CSN3 Gene Research Articles

Influence of polymorphism of complex of csn3 and lgb genes on the dairy productivity in holstein cattle

The study presents the results obtained in the course of DNA testing of allelic polymorphism by PCR-RFLP on CSN3 and LGB genes and the associations of their complex genotypes to the indicators of milk productivity of Holstein cow-heifers.Genetic polymorphism was detected by digestion with the endonucleases Hinf I (CSN3) and Hae III (LGB). All integrated CSN3/LGB genotypes were identified during the work. The maximum frequency of occurrence is observed in a combination of heterozygous AB/AB genotypes – 120 animals (25.5%), which is the fourth part of the studied cattle population. The group of animals with homozygous BB/BB genotypes of CSN3/LGB genes is the smallest one – 10 animals (2.1%). Milk productivity test, based on the complex CSN3 and LGB genotypes is observed in heifers with an integrated combination of CSN3BBLGBAAshowed that a significantly high level of productivity for 305 days of lactation – 7239.8 kg, and a minimum level of productivity (6222.9 kg) is observed in heifers with the complex CSN3BBLGBAB genotype, compared with indicators of milk yield of cow-heifers with other integrated genotypes. According to the content of the mass fraction of fat, the best indicators were established in the subpopulation of animals with the complex AA/AB genotype – 3.90%, and the worst result was recorded in the group of animals with the complex AB/AA genotype, which amounted to 3.44%. The high content of the mass fraction of protein was observed in individuals by the complexof the CSN3 and LGB genes BB/AA – 3.43%, and a reduced level of protein was observed in heifers with a combination of homozygous AA/AA and BB/BB genotypes by the genes under study – 3.18%. Thus, individuals carrying the homozygous A allele of the kappa-casein (CSN3) gene and the homozygous B allele of the beta-lactoglobulin (LGB) gene in the integrated genotype favorably differed on the main indicators of milk productivity.

СЫРОПРИГОДНОСТЬ МОЛОКА ЧЕРНО-ПЕСТРЫХ КОРОВ С РАЗНЫМИ ГЕНОТИПАМИ КАППА-КАЗЕИНА И ДИАЦИЛГЛИЦЕРОЛ О-АЦИЛТРАНСФЕРАЗЫ

The purpose of the research is improving the quality and technological properties of milk from black-and-white cows. The research was conducted on first-calf cows of a black-and-white breed in LLC «Dusym» of the Atninsky district of the Republic of Tatarstan. Three groups of experimental cows were formed depending on the genotype of CSN3 and DGAT1, which have equal lactation month. Cheese making milk obtained from cows with different allelic variants of the kappa-casein and diacylglycerol O-acyltransferase genes was studied. DNA testing was performed on the CSN3 and DGAT1 genes of experimental cows and allelic variants of these genes were determined by pol-ymerase chain reaction followed by analysis of restriction-site polymorphism. It was found that milk of cows with the genotype CSN3 AB and CSN3 BB, when exposed to rennet, showed a better yield of a dense casein clot (80 and 100%) and separation of serum (P <0.05), with a shorter coagulation time (20.3 and 16.5 min; P <0.05-0.01). Ac-cording to the DGAT1 gene, first-calf milk with the allelic variant DGAT1 AK and DGAT1 KK had the advantage, respectively, 86.7% and 85.7%, 24.0 and 25.7 minutes. The worst milk coagulation time is typical for CSN3 AA group – 29.5 minutes and DGAT1 AA one – 30.4 min. In terms of milk coagulation time, type II (15-40 min.) groups have the genotype CSN3 AB and CSN3 BB with the largest share of cows in the herd – 80.0 and 71.4% and with the genotype DGAT1 AK and DGAT1 KK – 86.6 and 85.7%, respectively. Thus, milk obtained from cows with the kappa-casein gene allele B and the diacylglycerol O-acyltransferase gene allele K gene in the genotype has the best cheese-making properties.

Investigation on the suitability of milk-derived primary bovine mammary epithelial cells grown on permeable membrane supports as an in vitro model for lactation.

