Novel SNP in 5' flanking region of EDG1 associated with marbling in Japanese Black beef cattle

Marbling, defined by the amount and distribution of intramuscular fat, is an economically important trait of beef cattle in Japan. The endothelial differentiation , sphingolipid G-protein-coupled receptor , 1 ( EDG1 ) gene has been considered as a positional functional candidate for the gene responsible for marbling. We have recently reported that 2 single nucleotide polymorphisms (SNPs), c.-312A > G in the 5' untranslated region (UTR) and c.*446G > A in the 3' UTR in EDG1 were associated with marbling in Japanese Black beef cattle, but this was not functional and a causal mutation for marbling. In the present study, we detected 2 novel SNPs, referred to as g.1475435G > A and g.1471620G > T , in the 5' flanking region of the EDG1 between low-marbled and high-marbled steer groups, which were previously shown to have EDG1 expression differences in musculus longissimus muscle. The g.1475435G > A SNP seemed not to segregate in Japanese Black beef cattle. The g.1471620G > T SNP was associated with the predicted breeding value for beef marbling standard number by the analyses using Japanese Black beef cattle population. Based on these findings, we hypothesized that the g.1471620G > T SNP might have an impact on EDG1 expression and also marbling.     

Association of a single nucleotide polymorphism in ribosomal protein L27a gene with marbling in Japanese Black beef cattle

Marbling, defined by the amount and distribution of intramuscular fat, is an economically important trait of beef cattle in Japan. The c2‐11#2 expressed sequence tag (EST) has been previously shown to possess expression difference in musculus longissimus muscle between low‐marbled and high‐marbled steer groups, and to be located within genomic region of a quantitative trait locus for marbling. Thus, the ribosomal protein L27a (RPL27A) gene containing the c2‐11#2 EST sequence was considered as a positional candidate for the gene responsible for marbling. In the present study, a single nucleotide polymorphism (SNP) in the promoter region of the RPL27A, referred to as g.3109537C>T, was detected between the 2 steer groups. The SNP was associated with the predicted breeding value for beef marbling standard number by the analyses using Japanese Black beef cattle population. The effect of genotypes of the SNP on the predicted breeding value for subcutaneous fat thickness was not statistically significant. These findings suggest that the RPL27A SNP may be useful for effective marker‐assisted selection to increase the levels of marbling in Japanese Black beef cattle.     

The g.841G>C SNP of FASN gene is associated with fatty acid composition in beef cattle

The objective of the current study is to evaluate the association between fatty acid composition and fatty acid synthase gene polymorphisms as responsible mutations. For this purpose, we selected seven previously reported single nucleotide polymorphisms (SNPs) in FASN gene, including one within promoter region (g.841G>C) and six non‐synonymous SNPs (g.8805C>T, g.13126C>T, g.15532A>C, g.16024A>G, g.16039C>T, g.17924A>G), and genotyped them in Japanese Black cattle. Genotyping results revealed that g.8805 C>T and g.17924 A>G were monomorphic loci. Genome‐wide association analysis including the other five SNPs revealed that only g.841G>C showed significant associations with the percentages of C14:0, C14:1, C16:1 and C18:1 at 5% genome‐wide significance level. In order to further evaluate the effect, we genotyped g.841G>C using additional three populations, including two Japanese Black populations and a Holstein cattle population. g.16024A>G was also genotyped and included in the analysis because it has been reported to be associated with fatty acid composition in Japanese Black cattle. In the result of analysis of variance, g.841G>C showed stronger effects on fatty acid percentage than those of g.16024A>G in all populations. These results suggested that g.841G>C would be a responsible mutation for fatty acid composition and contribute to production of high‐grade beef as a selection marker in beef cattle.     

