Alpha-lactalbumin Gene Research Articles

Differences between black-and-white cows with alternative genotypes of the alpha-lactalbumin gene in milk features

Differences between black-and-white cows with alternative genotypes of the alpha-lactalbumin gene in milk features

Polymorphism Detection in Alpha-Lactalbumin Gene and its Association with Some Productive Traits in Egyptian Goats

Polymorphism Detection in Alpha-Lactalbumin Gene and its Association with Some Productive Traits in Egyptian Goats

Polymorphism, Molecular Characteristics of Alpha-Lactalbumin (LALBA) Gene in River and Swamp Buffalo

Polymorphism, Molecular Characteristics of Alpha-Lactalbumin (LALBA) Gene in River and Swamp Buffalo

Screening and Association of Polymorphism in Alpha-Lactalbumin Gene in Holstein Friesian Crossbred Cattle

Screening and Association of Polymorphism in Alpha-Lactalbumin Gene in Holstein Friesian Crossbred Cattle

The effect of single-nucleotide polymorphism in the promoter region of bovinealpha-lactalbumin(LALBA) gene onLALBAexpression in milk cells and milk traits of cows

Polymorphisms of milk protein genes have been proposed as candidate markers for dairy production traits in cattle. In the present study, a polymorphism was detected in the 5'-flanking (promoter) region of the bovine alpha-lactalbumin (LALBA) gene, a T/C transition located at nucleotide -1,001 relative to the transcription start site g.-1001T > C (NC_037332.1:g.31183170T > C), which is recognizable with PstI restriction endonuclease. In silico analyses showed that this mutation created novel retinoid X receptor alpha and vitamin D receptor transcription factor binding sites. Real-time PCR found that cows with different genetic variants of the promoter demonstrated different levels of expression of LALBA mRNA in milk somatic cells (MSCs). The TT genotype cows demonstrated low expression, whereas those with CT demonstrated much higher expression (P < 0.05). ELISA analysis found milk LALBA protein levels also differed between the TT and CT cows (P < 0.05) and that these levels were not correlated with the mRNA abundance in MSC. Association analysis found that the g.-1001T > C polymorphism in the promoter region of the LALBA gene influenced milk production traits in Polish Holstein-Friesian cows. High daily milk yield and dry matter yield, and high lactose yield and concentration were associated with the TT genotype. The TT genotype cows also had a lower number of somatic cells in the milk, considered as an indicator of udder health status. Therefore, the TT genotype could be more desirable from the breeder's perspective.

A Novel SNPs in Alpha-Lactalbumin Gene Effects on Lactation Traits in Chinese Holstein Dairy Cows.

Simple SummaryAlpha-lactalbumin (α-LA) is a major whey protein component in mammalian milk, such as human (approximately 36%), bovine (approximately 17%), and other species, etc., It is involved in the regulation of lactose synthesis and has high nutritional value, especially in infant formula. Previous studies have confirmed that bovine α-LA gene 5′-flanking region has single nucleotide polymorphisms (SNPs), but little is known about polymorphisms in other regions, especially sequence coding for amino acids in protein (CDS) and their adjacent non-coding regions, including Chinese Holstein dairy cows. This study focused on investigated SNPs in the CDS and their adjacent non-coding regions of the α-LA gene in Chinese Holstein dairy cows, and assessed the association between SNPs and lactation traits. Sequence alignment showed that a potential SNPs (562th, G/A) in CDS2 region affect protein spatial structure, suggesting that this SNPs might affect the lactation traits of cows (milk type (Holstein and Jersey), and non-milk type (Bos Taurus)) need more in-depth study. More importantly, a novel SNPs at 1847th (T/C) bp in non-coding region near CDS4 was significantly associated with milk lactose composition, and lactose contents were significantly correlated with milk protein content, indicating that the SNPs could be used as a novel potential molecular marker for lactation traits in Chinese Holstein dairy cows.Alpha-lactalbumin (α-LA) is a major whey protein in bovine and other mammalian milk, which regulates synthesis of lactose. Little is known about its genetic polymorphism and whether can be used as a potential marker for dairy ingredients, milk yield traits, and milk properties. To investigate its polymorphisms and their relationship with milk lactation traits in Chinese Holstein dairy cows, single-strand conformation polymorphism method (PCR-SSCP) and direct sequencing method were used to mark the α-LA gene SNPs. AA (0.7402) and AB (0.2598) genotypes were screened out by PCR-SSCP bands analysis in two independent populations. Direct sequencing revealed that there is one SNP at 1847th (T/C) bp in noncoding region of α-LA gene with highly polymorphic (0.5 < PIC = 0.5623 or 0.5822), of which T is in AA genotype while C in AB. Association analysis also showed that lactose content (p < 0.05) was negatively correlated with fat and protein contents within subgroup, indicating that the SNPs (1847th, T/C) in α-LA gene could be used as a novel potential molecular marker for lactation traits in Chinese Holstein dairy cows.

