PLAG1 and NCAPG‐LCORL in livestock

A recent progress on stature genetics has revealed simple genetic architecture in livestock animals in contrast to that in humans. PLAG1 and/or NCAPG‐LCORL, both of which are known as a locus for adult human height, have been detected for association with body weight/height in cattle and horses, and for selective sweep in dogs and pigs. The findings indicate a significant impact of these loci on mammalian growth or body size and usefulness of the natural variants for selective breeding. However, association with an unfavorable trait, such as late puberty or risk for a neuropathic disease, was also reported for the respective loci, indicating an importance to discriminate between causality and association. Here I review the recent findings on quantitative trait loci (QTL) for stature in livestock animals, mainly focusing on the PLAG1 and NCAPG‐LCORL loci. I also describe our recent efforts to identify the causative variation for the third major locus for carcass weight in Japanese Black cattle.     

Genetic diversity and genomic inbreeding in Japanese Black cows in the islands of Okinawa Prefecture evaluated using single-nucleotide polymorphism array

Maintaining genetic diversity and inbreeding control are important in Japanese Black cattle production, especially in remote areas such as the islands of Okinawa Prefecture. Using a single-nucleotide polymorphism (SNP) array, we evaluated the genetic diversity and genomic inbreeding in Japanese Black cows from the islands of Okinawa Prefecture and compared them to those from other locations across Japan. Linkage disequilibrium decay was slower in cows in the islands of Okinawa Prefecture. The estimated effective population size declined over time in both populations. The genomic inbreeding coefficient (F_ROH) was estimated using long stretches of consecutive homozygous SNPs (runs of homozygosity; ROH). F_ROH was higher in the cows on the islands of Okinawa Prefecture than on other locations. In total, 818 ROH fragments, including those containing NCAPG and PLAG1, which are major quantitative trait loci for carcass weight in Japanese Black cattle, were present at significantly higher frequencies in cows in the islands of Okinawa Prefecture. This suggests that the ROH fragments are under strong selection and that cows in the islands of Okinawa Prefecture have low genetic diversity and high genomic inbreeding relative to those at other locations. SNP arrays are useful tools for evaluating genetic diversity and genomic inbreeding in cattle.     

Variation in the chicken LPIN2 gene and association with performance traits

1. The objective of the study was to investigate the distribution of LPIN2 variants and haplotypes among breeds and perform an association analysis of the variants and haplotypes with the broiler traits in chickens.

2. Six breeds were used to study the variation and distribution of chicken LPIN2, and an F₂ resource population was used to measure growth traits, carcass traits, meat quality traits and serum biochemistry parameters.

3. A c.-599G>A variant was located in the promoter region of LPIN2 and c.444G>A and c.1730A>T (E577D) coding variant mutations were detected. Linkage disequilibrium tests showed that these three variants were under moderate linkage disequilibrium in the 6 breeds and 7 haplotypes were constructed. The distribution of variation/haplotypes presented clear differences among breeds.

4. Association analysis showed that c.-599G>A was associated with leg muscle weight, jejunum length, ileum length, leg muscle fibre density and leg muscle fibre diameter; c.444G>A was associated with spleen weight, ileum length, body weight at hatch and metatarsus length at 8 weeks; c.1730T>A had significant effects on chicken liver weight, heart weight, body weight at 10 weeks, serum albumin and glucose.

5. Diplotypes were significantly associated with body weight at hatch, heart weight, pancreas weight, duodenum length, leg muscle fibre density and lactate dehydrogenase.

     

Whole-genome QTL scan for ultrasound and carcass merit traits in beef cattle using Bayesian shrinkage method

Fine mapping of quantitative trait loci (QTL) for 16 ultrasound measurements and carcass merit traits that were collected from 418 hybrid steers was conducted using 1207 SNP markers covering the entire genome. These SNP markers were evaluated using a Bayesian shrinkage estimation method and the empirical critical significant thresholds (α = 0.05 and α = 0.01) were determined by permutation based on 3500 permuted datasets for each trait to control the genome-wide type I error rates. The analyses identified a total of 105 QTLs (p < 0.05) for seven ultrasound measure traits including ultrasound backfat, ultrasound marbling and ultrasound ribeye area and 113 QTLs for seven carcass merit traits of carcass weight, grade fat, average backfat, ribeye area, lean meat yield, marbling and yield grade. Proportion of phenotypic variance accounted for by a single QTL ranged from 0.06% for mean ultrasound backfat to 4.83% for carcass marbling (CMAR) score, while proportion of the phenotypic variance accounted for by all significant (p < 0.05) QTL identified for a single trait ranged from 4.54% for carcass weight to 23.87% for CMAR.     