This study aimed to establish an in vitro model for lipid synthesis in primary bovine mammary epithelial cells (pbMECs) extracted from milk and cultured on Transwell permeable supports (TW culture). The suitability of these cells as a functional model for lactation was assessed by measuring κ-casein (CSN3) and diacylglycerol acyl transferase 1 (DGAT1) gene expression, the presence of intracellular lipid droplets, and the concentration of triacylglycerol in the cell lysates. The functionality of the milk-derived pbMECs cultured under lactogenic conditions, with and without oleic acid supplementation, was evaluated by comparing the cells grown on Transwell supports to cells grown on an extracellular matrix (ECM) gel (3D culture) or a plastic surface (2D culture). Furthermore, the functionality of milk-derived cells was compared to pbMECs obtained from bovine mammary tissue. Here, we show that in both tissue and milk-derived pbMECs, 3D culture offered the most suitable in vitro environment and led to increased levels of CSN3 and DGAT1 gene expression along with increased intracellular triacylglycerol content. The TW culture conditions also resulted in increased DGAT1 gene expression compared to the 2D conditions and milk-derived pbMECs cultured on TW inserts showed the highest viability compared to cells grown under 2D or 3D treatments. However, this was not observed for tissue-derived pbMECs, suggesting that TW culture may offer a beneficial environment specifically for milk-derived cells. We suggest that with further optimization of the culture conditions, TW culture may present a suitable model for the study of milk lipid synthesis in pbMECs.

PCR Based Genotyping of Lulu Cattle of Nepal for A1, A2 Type Beta-caseins

Lulu is an indigenous breed of cattle (Bos taurus) found in high altitude regions of western Nepal. Population of Lulu cattle has been declining due to introgression with other exotic breeds to increase milk productivity. Here we aimed at finding potential approach for conserving Lulu cattle and its assets by studying the milk contents and investigating which variant of beta-casein protein is present in this breed. Beta caseins are an abundant protein in cow milk with A1 and A2 being the most common genetic variants of this protein. Consumption of A1 type of milk has numerous health-related complications whereas A2 type of milk has numerous human health promoting factors. We used restriction fragment length polymorphism (RFLP) for determining the A1 and A2 variant of beta casein in Lulu cattle. For performing DNA extraction, we collected (n = 18) blood samples of Lulu from Mustang and (n=17) Nepal Agriculture research council farm. The amplified fragments in 3% agarose at 251bp and 213bp respectively confirmed the presence of both A1 and A2 gene in Lulu; however, A2 was of greater abundance. Our study indicated that Lulu has A2 variant of beta-casein predominantly. The gene frequency of A1A1 is 0, A1A2 is 0.06 and A2A2 is 0.94. We further found that the allele frequency of A1 and A2 is 0.03 and 0.97 respectively. We designed special primer for sequencing CSN2 genes since A2 type beta casein gene was predominantly seen on Lulu. The sequencing result further supports our RFLP result as most of our samples have “C” nucleotide SNP in amplified CSN2 gene sequence. The Chi-square value of the current study is 0.04 which supports Hardy-Weinberg equilibrium inferring that Lulu cattle are still in the pure state, where there is no genetic introgression with the exotic breed for the sake of improvement of productivity.

Allele and Genotype Frequencies of the Κappa-Casein (CSN3) Locus in Macedonian Holstein-Friesian Cattle

Abstract The bovine kappa-casein (κ-CN) is a phospho-protein with 169 amino acids encoded by the CSN3 gene. The two most common gene variants in the HF breed are CSN3*A and CSN3*B while CSN3*E has been found with lower frequency. The aim of this study was to optimize a laboratory method for genotyping of these three alleles as well as to determine their genotype and allele frequencies in the HF cattle population in the Republic of North Macedonia. Genomic DNA was extracted from full blood from 250 cows. The target DNA sequence was amplified with newly designed pair of primers and the products were subjected to enzymatic restriction with HindIII and HaeIII endonucleases. Genotype determination was achieved in all animals. The primers successfully amplified a fragment of 458 bp and the digestion of this fragment with both endonucleases enabled differentiation of five different genotypes with the following observed frequencies: AA (0.39), AB (0.29), BB (0.16), AE (0.10), and BE (0.06). The estimated allele frequencies were: CSN3*A (0.584), CSN3*B (0.336) and CSN3*E (0.08). The observed genotype frequencies differed significantly (P<0.01) from those that would be expected under HW equilibrium, while the fixation index (F=0.17) indicated moderate heterozygosity deficiency. Nevertheless, the CSN3*B allele was present with relatively high frequency which should be used to positively select for its carriers, since increasing its frequency could help to improve the rheological properties of the milk intended for cheese production.