Polymorphism association and expression analysis of alpha-lactalbumin (LALBA) gene during lactation in Nili Ravi buffalo.

Alpha-lactalbumin has been reported as a highly polymorphic gene that potentially alters the gene expression and is associated with milk composition in dairy breeds. Current study was conducted in two phases. In the first phase, polymorphisms identification in alpha-lactalbumin (LALBA) gene and its association with milk composition was performed. To identify the genetic polymorphism, Nili Ravi buffaloes at their second lactation were selected from Government livestock farm (Buffalo Research Institute, Pattoki). Genomic DNA was extracted from blood samples. After PCR amplification, products were sequenced, and data was analyzed. Results showed that the identified polymorphisms at chromosomal position 34310940 were found associated with major whey protein. In the second phase of study, milk samples were collected from five healthy mastitis-free Nili Ravi buffaloes in their second lactation for expression analysis of alpha-lactalbumin gene at their transition (day 15), mid (day 90), and late (day 250) lactation. Gene expression was observed highest in transition phase with a gradual decrease of expression in mid and late phase of lactation. Further studies are needed to explore the regulation of milk production genes and their translational efficiency during the course of lactation.

Discrimination by RT-Nested PCR of Alpha-Lactalbumin Transcript Expression in Mammary Tumors.

Alpha-lactalbumin is a protein of milk expressed by mammary epithelial cells and by some breast tumors. The alpha-lactalbumin gene has two transcripts. The expression of transcript 2 has not yet been studied. The main objective of this paper was to establish the expression profile of alpha-lactalbumin at the mRNA level and to develop a technique discriminating between the two transcripts. The study was performed on 46 fresh mammary biopsies: 17 malignant tumors, 13 benign tumors, and 16 adjacent healthy tissues. We developed an RT-nested PCR to detect LALBA gene expression without distinction between transcripts. We also designed an RT-nested PCR that detects transcript 2. Both nested PCRs avoid amplification of potentially contaminating genomic DNA. We show that benign tumors tend to appear at a young age contrarily to malignant tumors appearing later with p value = 0.002. Moreover, LALBA transcript expression varies among tissues more important in benign (69%) than in malignant (35%) tumors with significant p value = 0.041. The intensity of the bands reflected differential expression between patients with concordant data between both RT-nested PCRs. We also performed a qualitative analysis according to histopathological parameters and molecular subtypes. The expression of transcript 2 appeared associated with the HER2+ subtype, while transcript 1 was associated with the luminal A and triple-negative subtypes. In conclusion, we designed specific and sensitive methods to detect LALBA transcripts. We show for the first time a differential expression of these transcripts, but we need to confirm their potential use as markers in breast cancer.

Generating Goat Mammary Gland Bioreactors for Producing Recombinant Proteins by Gene Targeting.