Association of PCSK1 gene polymorphisms with abdominal fat content in broilers

Protein proteolytic enzymes (Proprotein Convertase, PC) is a Ca²⁺‐dependent serine protease family, whose main function is to cleave precursors of biologically inactive proteins or peptide chains into active functional molecules. Proprotein convertase subtilisin/kexin type 1 (PCSK1) gene is mainly expressed in nerve and endocrine tissues. In this study, PCSK1 was selected as an important candidate gene for abdominal fat content in broilers. We cloned the exon region of chicken PCSK1 gene and found six single‐nucleotide polymorphisms (SNPs). Association analysis was carried out and we found that the polymorphisms of these six SNPs were significantly associated with abdominal fat content in G19 and G20 populations. Five of these SNPs were significantly associated with abdominal fat content in G19 and G20 combined population. The polymorphism of these five SNPs was significantly correlated with the abdominal fat content of AA broilers. Together, our study demonstrated that c.927T>C, c.1880C>T, c.*900G>A, and c.*1164C>T were significantly associated with abdominal fat content in populations used in this study, which means that these SNPs in PCSK1 gene could be used as candidate markers to select lean broiler lines.     

The T allele at the g.1471620G>T in the EDG1 gene associated with high marbling in Japanese Black cattle is at a low frequency in breeds not selected for marbling

Our previous study detected a single nucleotide polymorphism (SNP), g.1471620G > T , in the 5' flanking region of the endothelial differentiation sphingolipid G-protein-coupled receptor 1 ( EDG1 ) gene, which has been considered as a positional functional candidate for the gene responsible for marbling, and showed association of the g.1471620G > T SNP with marbling in Japanese Black beef cattle. In the present study, we investigated the allele frequency distribution of the g.1471620G > T SNP among the 5 cattle breeds, Japanese Black, Japanese Brown, Japanese Short Horn, Holstein, and Brown Swiss breeds. The T allele at the g.1471620G > T SNP associated with high marbling was found at high frequency in Japanese Black breed that has been subjected to a strong selection for high marbling, while the allele was absent or at very low frequencies in the other breeds that have not been strongly selected for high marbling. Based on this finding, we hypothesized that the pressure of the strong selection for high marbling in Japanese Black breed has increased the frequency of the T allele at the g.1471620G > T SNP in the EDG1 .     

UTS2R gene polymorphisms are associated with fatty acid composition in Japanese beef cattle

Fatty acid composition of beef adipose tissue is one of its important traits because a high proportion of monounsaturated fatty acid is related to favorable beef flavor and tenderness. In this study, we searched polymorphisms in full length coding DNA sequence of urotensin 2 recepter and investigated the effects on fatty acid composition (C14:0, C14:1, C16:0, C16:1, C18:0, C18:1, C18:2, monounsaturated fatty acid, saturated fatty acid). Eight single nucleotide polymorphisms (SNP) were identified by sequence comparison among eight animals, including five Japanese Black and three Holstein cattle. One of these SNP (c.866C>T) was predicted to cause amino acid substitutions (P289L) and the other seven synonymous SNP, including c.267C>T, were presumed to be in linkage disequilibrium. Therefore we selected two SNP (c.267C>T and c.866C>T) for further analysis. We investigated associations between these genotypes and fatty acid composition in three Japanese Black populations (n = 560, 245 and 287) and a Holstein population (n = 202). Tukey‐Kramer's honestly significant difference test revealed that CC genotype in c.267C>T indicated lower C14:0 and higher C18:1 than the other genotypes in Japanese Black cattle and CC genotype in c.866C>T showed lower C16:1 than CT genotype in Holstein cattle (P < 0.05). These results suggested that these genotypes would contribute to production of high‐grade meat as selection markers in beef cattle.     