A meta-analysis on association between CSN3 gene variants and milk yield and composition in cattle.

A meta-analysis on the effects of A and B alleles, the most frequent alleles of CSN3 gene, on milk yield and composition traits was conducted by pooling a large dataset consisting of 30 471 genotyped cattle. Four genetic models, comprising dominant (AA+AB vs. BB), recessive (AA vs. AB+BB), additive (AA vs. BB) and co-dominant (AA+BB vs. AB), were employed to analyze data. Standardized mean difference (SMD) was used to measure the size of the effects of A and B alleles of CSN3 on studied traits. Effect sizes of 0.2, 0.5 and 0.8 represent small, medium and large effects, respectively. The results indicate that B allele, in the form of BB genotype, has a significant, but medium effect on lactation yield under dominant (SMD=0.259, P-value=0.006) and additive (SMD=0.279, P-value=0.035) models. Moreover, a small decrease in the fat percentage occurred in cows having A allele under dominant (SMD=-0.077, P-value=0.006) and additive (SMD=-0.106, P-value=0.035) models. Furthermore, CSN3 variants significantly but slightly affect protein percentage under dominant (SMD=-0.146, P-value=0.000), recessive (SMD=-0.077, P-value=0.000) and additive (SMD=-0.219, P-value=0.000) models, showing the negative effect of A allele on this trait. Meta-analysis results reveal that daily milk yield is slightly affected by CSN3 variants under recessive (SMD=0.056, P-value=0.033) and additive (SMD=0.061, P-value=0.013) genetic models. There is no effect of CSN3 variants on either protein or fat yield. In addition, the effects of CSN3 variants on milk-related traits were not observed under the co-dominant model. Sensitivity and publication bias analyses were carried out to confirm the stability of meta-analyses results.

Frequencies Evaluation of β-Casein Gene Polymorphisms in Dairy Cows Reared in Central Italy.

Simple SummaryBovine milk contains several β-casein variants, with the A1 and A2 variants occurring most frequently. The presence of some variants, such as A1, B, and C, is considered a risk factor for disease in humans who consume milk. These variants are probably involved in intolerance to milk and some human diseases due to the production of a bioactive peptide with opioid activity during digestion, β-casomorphin 7 (BCM-7). In contrast, the A2 variant is not involved in pathogenetic mechanisms; thus, its presence in milk is a desirable feature. The difference between the A1 and A2 variants is a mutation at position 67 of the β-casein gene (CSN2), which causes an amino acid to change from histidine (in the A1, B, and C variants) to proline (in the A2 variant). To select dairy cows on the basis of the presence of the β-casein variant A2, allele frequencies of CSN2 variants were evaluated in Italian dairy cows reared in central Italy. The results of this study may help with the selection of animals with the β-casein gene variant A2 to produce a more digestible milk that only contains the β-casein variant A2.The majority of proteins in cow’s milk are caseins, which occur in four groups (α-s1, α-s2, β, and k) encoded by different genes (CSN1S1, CSN1S2, CSN2, and CSN3, respectively). In this study, we focused on the β-casein allele variants A1 and A2 due to their influence on milk’s technological characteristics and human health. Digestion of the β-casein variant A1 leads to the formation of β-casomorphin 7 (BCM-7), a bioactive peptide that has been suggested to be a possible cause of various human diseases and associated with low milk digestibility. The potential negative role of the β-casein variant A1 in human health has stimulated the planning of cattle breeding programs based on genetic selection to increase the frequency of the A2 variant, which is associated with increased milk digestibility. The aim of this work was to evaluate the frequencies of the different β-casein variants in Italian Holstein Friesian dairy cows from cattle farms located in central Italy to select a population of A2 homozygous animals. β-casein genotypes were identified by evaluating the presence of single nucleotide polymorphisms (SNPs) of the CSN2 gene using PCR and sequencing analysis. The frequency of the desirable β-casein variant A2 in the studied bovine population was 0.61. The frequency of the undesirable A1 variant in the studied bovine population was 0.30. The frequency of the A2 allele was higher than expected for the breed; therefore, genetic selection for the A2 variant in these animals could be achieved in a fairly short time using A2 homozygous bulls.