Exogenous genes can be site-specifically integrated into the genomic DNA of animals by homologous recombination, generating transgenic animals. These animals have a clear hereditary background, although position effects of the exogenous genes and potential functional disruption of host genes can be caused by the genetic inserts. Therefore, the generation of mammary gland bioreactors via gene-targeting methods is a great asset for producing recombinant proteins in milk. Here, we describe a method of generating gene-targeted goats with the human alpha-lactalbumin gene (hα-LA) integrated into the beta-lactoglobulin gene (BLG) locus. The milk from these goats will be less allergenic and will be enriched with components of human milk protein.

Allele mining in the caprine alpha-lactalbumin (LALBA) gene of native Saudi origin

ABSTRACTAlpha-lactalbumin (α-LA) is a distinctive whey protein found in bovine milk and the milk of other mammalian species. Its main function in the lactating mammary glands is involved in lactose biosynthesis, which makes it a potential quantitative trait locus. The aim of this study is to determine the incidence of single nucleotide polymorphisms (SNPs) within the exonic and intronic portions of the caprine α-LA gene (LALBA) in native Saudi breeds (Ardi, Habsi and Harri) in relation to utility traits. Blood samples were collected from 300 animals (100 for each breed). Genomic DNA was extracted and a 268 bp fragment of the LALBA gene comprising exon 3 and its flanking area was amplified. Subsequent digestion with MvaI restriction endonuclease resulted in two different banding patterns: αLA A1/αLA A1 and αLA A1/αLA A2, and two allelic forms, i.e. αLA A1 and αLA A2. Nucleotide sequencing of the designated LALBA amplicons was done and, following successful BLAST analysis, the sequences were submitted to GenBank (NCBI Accession No. KM267632, KM267633, KM267634 and KP940442). SNPs were searched within breeds of Saudi origin and homology was sought among caprine, ovine and bovine species. A C > T transition was identified within the fifth nucleotide of the silent α-LA A2 allele. Phylogenetic analysis on the basis of LALBA nucleotide sequence of Saudi goats indicated similarity with evolutionarily related sheep, while the origin of bovine species and deer was located some distance away.

Prenatal exposure to BPA alters the epigenome of the rat mammary gland and increases the propensity to neoplastic development.

Exposure to environmental estrogens (xenoestrogens) may play a causal role in the increased breast cancer incidence which has been observed in Europe and the US over the last 50 years. The xenoestrogen bisphenol A (BPA) leaches from plastic food/beverage containers and dental materials. Fetal exposure to BPA induces preneoplastic and neoplastic lesions in the adult rat mammary gland. Previous results suggest that BPA acts through the estrogen receptors which are detected exclusively in the mesenchyme during the exposure period by directly altering gene expression, leading to alterations of the reciprocal interactions between mesenchyme and epithelium. This initiates a long sequence of altered morphogenetic events leading to neoplastic transformation. Additionally, BPA induces epigenetic changes in some tissues. To explore this mechanism in the mammary gland, Wistar-Furth rats were exposed subcutaneously via osmotic pumps to vehicle or 250 µg BPA/kg BW/day, a dose that induced ductal carcinomas in situ. Females exposed from gestational day 9 to postnatal day (PND) 1 were sacrificed at PND4, PND21 and at first estrus after PND50. Genomic DNA (gDNA) was isolated from the mammary tissue and immuno-precipitated using anti-5-methylcytosine antibodies. Detection and quantification of gDNA methylation status using the Nimblegen ChIP array revealed 7412 differentially methylated gDNA segments (out of 58207 segments), with the majority of changes occurring at PND21. Transcriptomal analysis revealed that the majority of gene expression differences between BPA- and vehicle-treated animals were observed later (PND50). BPA exposure resulted in higher levels of pro-activation histone H3K4 trimethylation at the transcriptional initiation site of the alpha-lactalbumin gene at PND4, concomitantly enhancing mRNA expression of this gene. These results show that fetal BPA exposure triggers changes in the postnatal and adult mammary gland epigenome and alters gene expression patterns. These events may contribute to the development of pre-neoplastic and neoplastic lesions that manifest during adulthood.