Estimation of beef marbling in the Longissimus muscle with computer image analysis of ultrasonic pictures of the Iliocostalis muscle area

This study investigated an objective method for estimating beef marbling using ultrasonic images of the Iliocostalis muscle and the Lomgissimus muscle area sections. Thirty‐one Japanese Black cattle steers were used in this study. The end of the left side shoulder blade bone was scanned using an ultrasonic device. Ultrasonic images were captured of the Longissimus muscle area and that around the Iliocostalis muscle area. Twenty items were measured in the two images using computer image analysis software. The level of beef marbling was measured according to the Beef Marbling Standard (BMS) for carcass grading, and the percentage of ether‐extractable fat content in the Longissimus muscle (EE). The difference in the gray level between the Iliocostalis muscle and intermuscular fat (X10) was used to estimate the BMS and the EE, which were highly correlated (r² = 67.72% and 61.30%). An equation was developed using four parameters from the two ultrasonic images, which could estimate the BMS (r² = 85.88%). This equation could also estimate the EE (r² = 68.98%). The equations used to estimate beef marbling were based on one to four parameters that included X10. Thus, ultrasonic images of the Iliocostalis muscle area section are important for estimating beef marbling.     

Polymorphisms of the melatonin receptor 1A gene that affects the reproductive seasonality and litter size in Small Tail Han sheep

Previous researches have shown that MTNR1A plays an essential role in sheep reproduction. However, most researches focused more on the reproductive seasonality of sheep, and few scientists had studied the association of polymorphisms of the MTNR1A gene with ovine litter size and reproductive seasonality. Therefore, we chose MTNR1A gene to detect its novel sequence polymorphisms and population genetics and analyse their association with seasonal reproduction and litter size in ewes. The mRNA expression level in hypothalamus, pituitary and ovary was also detected. In this study, five polymorphisms (g.15118664G > T, g.15118683C > T, g.15118756C > T, g.15118774C > T and g.15118951G > A) were identified in exon 2. Most importantly, the g.15118683C > T and g.15118951G > A were significant difference between year‐round oestrous sheep and seasonal oestrous sheep (p < .01), and g.15118756C > T had a great effect on litter size of Small Tail Han sheep (p < .05). In addition, the mRNA expression level of MTNR1A in the hypothalamus of polytocous Small Tail Han sheep was significantly higher than that in monotocous Small Tail Han sheep (p < .01) and the expression of MTNR1A in the hypothalamus of year‐round oestrous sheep was significantly higher than that in seasonal oestrous sheep (p < .01). Polymorphisms in exon 2 may regulate the reproductive seasonality and litter size of ewes by influencing gene expression to regulate the reproductive seasonality and litter size of ewes. Our studies provided useful guidance in marker‐assisted selection of the litter size in Small Tail Han sheep.     

Genetic association between GHSR1a 5′UTR‐microsatellite and nt‐7(C>A) loci and growth and carcass traits in Japanese Black cattle

We carried out a genetic association study between five nucleotide polymorphisms (5′UTR microsatellite ((TG)_n), nt‐7(C>A), L24V, DelR242 and Intron 1 microsatellite) of the GHSR1a gene and growth and carcass traits in 1285 steers sired by 117 Japanese Black bulls in a progeny testing program. We report herein, a significant association between the 5′UTR microsatellite and nt‐7(C>A) loci and growth and carcass traits. We also propose a translational hypothesis that the association is due to differences in the secondary structure of GHSR1b mRNA (the non‐spliced type with the 5′UTR microsatellite) among the GHSR1a gene haplotypes. Furthermore, we predicted the potential increase in profitability due to increased carcass weight in cow‐calf fattening enterprises through planned matings based on DNA testing of the 5′UTR microsatellite. Statistical analysis revealed that the 5′UTR microsatellite locus had a significant additive effect on carcass weight (CW) and average daily gain (ADG), but not on beef marbling score (BMS). One of the four major microsatellite alleles (19‐TG allele) with an allele frequency of 0.145, had a significantly (P < 0.0007) desirable effect on CW and ADG. We concluded that the 19‐TG allele could potentially be economically useful nucleotide markers for growth and carcass traits in Japanese Black cattle.     