Genotyping of selection-significant polymorphisms of cattle of the Western Siberia

The article presents the results of genotyping of 156 Simmental cows for the genes of kappa casein (CSN3), beta-lactoglobulin (BLG), alpha-lactalbumin (LALBA) and leptin (LEP). Studies have shown that the population for all genes is in a state of equilibrium distribution of genotypes, which indicates the absence of factors that change allele frequencies. For the CSN3 gene, the frequency of genotypes A/A, A/B, and B/B is 0.583, 0.385, and 0.032; alleles A and B - 0.776 and 0.224. For the BLG gene, the frequency of the corresponding genotypes is 0.449, 0.455, and 0.096; alleles A and B - 0.676 and 0.324. The occurrence of the LALBA gene genotypes is as follows: A/A – 0.513, A/B – 0.429, B/B – 0.058; allele A – 0.728, allele B – 0.272. Distribution of genotypic classes of the LEP gene: C/C – 0.429, C/T – 0.481, T/T – 0.09; frequency of alleles C and T – 0.67 and 0.33. The observed population structure with a low frequency of B/B genotypes for the CSN3 and BLG genes associated with protein-milkability, but a relatively high level of heterozygosity, demonstrates a stable equilibrium of allele frequencies, which is possible in the case of selection in favor of heterozygotes.

Productive longevity of white-and-black cows of different genotypes CSN3 and DGAT1

The article studies the duration of use and life-long productivity black-motley breed cows with different allelic variants of the kappa-casein and diacylglycerol O-acyltransferase genes. The research consists the data study on cows dropped out the herd during 2012–2016 in the Dusym LLC of Atninsky District in the Republic of Tatarstan. The cows were previously genotyped for CSN3 and DGAT1. The herd of black-and-white cattle consisted the genotypes CSN3 AA and DGAT1 AK with a frequency of 60.9 and 53.5 % and less often CSN3 BB and DGAT1 KK – 4.0 and 6.3 %. Moreover, the frequency of occurrence of the desired B allele of the CSN3 gene was 0.22, and the K allele of DGAT1 was 0.33. Cows with the CSN3 AB genotype reliably surpass animals with the CSN3 AA genotype in the duration of use by 0.39 lactations (P <0.05), life-long milk yield by 2401 kg of milk (P <0.05), and the amount of milk fat by 90 kg (P <0.05), mass fraction of protein in milk – by 0.04 % (P <0.01), the amount of milk protein – by 83 kg (P <0.05). Animals with the DGAT1 KK genotype also have an excess over other groups of cows in terms of productive longevity, but the difference is significant only in fat milk (P <0.01–0.001).

Влияние полиморфных вариантов гена CSN3 на технологические свойства молока

Влияние полиморфных вариантов гена CSN3 на технологические свойства молока

Polymorphisms of the kappa casein (<i>CSN</i>3) gene and inference of its variants in water buffalo (<i>Bubalus bubalis</i>)

Kappa casein plays a crucial role in the formation of stable casein micelles and has a key influence on milk-clotting properties. However, current understanding of buffalo CSN3 gene polymorphisms is not sufficient. In this study, the polymorphisms in the complete coding sequence (CDS) of the buffalo CSN3 were detected using PCR product direct sequencing. The CDS of CSN3 for river and swamp buffalo was the same in length, which contained an open reading frame of 573 nucleotides encoding a peptide containing 190 amino acid residues. A total of eight single nucleotide polymorphisms (SNPs) was identified in two types of buffalo. Among them, c.86C>T, c.252G>C, c.445G>A, c.467C>T and c.516A>C were non-synonymous, which leads to p.Pro8Leu, p.Lys63Asn, p.Val128Ile, p.Thr135Ile and p.Glu151Asp substitutions in buffalo kappa casein (-CN), respectively. The substitution of p.Thr135Ile may exert a vital effect on the function of buffalo -CN. Eleven haplotypes were defined based on the SNPs found in buffalo, and accordingly, seven protein variants and four synonymous variants of buffalo -CN were inferred, called variants A, B, B, C, C, C, D, E, F, F and G. The variants observed in water buffalo did not exist in the Bos genus. In addition, 14 amino acid differential sites of -CN between buffalo and the Bos genus were identified, of which 3 were located at glycosylation sites (80S, 96T, 141S) and 4 at phosphorylation sites (19S, 80S, 96T, 141S). It is speculated that they may lead to differences in the physicochemical properties of -CN between buffalo and the Bos genus. This study will lay a foundation for exploring the association between the variation in the CSN3 gene and the lactation traits of buffalo.

