = 0.40 ± 0.05) had numerically higher estimates than did monounsaturated (h² = 0.09 ± 0.09; r = 0.22 ± 0.05) and polyunsaturated fatty acids (h² = 0.08 ± 0.09; r = 0.27 ± 0.05). For relative proportions of individual fatty acids, the greatest heritability and repeatability estimates were obtained for C8:0 (h² = 0.18 ± 0.12; r = 0.36 ± 0.05), C10:0 (h² = 0.22 ± 0.13; r = 0.46 ± 0.05), C12:0 (h² = 0.18 ± 0.12; r = 0.46 ± 0.05), C16:0 (h² = 0.09 ± 0.12; r = 0.48 ± 0.05), C16:1 (h² = 0.49 ± 0.13; r = 0.49 ± 0.05), and C18:0 (h² = 0.24 ± 0.11; r = 0.39 ± 0.05). Our results suggest the existence of genetic variability of milk fatty acids, in particular of medium-and long-chain fatty acids (C8:0 to C18:0), which could be used to improve the nutritional and textural properties of milk fat by selective breeding.