Associations between candidate SNPs in the calpain 1, calpastatin and leptin genes and meat tenderness among Swedish beef populations

Abstract

The contribution of three candidate genes to the variation in meat tenderness was tested in muscle samples from 243 pure-bred, young, beef bulls of Angus, Charolais, Hereford, Limousin and Simmental breeds, raised in Swedish commercial herds. The animals were genotyped for single nucleotide polymorphisms (SNPs) in the calpain 1 (CAPN1), calpastatin (CAST) and leptin genes. The frequent calpain 1 CAPN1:c.947G>C G allele showed an unfavourable association with tenderness. The calpastatin CAST:c.155C>T T allele, which was the most common allele, showed a favourable association with Warner–Bratzler shear force (WBSF) and compression tests. An association was observed between the leptin UASMS2C>T SNP and compression tests.     

Genetic parameter estimation and gene association analyses for meat quality traits in open-air free-range Iberian pigs

Meat quality of Iberian pigs is defined by the combination of their genetic characteristics and the particular production system. To carry out a genetic analysis of the main meat quality traits, we estimated their heritabilities, genetic correlations and the association effects of 32 selected SNPs of 12 candidate genes. A total of ten traits were measured in longissimus dorsi samples from 1,199 Iberian pigs fattened in the traditional free-range system: water holding capacity (thawing, cooking and centrifuge force water losses), instrumental colour (lightness L*, redness a* and yellowness b*), myoglobin content, shear force on cooked meat, and shear force and maximum compression force on dry-cured loin. Estimated heritability values were low to moderate (0.01 to 0.43) being the lowest for L* and the highest for cooking loss. Strong genetic correlations between water holding capacity traits (0.93 to 0.96) and between myoglobin content and a* (0.94) were observed. The association analyses revealed 19 SNPs significantly associated with different traits. Consistent and strong effects were observed between PRKAG3 SNPs (rs319678464G > C and rs330427832C > T), MYH3_rs81437544T > C, CASP3_rs319658214G > T and CTSL_rs332171512A > G and water losses. Also for CAPN1_rs81358667G > A and CASP3_rs319658214G > T and shear force. The SNPs mapping on PRKAG3 showed the highest effects on Minolta colour traits. Genotyping of these SNPs could be useful for the selection of Iberian young boars with similar estimated breeding values for productive traits.     

Genome‐wide association studies identified variants for taurine concentration in Japanese Black beef

We performed genome‐wide association studies (GWAS) using the BovineSNP50 array to detect significant single nucleotide polymorphisms (SNPs) that may affect the concentration of 22 free amino acids and three peptides in Japanese Black beef cattle. A total of 574 Japanese Black cattle and 40,657 SNPs from the array were used for this study. Genome‐wide significant SNPs were detected for β‐alanine (three SNPs on chromosomes 22 and 29) and taurine (26 SNPs on chromosome 22). Importantly, the top two SNPs for taurine were highly significant (p = 6.2 × 10^?21), and the frequency of the increase‐concentration allele (Q) for taurine was found to be 0.73. The Q allele frequency of this population was similar to that of the other unrelated Japanese Black cattle, but different from that of the other breeds. In addition, the significant SNPs were not associated with carcass traits or fatty acid compositions. Interestingly, the top three of the four most significant SNPs for taurine were located near solute carrier family 6, member 6 (SLC6A6), which is a membrane transporter for taurine. We also found two associated variants in the 5′‐upstream region of SLC6A6; however, they were less significantly associated than the SNPs from the BovineSNP50 array.     