DNA Sequence Variants and Protein Haplotypes of Casein Genes in German Black Pied Cattle (DSN).

Casein proteins were repeatedly examined for protein polymorphisms and frequencies in diverse cattle breeds. The occurrence of casein variants in Holstein Friesian, the leading dairy breed worldwide, is well known. The frequencies of different casein variants in Holstein are likely affected by selection for high milk yield. Compared to Holstein, only little is known about casein variants and their frequencies in German Black Pied cattle (“Deutsches Schwarzbuntes Niederungsrind,” DSN). The DSN population was a main genetic contributor to the current high-yielding Holstein population. The goal of this study was to investigate casein (protein) variants and casein haplotypes in DSN based on the DNA sequence level and to compare these with data from Holstein and other breeds. In the investigated DSN population, we found no variation in the alpha-casein genes CSN1S1 and CSN1S2 and detected only the CSN1S1*B and CSN1S2*A protein variants. For CSN2 and CSN3 genes, non-synonymous single nucleotide polymorphisms leading to three different β and κ protein variants were found, respectively. For β-casein protein variants A 1, A 2, and I were detected, with CSN2*A 1 (82.7%) showing the highest frequency. For κ-casein protein variants A, B, and E were detected in DSN, with the highest frequency of CSN3*A (83.3%). Accordingly, the casein protein haplotype CSN1S1*B-CSN2*A 1-CSN1S2*A-CSN3*A (order of genes on BTA6) is the most frequent haplotype in DSN cattle.

A Novel Test System for Genotyping rs43703016 Single-nucleotide Substitutions in the Bovine CSN3 Gene

Aims: Caseins are among the main milk proteins that determine many of its properties. Bovine kappa-casein (CSN3) is associated with the qualitative composition of milk, as well as with the quality of cheese obtained from this milk. The rs43703016 single-nucleotide substitution (g.88532332A>C; Asp148Ala) in exon 4 of the bovine CSN3 gene plays an important role in the production of quality hard cheeses. Various methods for the DNA testing of this substitution have been developed in the last three decades. Emergent DNA technologies provide an opportunity to modernize methods of genotyping single-nucleotide polymorphisms.
 Results: We have developed and verified a method to differentiate A/C alleles of the rs43703016 substitution in the bovine CSN3 gene by real-time PCR using allele-specific fluorescent probes.
 Conclusion: Our new method allows fast genotyping of animals, and may be used for selection of cows carrying the CC genotype, which determines good cheese-making properties of milk.

Casein haplotype diversity in seven dairy goat breeds.

Selection, drift, gene flow and breeding have extensively shaped the genomic variability of domestic animals. In goat species, several mutations identified within the casein genes have been shown to affect the level of gene expression of milk production traits. The four casein genes – CSN1S1, CSN2, CSN1S2 and CSN3 – are organized in a cluster of 250 kb located in chromosome 6, and due to tight linkage, their genetic variability is well depicted by haplotypes which are transmitted to the progeny. Thirty single nucleotide polymorphisms (SNPs) located within the casein gene cluster were used to characterize the haplotype variability of six southern Italian goat breeds (Girgentana, Maltese, Rossa Mediterranea, Argentata dell'Etna, Messinese, Capra dell'Aspromonte). A representative sample of the Norwegian dairy goat breed (Norsk melkegeit) has been used as an out-group to obtain a weighted measure of genetic diversity in the metapopulation. A total of 54 haplotypes were detected among the seven breeds: 26, 9, 8 and 11 haplotypes were found at CSN1S1, CSN2, CSN1S2 and CSN3 respectively. The number of haplotypes per breed was 14 (Norwegian), 26 (Messinese), 27 (Rossa Mediterranea and Girgentana) and 31 (Maltese, Argentata dell'Etna and Capra dell'Aspromonte). The Maltese breed showed the highest number of private haplotypes, whereas the Norwegian goat recorded the highest number of shared haplotypes. The linkage disequilibrium analysis showed higher levels of association for the SNP pairs within casein loci than SNP pairs between casein loci, likely reflecting low levels of intra-genic recombination. The highest linkage disequilibrium values were found in CSN1S1 and CSN2 genes in all the breeds, except for Argentata dell'Etna and Rossa Mediterranea. The resolution of the haplotype diversity at the casein cluster can be exploited both for selective and conservative plans.