Mva I PCR-RFLP identifies single nucleotide polymorphism of the alpha-lactalbumin gene in some goat breeds reared in Turkey

Alpha-lactalbumin plays a key role in lactose synthesis in the mammary glands of domestic animals. This study evaluated Hair (188), Saanen (82), Kilis (63), and Honamli (183) goat breeds, with the primary goal of investigating DNA polymorphism of Mva I RFLP at exon 3 of the alpha-lactalbumin gene. Polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP) was employed to genotype a total of 516 goats for Mva I polymorphism of alpha-lactalbumin. In the breeds that were studied, digestion of the amplification product with the Mva I restriction enzyme revealed 2 alleles (A1 and A2) and 3 genotypes (A1A1, A1A2, and A2A2). Allelic frequencies for Hair, Honamli, Saanen, and Kilis breeds were found to be 0.87, 0.92, 0.87, and 0.85, respectively, for the A1 allele and 0.13, 0.08, 0.13, and 0.15, respectively, for the A2 allele. None of the studied breeds exhibited significant deviation from Hardy-Weinberg equilibrium. Consequently, this study shed light on the polymorphism of alpha-lactalbumin in 4 goat breeds. Another finding of this study was the presence of genetic polymorphism at the alpha-lactalbumin gene in Kilis and Honamli goat breeds, a fact that was not known previously.

GWA Analysis for Milk Production Traits in Dairy Sheep and Genetic Support for a QTN Influencing Milk Protein Percentage in the LALBA Gene

In this study, we used the Illumina OvineSNP50 BeadChip to conduct a genome-wide association (GWA) analysis for milk production traits in dairy sheep by analyzing a commercial population of Spanish Churra sheep. The studied population consisted of a total of 1,681 Churra ewes belonging to 16 half-sib families with available records for milk yield (MY), milk protein and fat yields (PY and FY) and milk protein and fat contents (PP and FP). The most significant association identified reached experiment-wise significance for PP and FP and was located on chromosome 3 (OAR3). These results confirm the population-level segregation of a previously reported QTL affecting PP and suggest that this QTL has a significant pleiotropic effect on FP. Further associations were detected at the chromosome-wise significance level on 14 other chromosomal regions. The marker on OAR3 showing the highest significant association was located at the third intron of the alpha-lactalbumin (LALBA) gene, which is a functional and positional candidate underlying this association. Sequencing this gene in the 16 Churra rams of the studied resource population identified additional polymorphisms. One out of the 31 polymorphisms identified was located within the coding gene sequence (LALBA_g.242T>C) and was predicted to cause an amino acid change in the protein (Val27Ala). Different approaches, including GWA analysis, a combined linkage and linkage disequilibrium study and a concordance test with the QTL segregating status of the sires, were utilized to assess the role of this mutation as a putative QTN for the genetic effects detected on OAR3. Our results strongly support the polymorphism LALBA_g.242T>C as the most likely causal mutation of the studied OAR3 QTL affecting PP and FP, although we cannot rule out the possibility that this SNP is in perfect linkage disequilibrium with the true causal polymorphism.

SSCP typing of alpha-lactalbumin and beta-lactoglobulin gene and its association with milk production and constituent traits in Indian riverine buffalo