Effects of intramuscular fat deposition on the beef traits of Japanese Black steers (Wagyu)

Effects of intramuscular fat deposition on the chemical composition, tenderness, and free amino acids (FAA) concentration of beef were studied using various classified carcasses of 21 Japanese Black (Wagyu) steers. The Beef Marbling Standard (BMS) No., judged on the market in 1998, and fat content, ranged from 2 to 10 and 4.8 to 39.0% in the M. longissimus thoracis, respectively. Moisture content was negatively correlated with the fat content (r =?0.98, P < 0.01). In contrast, protein content was relatively constant up to approximately 23% fat, corresponding to BMS No.4, and decreased as the fat content increased. Cooking loss was also constant up to approximately 28% fat, corresponding to BMS No.4, and markedly decreased with fat content increase. The beef in these cases were, however, classified into BMS No. around 9 and 11, respectively, according to the BMS model in 1988. A negative correlation was found between the shear‐force value and the fat content (r =?0.83, P < 0.05). Most FAA concentrations on the wet weight of meat were correlated negatively with the fat content, except glutamine, and this negative relationship was still observed when the concentrations were recalculated on the basis of protein. These results indicate that higher marbling Wagyu beef (above approx. 23% fat) would have an extremely lower content of protein, which would partly explain the lower cooking loss and FAA with fat increase. In addition, some other reasons not relating to protein content (e.g. high fat content preventing the breakdown of protein to FAA) seem to explain the negative correlation between fat and FAA.     

Development of discrimination markers between Japanese domestic and imported beef

In the meat industry, correct labeling of beef origins or breed is required to assure quality and safety. This paper describes the development of discrimination markers between Japanese domestic and imported beef from the United States (US) and Australia (AUS) based on a bovine 50K single nucleotide polymorphism (SNP) array using a total of 110 samples: Japanese Black (n = 50), Japanese Holstein (n = 50) and US cattle (n = 10). Genotyping information revealed 1081 SNPs as candidate markers that were polymorphic only in US cattle. The genotyping results by PCR – restriction length polymorphism in Japanese Black (n = 300) and Holstein cattle (n = 146) revealed that 11 SNPs had alleles specific to US cattle. Their allelic frequencies in US cattle (n = 108) ranged from 0.097 to 0.250 with an average of 0.178 and the combined identification probability of US cattle was 0.987. In addition, we also verified the applicability of these US‐specific markers to AUS cattle. Their allelic frequencies in AUS cattle (n = 280) ranged from 0.063 to 0.224 with an average of 0.137 and the combined identification probability of AUS cattle was 0.963. In conclusion, a set of these markers could be useful for discriminating between Japanese domestic and imported beef and would contribute to identify origins and prevent falsified labeling of beef.     

Lack of the Associations of the Polymorphisms in IGF2, MC4R and GNAS Genes with Reproduction Traits in Pigs and Imprinting Analysis of IGF2 Gene in Ovary and Cornus Uteri

In recent years, intensive attention has been put on improving reproductive performance of pigs. Several experiments aimed to identify markers associated with prolificacy, but this issue still remains open. In our study, we investigated associations between polymorphisms in IGF2, GNAS and MC4R genes with reproductive traits of Polish Landrace and Large White pigs. We did not find any significant associations for g. GNAS314T > 324C, IGF2 intron3‐g.3072G > A or g. MC4R 1426G > A in Polish Landrace and Large White pigs. In the case of IGF2 intron3‐g.3072G > A, this information is of great importance, because this marker is widely implemented in pigs breeding and previous experiments suggested its role in prolificacy of pigs. We also investigated expression of IGF2 gene and showed that this gene is monoallelically expressed in reproductive organs (ovary and cornus uteri).     

Two novel intronic polymorphisms of bovine FGF21 gene are associated with body weight at 18 months in Chinese cattle

Fibroblast growth factor 21 (FGF21) is a hepatic hormone that regulates peripheral glucose tolerance, energy balance and lipid metabolism. Prior evidence suggests that FGF21 may have the potential to favorably reduce obesity. The objective of the present study was to identify single nucleotide polymorphisms (SNPs) of bovine FGF21 using 1255 animals representing the five main Chinese breeds and to investigate the effect of these SNPs on economic traits in Nanyang cattle. Four significant SNPs were identified, one was a synonymous mutation and the other three were in intronic regions. The polymorphism information content (PIC) analysis showed that four beef cattle populations (NY, JX, LX and QC) had a moderate genetic diversity at the four loci while the beef and dairy population (CRS) had a low level. Additionally, allele and genotype frequencies for the beef breeds were significantly different from CRS, implying that these mutations are possibly associated with some quantitative traits. Moreover, linkage disequilibrium analysis and haplotype frequencies were also reported. Seven different haplotypes were identified and haplotype TCCC was predominant in all five cattle breeds. Association analysis suggested that SNPs g.297C>G and g.940C>T of bovine FGF21 were associated with higher body weight at 18 months within NY cattle, which would contribute to cattle breeding and genetics through marker-assisted selection (MAS).     