THE ANALYSIS OF POLYMORPHISM OF KAPPA-CASEIN, Β-LACTOGLOBULIN AND PROLACTIN GENES AMONG YAKUTIAN CATTLE AND ITS INFLUENCE ON MILK PRODUCTION

DNA samples of cows of Simmental and Kholmogory breeds served as the material for the research. Release of DNA was carried out by a standard method of phenolchloroform extraction. Polymorphisms of rs43703015 of the CSN3 gene, rs109625649 of the LGB gene and rs134028641 of the PRL gene are studied by means of PCR-RFLP analysis. In all studied selections, the A allele prevails on the CSN3 gene. The frequency of the A allele prevailed over the frequency of the B allele on the PRL gene. The animals with the greatest milk yield and fat content of milk on the CSN3 gene had the genotype including the A allele. Depending on the LGB genotype, there was established that the largest size of milk yields have individuals with the AA and AB genotype in selections of the Simmental breed of both selections. In selection of the Kholmogory breed - with the B genotype. Adaptation process is fraught with a serious pressure, that, undoubtedly, affects the efficiency; while long-acting it leads to disorder of physiological functions. Therefore, realization of genetic potential of the delivered cattle to Yakutia will require time.

Evaluation of candidate gene effects and environmental factors on reproductive performance of Holstein cows

This study investigated the impact of nine polymorphisms located in the CSN2 , CSN3 , CSN1S1 , CSN1S2 , OLR1 , LALBA , STAT1 , DGAT1 and LGB genes, and environmental factors including calving year, season and parity on reproductive traits. The analysis was conducted on 165 Holstein-Friesian cows. Genotypes were identified using PCR-RFLP. The data of reproductive traits for four lactations were evaluated. Statistical analysis was carried out using least squares of the GLM procedures. Results indicated that CSN2 had significant effects on days before first insemination and first insemination to pregnancy interval. The SNP at the CSN3 was significantly associated with gestation length. A novel effect of OLR1 -C223A on age at first calving was observed in the present study. Moreover, DGAT1 and LGB markers were significantly associated with calving interval and days before first insemination, respectively. In addition, significant environmental effects were as follows: calving year with days before first oestrus, days open, and first insemination to pregnancy interval; season with days open, first insemination to pregnancy interval, and calving interval; parity with days before first oestrus and days open. The present results and novel associations may therefore be useful and indicative for future studies on a genetic basis of cattle reproduction traits. Keywords: cattle, environmental effects, genetic marker, Holstein-Friesian, reproduction parameters

Genomic analysis of the origins of extant casein variation in goats

The variation in the casein genes has a major impact on the milk composition of goats. Even though many casein polymorphisms have been identified so far, we do not know yet whether they are evolutionarily ancient (i.e., they existed before domestication) or young (i.e., they emerged after domestication). Herewith, we identified casein polymorphisms in a data set of 106 caprine whole-genome sequences corresponding to bezoars (Capra aegagrus, the ancestor of domestic goats) and 4 domestic goat (Capra hircus) populations from Europe, Africa, the Far East, and the Near East. Domestic and wild goat populations shared a substantial number of casein SNP, from 36.1% (CSN2) to 55.1% (CSN1S2). The comparison of casein variation among bezoars and the 4 domestic goat populations demonstrated that more than 50% of the casein SNP are shared by 2 or more populations, and 18 to 44% are shared by all populations. Moreover, the majority of casein alleles reported in domestic goats also segregate in the bezoar, including several alleles displaying significant associations with milk composition (e.g., the A/B alleles of the CSN1S1 and CSN3 genes, the A allele of the CSN2 gene). We conclude that much of the current diversity of the caprine casein genes comes from ancient standing variation segregating in the ancestor of modern domestic goats.

Polymorphisms in CSN3, CSN2 and LGB Genes and Their Relation to Milk Production in Dairy Cattle in the Czech Republic

The principal objective of this study was to perform genotyping of the CSN3, CSN2 and LGB genes in selected dairy cattle breeds in the Czech Republic, to determine genotype and allele frequencies and to perform an associative analysis on milk yield in kg, the percentage of fat and protein content, and fat and protein production in kg. The AA was the most frequent genotype for CSN3 with a frequency of 0.474 and the A allele (0.694). The BB genotype (0.814) and the B allele (0.895) predominated in CSN2 gene. Other genotyping was performed for the A1 and A2 alleles, the most frequent were the A2A2 genotype (0.475) and the A2 allele (0.680). The heterozygous AB genotype with a frequency of 0.900 was the most frequent in the LGB gene. A statistically significant higher protein content was found in the AB genotype in comparison to the BB in gene for CSN2. In the remaining cases, the effect of CSN3, CSN2 and LGB polymorphic variations on the milk production indicators was not statistically significant.