A study was conducted to explore genetic polymorphism in alpha-lactalbumin and beta-lactoglobulin gene and to elucidate the effect of polymorphism on milk production and constituent traits in Indian riverine buffaloes namely, Murrah, Bhadawari, Mehsana and Surti. Single strand conformation polymorphism (SSCP) was conducted on exon II of alpha-lactalbumin and exon I of beta-lactoglobulin gene. The study on alpha-lactalbumin gene revealed the presence of a number of genotypes namely, AB, AC, BB, BC, BD and BE in Mehsana; AC, BB, BC, BD, BE and CC in Surti; AC, BB, BC, BE and CC in Bhadawari and BC, BE, CC and CD in Murrah buffalo. In beta-lactoglobulin gene, genotypes like AA, BB, CC, AB and AC were observed in Murrah buffalo; AA, BB, AB and AC in Mehsana; AA, BB, CC, AB and AC in Surti and BB, CC, AB and AC in Bhadawari buffalo. All the alleles were sequenced and found to be differed at different locations. The alpha-lactalbumin genotypes had significant effect (P&lt;0.05) on fat%, daily fat yield, total fat yield, SNF%, daily solid not fat and total solid not fat yield in Murrah buffalo; whereas, in Surti breed total milk yield, daily solid yield, total solid, daily solid not fat, total solid not fat yield, daily and total protein yield were significantly associated with the genotypes. The beta-lactoglobulin genotypes showed significant effect (P&lt;0.05) on daily milk yield in Murrah, Bhadawari and Mehsana buffalo. In all the breeds, BB genotypes produced highest daily milk yield. In Murrah, Bhadawari and Mehsana breed, genotypes AA, CC and AC were associated with lowest production.

Single nucleotide polymorphism (SNP) in alpha-lactalbumin gene of Indian Jamunapari breed of Capra hircus

Alpha-lactalbumin is a major whey protein found in milk. It influences lactose synthesis by modifying the substrate specificity of galactosyl-transferase, is important to milk synthesis since lactose, an impermeable disaccharide, is the major osmole of milk. The present study was undertaken to detect polymorphism in the full coding region of alpha-lactalbumin at the genetic level and to explore allelic variability of this gene. Samples of Jamunapari breed of goat ( n = 50) were included under the present study. Jamunapari is the highest milk producer among local Indian goat breeds. PCR-SSCP of all four exons of alpha-lactalbumin (ALA) revealed a total of 9 gel phenotypes of Jamunapari breed of goat. These were sequenced, analyzed and deposited in GenBank, NCBI (accession nos. EU573193– EU573195). Nucleotide and amino acid variations were searched within breeds of Indian goats and homology between caprine, ovine, bovine, bubaline and human. In the present study we describe for the first time two novel gene variants on the goat alpha-lactalbumin gene exon 4.

Protein profile and alpha-lactalbumin concentration in the milk of standard and transgenic goats expressing recombinant human butyrylcholinesterase

The expression of recombinant proteins of pharmaceutical interest in the milk of transgenic farm animals can result in phenotypes exhibiting compromised lactation performance, as a result of the extraordinary demand placed on the mammary gland. In this study, we investigated differences in the protein composition of milk from control and transgenic goats expressing recombinant human butyrylcholinesterase. In Experiment 1, the milk was characterized by gel electrophoresis and liquid chromatography/mass spectrometry in order to identify protein bands that were uniquely visible in the transgenic milk and/or at differing band densities compared with controls. Differences in protein content were additionally evaluated by computer assisted band densitometry. Proteins identified in the transgenic milk only included serum proteins (i.e. complement component 3b, ceruloplasmin), a cytoskeleton protein (i.e. actin) and a stress-induced protein (94 kDA glucose-regulated protein). Proteins exhibiting evident differences in band density between the transgenic and control groups included immunoglobulins, serum albumin, beta-lactoglobulin and alpha-lactalbumin. These results were found to be indicative of compromised epithelial tight junctions, premature mammary cell death, and protein synthesis stress resulting from transgene expression. In Experiment 2, the concentration of alpha-lactalbumin was determined using the IDRing assay and was found to be significantly reduced on day 1 of lactation in transgenic goats (4.33 +/- 0.97 vs. 2.24 +/- 0.25 mg/ml, P < 0.01), but was not different from non-transgenic controls by day 30 (0.99 +/- 0.46 vs. 0.90 +/- 0.11 mg/ml, P > 0.05). We concluded that a decreased/delayed expression of the alpha-lactalbumin gene may be the cause for the delayed start of milk production observed in this herd of transgenic goats.