Genetic and genomic analyses for predicted methane‐related traits in Japanese Black steers

The objectives of this study were to estimate genetic parameters and to perform a genome‐wide association study (GWAS) for predicted methane‐related traits in Japanese Black steers. The methane production and yield traits were predicted using on‐farm measurable traits, such as dry matter intake and average daily gain. A total of 4,578 Japanese Black steers, which were progenies of 362 sires genotyped with imputed 551,995 single nucleotide polymorphisms (SNPs), had phenotypes of predicted methane‐related traits during the total fattening period (52 weeks). For the estimation of genetic parameters, the estimated heritabilities were moderate (ranged from 0.57 to 0.60). In addition, the estimated genetic correlations of methane production traits with most of carcass traits and feed‐efficiency traits were unfavorable, but those of methane yield traits were favorable or low. For the GWAS, no genome‐wide significant SNP was detected, but a total of four quantitative trait locus (QTL) regions that explained more than 5.0% of genetic variance were localized on the genome, and some candidate genes associated with growth and feed‐efficiency traits were located on the regions. Our results suggest that the predicted methane‐related traits are heritable and some QTL regions for the traits are localized on the genome in Japanese Black steers.     

Effects of clay on fat necrosis and carcass characteristics in Japanese Black steers

Twenty 10‐month‐old Japanese Black steers were used to evaluate the effects of clay on fat necrosis and carcass characteristics. Ten steers (Clay group) were fed the clay (50 g/day) during 10–30 months of age. The other 10 steers (Control group) were not fed it. There was no significant difference in body weight or average daily gain between the two groups (P > 0.05). The occurrence of fat necrotic mass in the Clay group (30%) was lower (P < 0.05) than that in the Control group (90%) at slaughter. The size of necrotic masses in the Clay group was smaller (P < 0.05) than that in the Control group. There was no significant difference in the marbling score, beef color, Longissimus muscle area or subcutaneous fat thickness between the two groups. These results suggest that the clay prevented the occurrence of fat necrosis and did not affect the carcass characteristics in Japanese Black steers.     

Polymorphisms of the PNPLA3 gene and their associations with chicken growth and carcass traits

1. An F₂ resource population of Gushi chickens crossed with Anka broilers was used to investigate the genetic effects of the chicken PNPLA3 gene on growth and adipose accumulation.

2. Associations between three SNPs (g.40006G?>?T, g.42344T?>?C and g.42404A?>?T) and broiler traits were determined using linkage disequilibrium, haplotype construction and association analysis.

3. The g.40006G?>?T mutation was associated with body weights at 4, 6, 8, 10 and 12 weeks of age, carcass weight, evisceration weight and semi-evisceration weight (P? 4. Haplotypes of the g.42344T?>?C and g.42404A?>?T mutations were associated with body weight at 12 weeks, carcass weight, evisceration weight, and semi-evisceration weight (P? 5. The results suggest that the PNPLA3 gene may be in linkage with the causative mutation or a QTL controlling growth traits in chickens. In contrast to human studies, the polymorphisms were not associated with fat related traits.     

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.     

Associations between variants of bone morphogenetic protein 7 gene and growth traits in chickens

1. Enhancing bone strength to solve leg disorders in poultry has become an important goal in broiler production. Bone morphogenetic protein 7 (BMP7), a member of the BMP family, represents an attractive therapeutic target for bone regeneration in humans and plays critical roles in skeletal development.

2. The objective of this study was to investigate the relationship between BMP7 gene expression, single-nucleotide polymorphisms (SNPs) and growth traits in chickens. Here, a SNP (c.1995T>C) in the chicken (Gallus gallus) BMP7 gene was identified, that was associated with growth and carcass traits.

3. Genotyping revealed that the T allele occurred more frequently in breeds with high growth rates, whereas the C allele was predominant in those with low growth rates. The expression level of BMP7 in the thigh bone of birds with the TT genotype was significantly higher than in those with the CC genotype at 21, 42 and 91 d of age.

4. These findings suggest that selecting the birds with the TT genotype of SNP c.1995T>C could improve bone growth, could reduce leg disorders in fast-growing birds. The SNP c.1995T>C may serve as a selective marker for improving bone growth and increasing the consistency of body weights in poultry breeding.     

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.     

Mapping the quantitative trait loci (QTL) for body shape and conformation measurements on BTA1 in Japanese Black cattle

The detection and mapping of segregating quantitative trait loci (QTL) that influence withers height, hip height, hip width, body length, chest width, chest depth, shoulder width, lumbar width, thurl width, pin bone width, rump length, cannon circumference, chest girth, abdominal width and abdominal girth at weaning was conducted on chromosomal regions of bovine chromosome one. The QTL analysis was performed by genotyping half‐sib progeny of five Japanese Black sires using microsatellite DNA markers. Probability coefficients of inheriting allele 1 or 2 from the sire at specific chromosomal locations were computed. The phenotypic data of progeny were regressed on these probability coefficients in a within‐common‐parent regression analysis using a linear model that included fixed effects of sex, parity and season of birth, as well as age as a covariate. F‐statistics were calculated every 1 cM on a linkage map. Permutation tests of 10 000 iterations were conducted to obtain chromosome‐wide significance thresholds. A significant QTL for chest width was detected at 91 cM in family 3. The detection of this QTL boosts the prospects of implementing marker‐assisted selection for body conformation traits in Japanese Black beef cattle.     