Genetic diversity of κ-casein (CSN3) and lactoferrin (LTF) genes in the endangered Turkish donkey (Equus asinus) populations.

In this study, the -casein (CSN3) and lactoferrin (LTF) genes which were found in association with milk production traits in different animal species were studied firstly in Turkish donkey populations. A total of 108 donkeys from different regions of Turkey were used in order to reveal the different genotypes of CSN3 and LTF genes by using polymerase chain reaction – restriction fragment length polymorphism (PCR-RFLP) and DNA sequencing methods. To determine the genetic polymorphism, we attempted to digest a fragment of 235 bp of the CSN3 gene and a fragment of 751 bp of the LTF gene using PstI, and DraII, EagI and MboI restriction enzymes, respectively. Neither the CSN3 gene nor the LTF gene had enzyme recognition sites with the PstI, DraII and MboI restriction enzymes in all of the studied samples. However, the LTF gene was only distinguished with the EagI restriction enzyme. Three genotypes were identified in the LTF gene with the EagI restriction enzyme: GG homozygotes (667, 84 bp), AG heterozygotes (751; 667, 84 bp) and AA homozygotes (751 bp). The transition from guanine to adenine in 89 bp of the LTF gene lacks the restriction site and different genotypes are obtained. This novel single nucleotide polymorphism (SNP) has been firstly detected in donkeys. According to the results, the G allele was predominant in the LTF-EagI gene in the studied Turkish donkey populations. In this study, all the genotype distributions of LTF-EagI were not found in Hardy–Weinberg equilibrium (). The CSN3 and LTF genes have not been studied before in donkeys, so the results are the preliminary results of these gene regions in donkeys.

Genome-Wide Association Study for Milk Protein Composition Traits in a Chinese Holstein Population Using a Single-Step Approach.

Genome-wide association studies (GWASs) have been widely used to determine the genetic architecture of quantitative traits in dairy cattle. In this study, with the aim of identifying candidate genes that affect milk protein composition traits, we conducted a GWAS for nine such traits (αs1-casein, αs2-casein, β-casein, κ-casein, α-lactalbumin, β-lactoglobulin, casein index, protein percentage, and protein yield) in 614 Chinese Holstein cows using a single-step strategy. We used the Illumina BovineSNP50 Bead chip and imputed genotypes from high-density single-nucleotide polymorphisms (SNPs) ranging from 50 to 777 K, and subsequent to genotype imputation and quality control, we screened a total of 586,304 informative high-quality SNPs. Phenotypic observations for six major milk proteins (αs1-casein, αs2-casein, β-casein, κ-casein, α-lactalbumin, and β-lactoglobulin) were evaluated as weight proportions of the total protein fraction (wt/wt%) using a commercial enzyme-linked immunosorbent assay kit. Informative windows comprising five adjacent SNPs explaining no < 0.5% of the genomic variance per window were selected for gene annotation and gene network and pathway analyses. Gene network analysis performed using the STRING Genomics 10.0 database revealed a co-expression network comprising 46 interactions among 62 of the most plausible candidate genes. A total of 178 genomic windows and 194 SNPs on 24 bovine autosomes were significantly associated with milk protein composition or protein percentage. Regions affecting milk protein composition traits were mainly observed on chromosomes BTA 1, 6, 11, 13, 14, and 18. Of these, several windows were close to or within the CSN1S1, CSN1S2, CSN2, CSN3, LAP3, DGAT1, RPL8, and HSF1 genes, which have well-known effects on milk protein composition traits of dairy cattle. Taken together with previously reported quantitative trait loci and the biological functions of the identified genes, we propose 19 novel candidate genes affecting milk protein composition traits: ARL6, SST, EHHADH, PCDHB4, PCDHB6, PCDHB7, PCDHB16, SLC36A2, GALNT14, FPGS, LARP4B, IDI1, COG4, FUK, WDR62, CLIP3, SLC25A21, IL5RA, and ACADSB. Our findings provide important insights into milk protein synthesis and indicate potential targets for improving milk quality.