Development of SNP markers for individual identification and parentage test in a Japanese Black cattle population

This study describes the development of efficient single nucleotide polymorphism (SNP) markers for individual identification and parentage tests in a Japanese Black cattle population. An amplified fragment length polymorphism method was employed to detect informative candidate markers, and yielded 44 SNP markers from 220 primer combinations. 29 unlinked SNPs were finally selected as diagnostic markers. The allelic frequencies for each marker were estimated by using PCR‐RFLP in the Japanese Black population. Based on the frequency data, the estimated identity power of these markers was 2.73 × 10^?12. Parentage exclusion probabilities, when both suspected parents' genotypes were known and when only one suspected parent was genotyped, were estimated as 0.96929 and 0.99693, respectively. This panel of SNP markers is theoretically sufficient for individual identification, and would also be a powerful tool for a parentage test in Japanese Black cattle. The markers could contribute to the management of the beef industry in Japan.     

Application of highly differentiated SNPs between Japanese Black and Holstein to a breed assignment test between Japanese Black and F1 (Japanese Black x Holstein) and Holstein

Two taurine breeds, Japanese Black and Holstein, established from geographically distant origins and selected for different uses, beef and dairy, were extensively genotyped using a genome‐wide single nucleotide polymorphism (SNP) chip with more than 1000 animals of each breed. The genetic structure was examined by principal component analysis, in which the first principal component clearly separated the two breeds and explained more than 15% of the variance. Highly differentiated SNPs were detected throughout the genome, some of which were clustered within small regions on BTA4 (79.2–79.7 Mb, Btau4.0) and BTA26 (22.2–23.6 Mb). A breed assignment test was developed using 18 highly differentiated SNPs to distinguish Japanese Black from F₁ (Japanese Black × Holstein) and Holstein. The error rate that an F₁ or Holstein animal is misjudged as Japanese Black was expected to be < 0.8%, while the error rate that a Japanese Black animal is misjudged as F₁ or Holstein was expected to be < 0.001%. This test provides a reliable and powerful method to detect breed label falsification in retail beef.     

Relationship between perilipin genes polymorphisms and growth,carcass and meat quality traits in pigs

The perilipins (PLIN) belong to a family of structural proteins that play a role regulating intracellular lipid storage and mobilization. Here, PLIN1 and PLIN2 have been evaluated as candidate genes for growth, carcass and meat quality traits in pigs. A sample of 607 Duroc pigs were genotyped for two single‐nucleotide polymorphisms, one in intron 2 of the PLIN1 gene (JN860199:g.173G>A) and the other at the 3′ untranslated region of the PLIN2 gene (GU461317:g.98G>A). Using a Bayesian approach, we have been able to find evidence of additive, dominant and epistatic associations of the PLIN1 and PLIN2 polymorphisms with early growth rate and carcass length. However, the major effects were produced by the dominant A allele at the PLIN2 polymorphism, which also affected the carcass lean weight. Thus, pigs carrying an additional copy of the A allele at the g.98G>A PLIN2 polymorphism had a probability of at least 98% of producing carcasses with heavier lean weight (+0.41 kg) and ham weight (+0.10 kg). The results obtained indicate that the PLIN2 polymorphism could be a useful marker for lean growth. In particular, it may help to reduce the undesired negative correlated response in lean weight to selection for increased intramuscular fat content, a common scenario in some Duroc lines involved in the production of high quality pork products.