Plasma hormone and metabolite concentrations involved in the somatotropic axis of Japanese Black heifers in association with growth hormone gene polymorphism

Bovine growth hormone (bGH) gene polymorphism of leucine (Leu)-threonine (Thr) (allele A), valine (Val)-Thr (allele B), and Val-methionine (Met) (allele C) at codons 127 and 172 was shown to relate with carcass trait variations in Japanese Black cattle. In this study, 10-mo-old Japanese Black heifers with growth hormone (GH) genotypes AA, AB, BB, AC, BC, and CC (N = 141) were compared for basal GH, insulin-like growth factor-1 (IGF-1), insulin, ghrelin, glucose, and nonesterified fatty acid (NEFA) concentrations. Growth hormone release was also measured as response to growth hormone–releasing hormone (GHRH) (0.4 μg/kg body weight [BW]) using 18 heifers with GH genotypes AA, BB, and CC (n = 6 for each group). The genotype AA heifers showed the greatest BW among genotypes (P < 0.05). Genotype AC, BC, and CC heifers showed greater GH concentrations than genotype AA, AB, or BB heifers, in which genotype CC heifers had the highest concentrations (P < 0.05). However, IGF-1 concentrations did not significantly differ. The genotype AA and BB heifers had a greater GH release at 60 min following GHRH injection than did the genotype CC heifers. The area under the curve (AUC; P < 0.07) and incremental area (IA; P < 0.08) of GH responses to the GHRH challenge tended to be the highest in the genotype AA heifers and the lowest in the genotype CC heifers. In conclusion, GH gene polymorphism altered GH, which may have contributed to differences in BW and carcass traits among genotypes.     

Pool‐based genome‐wide association study identified novel candidate regions on BTA9 and 14 for oleic acid percentage in Japanese Black cattle

Fatty acid composition is an important indicator of beef quality. The objective of this study was to search the potential candidate region for fatty acid composition. We performed pool‐based genome‐wide association studies (GWAS) for oleic acid percentage (C18:1) in a Japanese Black cattle population from the Hyogo prefecture. GWAS analysis revealed two novel candidate regions on BTA9 and BTA14. The most significant single nucleotide polymorphisms (SNPs) in each region were genotyped in a population (n = 899) to verify their effect on C18:1. Statistical analysis revealed that both SNPs were significantly associated with C18:1 (p = .0080 and .0003), validating the quantitative trait loci (QTLs) detected in GWAS. We subsequently selected VNN1 and LYPLA1 genes as candidate genes from each region on BTA9 and BTA14, respectively. We sequenced full‐length coding sequence (CDS) of these genes in eight individuals and identified a nonsynonymous SNP T66M on VNN1 gene as a putative candidate polymorphism. The polymorphism was also significantly associated with C18:1, but the p value (p = .0162) was higher than the most significant SNP on BTA9, suggesting that it would not be responsible for the QTL. Although further investigation will be needed to determine the responsible gene and polymorphism, our findings would contribute to development of selective markers for fatty acid composition in the Japanese Black cattle of Hyogo.     

Allelic frequencies and association with carcass traits of six genes in local subpopulations of Japanese Black cattle

Marker‐assisted selection (MAS) is expected to accelerate the genetic improvement of Japanese Black cattle. However, verification of the effects of the genes for MAS in different subpopulations is required prior to the application of MAS. In this study, we investigated the allelic frequencies and genotypic effects for carcass traits of six genes, which can be used in MAS, in eight local subpopulations. These genes are SCD, FASN and SREBP1, which are associated with the fatty acid composition of meat, and NCAPG, MC1R and F11, which are associated with carcass weight, coat color and blood coagulation abnormality, respectively. The frequencies of desirable alleles of SCD and FASN were relatively high and that of NCAPG was relatively low, and NCAPG was significantly associated with several carcass traits, including carcass weight. The proportions of genotypic variance explained by NCAPG to phenotypic variance were 4.83 for carcass weight. We thus confirmed that NCAPG is a useful marker for selection of carcass traits in these subpopulations. In addition, we found that the desirable alleles of six genes showed no negative effects on carcass traits. Therefore, selection using these genes to improve target traits should not have negative impacts on carcass traits.     