Transformation of Human Alpha-lactalbumin Gene into Tobacco and Improvement of Cysteine Contents in Transgenic Tobacco Plants

The plant expression vector containing human alpha-lactalbumin gene was constructed and its gene was transformed into tobacco using leaf disc mediated by Agrobacterium tumefaciens LBA 4404 as a vehicle.PCR and Southern blot analyses showed that human alpha-lactalbumin gene was integrated into the tobacco genome.RT-PCR and histochemical staining for GUS activity demonstrated that human alpha-lactalbumin gene was expressed in transgenic tobacco plants.Further analysis of the data indicated that the average of cysteine contents in 9 transgenic tobacco plants was greatly increased by 166.40% besides one of them which was increased by 318.02%.

Single strand conformation polymorphism detection in alpha-lactalbumin gene of Indian Jakhrana milk goats

Alpha-lactalbumin (α-LA) (ALA), one of the two major whey proteins, is strongly correlated with the nutritional value and the functional properties of whey and whey products. The genetic variations of ALA gene in Jakhrana Goat breed (Capra hircus) were investigated using an optimized non-radioactive polymerase chain reaction–single strand conformation polymorphism (PCR–SSCP) analysis of four amplified fragments covering all four exons of the gene. A total of eight SSCPs patterns were detected in three of these exons in a sample of 50 Jakhrana goats, indicating that this breed has high genetic variability in the ALA gene. This result opens interesting prospects for future breeding programs and conservation strategies. These ALA gene variants can be sequenced and screened in the entire population to develop single nucleotide polymorphisms for association studies with different productive and reproductive performances and marker assisted selection. In addition, our data show that PCR–SSCP is an appropriate tool for evaluating genetic variability.

Correlations between SNP of &lt;I&gt;LALBA&lt;/I&gt; gene and economic traits in Inner Mongolian white cashmere goat

PCR-SSCP and DNA sequencing methods were conducted to detect single nucleotide polymorphism of alpha-lactalbumin (LALBA) gene in 452 Inner Mongolian white cashmere goats (IMWC). Correlations between SNP of goat LALBA gene and economic traits, e.g., cashmere yield, cashmere thickness, length and weight, were analyzed. The SSCP in P2 primer locus, which was caused by the point mutation M63868:g.1897T>C in the exon 3 of LALBA gene was detected. At this locus, the genotype TT and allele T were predominant in the IMWC population, which agreed with Hardy-Weinberg equilibrium. Moreover, there was a significant correlation between polymorphism of goat M63868:g.1897 locus and cashmere yield of IMWC (P=0.017). The individuals with genotype TC had more cashmere yield than those with geontype TT. Hence, genotype TC of LALBA gene can be used as a molecular marker for breeding superior cashmere yield in goat marker-assisted selection.

Uteroplacental insufficiency alters the mammary gland response to lactogenic hormones in vitro

Adequate mammary development and coordinated actions of lactogenic hormones are essential for the initiation of lactation. Pregnancies compromised by uteroplacental insufficiency impair mammary development and lactation, further slowing postnatal growth. It is not known whether the initiation of lactation or galactopoesis is compromised. Uteroplacental insufficiency induced in rats by bilateral uterine vessel ligation (Restricted) or sham surgery (Control) on Day 18 of gestation preceded collection of mammary tissue on Day 20 of pregnancy. Mammary explants were cultured with combinations of insulin, cortisol and prolactin and analysed for alpha-lactalbumin and beta-casein gene expression. Mammary tissue from late pregnant Restricted rats had elevated alpha-lactalbumin, but not beta-casein, mRNA, which is consistent with premature lactogenesis resulting from an early decline in peripheral maternal progesterone. Explants from Restricted rats were more responsive to hormone stimulation after 3 days in culture, indicating that compromised galactopoesis, not lactogenesis, most likely leads to the reduced growth of suckled pups.