Genome wide association study on beef production traits in Marchigiana cattle breed

A genomewide association study was carried out on a sample of Marchigiana breed cattle to detect markers significantly associated with carcass and meat traits. Four hundred and nine young bulls from 117 commercial herds were genotyped by Illumina 50K BeadChip assay. Eight growth and carcass traits (average daily gain, carcass weight, dressing percentage, body weight, skin weight, shank circumference, head weight and carcass conformation) and two meat quality traits (pH at slaughter and pH 24 h after slaughter) were measured. Data were analysed with a linear mixed model that included fixed effects of herd, slaughter date, fixed covariables of age at slaughter and SNP genotype, and random effects of herd and animal. A permutation test was performed to correct SNP genotype significance level for multiple testing. A total of 96 SNPs were significantly associated at genomewide level with one or more of the considered traits. Gene search was performed on genomic regions identified on the basis of significant SNP position and level of linkage disequilibrium. Interesting loci affecting lipid metabolism (SOAT1), bone (BMP4) and muscle (MYOF) biology were highlighted. These results may be useful to better understand the genetic architecture of growth and body composition in cattle.     

Estimation of the carcass composition from a cross-section of the rib-loin from crossbred Japanese Black × Limousin F2 cattle by computer image analysis

The carcass composition of crossbred Japanese Black × Limousin F2 cattle was examined in order to find an accurate carcass composition equation. The test animals included 17 steers and 17 heifers. The 28 image measurements from the area encircling the vertical line to the thoracic vertebra and the line from the thoracic vertebra between the sixth and seventh rib‐bones were measured by computer image analysis. The relationships between the 29 parameters that added the carcass left side weight of the animal and the carcass composition were suggested. The carcass composition included muscle weight, muscle ratio, fat weight and fat ratio. The carcass composition from steers was estimated by an equation composed of these three or four parameters (R² = 90.80%, 79.30%, 90.75% and 73.70%, respectively). The selected parameters were measured without cutting the thoracic vertebra. The carcass composition from heifers was estimated by an equation composed of two to four parameters (R² = 96.15%, 90.98%, 93.60% and 88.22%, respectively). The parameters for the estimation of the muscle and fat weight, and muscle and fat ratio are very similar. Furthermore, the equations using the parameters could estimate the carcass composition from the Japanese Black × Limousin cattle resource population.     

Whole‐genome resequencing to identify candidate genes for the QTL for oleic acid percentage in Japanese Black cattle

In our previous study, we detected a QTL for the oleic acid percentage (C18:1) on BTA9 in Japanese Black cattle through a genome‐wide association study (GWAS). In this study, we performed whole‐genome resequencing on eight animals with higher and lower C18:1 to identify candidate polymorphisms for the QTL. A total of 39,658 polymorphisms were detected in the candidate region, which were narrowed to 1993 polymorphisms within 23 genes based on allele differences between the high and low C18:1 groups. We subsequently selected three candidate genes, that is, CYB5R4, MED23, and VNN1, among the 23 genes based on their function in fatty acid metabolism. In each candidate gene, three SNPs, that is, CYB5R4 c.*349G > T, MED23 c.3700G > A, and VNN1 c.197C > T, were selected as candidate SNPs to verify their effect on C18:1 in a Japanese Black cattle population (n = 889). The statistical analysis showed that these SNPs were significantly associated with C18:1 (p < 0.05), suggesting that they were candidates for the QTL. In conclusion, we successfully narrowed the candidates for the QTL by detecting possible polymorphisms located within the candidate region. It is expected that the responsible polymorphism can be identified by demonstrating their effect on the gene's function.     

Quantitative trait loci analysis for growth and carcass traits in a half-sib family of purebred Japanese Black (Wagyu) cattle

We used a half-sib family of purebred Japanese Black (Wagyu) cattle to locate economically important quantitative trait loci. The family was composed of 348 fattened steers, 236 of which were genotyped for 342 microsatellite markers spanning 2,664 cM of 29 bovine autosomes. The genome scan revealed evidence of 15 significant QTL (<5% chromosome-wise level) affecting growth and carcass traits. Of the 15 QTL, six QTL were significant at the 5% experiment-wise level and were located in bovine chromosomes (BTA) 4, 5, and 14. We analyzed these three chromosomes in more detail in the 348 steers, with an average marker interval of 1.2 cM. The second scan revealed that the same haplotype of the BTA 4 region (52 to 67 cM) positively affected LM area and marbling. We confirmed the QTL for carcass yield estimate on BTA 5 in the region of 45 to 54 cM. Five growth-related QTL located on BTA 14, including slaughter and carcass weights, were positively affected by the same region of the haplotype of BTA 14 (29-51 cM). These data should provide a useful reference for further marker-assisted selection in the family and positional cloning research. The research indicates that progeny design with moderate genotyping efforts is a powerful method for detecting QTL in a purebred half-sib family.