Preliminary genetic parameter estimates of meat quality traits in Hu sheep.

Because substantial numbers of Chinese consumers are prepared to pay for tender and quality lamb, meat quality traits are becoming more relevant for breeding programs for Chinese sheep breeds. The current study estimated heritabilities and genetic correlations for 13 meat quality traits recorded on lamb loins from Hu sheep. Heritability estimates ranged from 0.04 ± 0.06 for meat redness at 45 min to 0.57 ± 0.10 for drip loss, with most of the meat quality traits having moderate heritabilities. Positive genetic correlations were observed among meat color traits. Intramuscular fat (IMF) was genetically correlated with most meat quality traits, indicating that increasing IMF can favor meat pH, color, and tenderness, but would lead to increased cooking loss. Direct selection to increase IMF of loins is recommended to be included in breeding programs for Hu sheep, as it was more efficient than indirect selection on the other meat quality traits. The genetic parameters presented in this preliminary study provide valuable genetic information needed to design a breeding program aimed at improving the quality of lamb meat from Hu sheep.

Transcriptomic analysis of the HPG axis-related tissues reveals potential candidate genes and regulatory pathways associated with testicular size in Hu sheep.

Testicular size is an excellent proxy for selecting high-fertility rams. The hypothalamus-pituitary-gonadal (HPG) axis plays an important role in regulating reproductive capacity in vertebrates, while key genes and regulatory pathways within the HPG axis associated with testicular size remain largely unknown in sheep. This study comprehensively compared the transcriptomic profiles in the hypothalamus, pituitary and testis of rams after sexual maturity between the large-testis group (LTG, testicular weight = 454.29 ± 54.24 g) and the small-testis group (STG, testicular weight = 77.29 ± 10.76 g). In total, 914, 795 and 10518 differentially expressed genes (DEGs) were identified in the hypothalamus, pituitary and testis between LTG and STG, respectively. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses showed that these DEGs were mainly involved in the biological processes of reproduction, biological regulation, and development process. Notably, the neuroactive ligand-receptor interaction and cAMP signaling pathways, commonly enriched by the DEGs in the hypothalamus and pituitary between two groups, were considered as two key signal pathways regulating testicular development through the HPGs axis. Weighted gene co-expression network analysis (WGCNA) identified two modules that were significantly associated with testicular size, and 97 key genes were selected with high module membership (MM) and gene significance (GS) in these two modules. Finally, a protein-protein interaction (PPI) network was constructed, and ten genes with the highest degree were represented as hub genes, including FOS, NPY, SST, F2, AGT, NTS, OXT, EDN1, VIP and TAC1. Taken together, these results provide new insights into the molecular mechanism underlying the HPG axis regulating testicular size of Hu sheep.

Localization and expression of phospholipase A2 and polyunsaturated fatty acid profile in the testis tissues of Hu sheep.

The fatty acid content and the localization and expression of phospholipase A2 (PLA2) in the testis of Hu sheep were investigated. A total of 18 six-month-old Hu sheep were divided into small group (S, with left testis weight < 50 g), medium group (M, with left testis weight among 90-110 g), and large group (L, with left testis weight >160 g), which had six individuals each. The expression of PLA2 in testicular tissues of different sizes was analyzed by immunohistochemistry, RT-qPCR, and Western blot. The fatty acid profile was detected by gas chromatography. Immunohistochemical labeling determined that PLA2 protein was expressed in the Leydig and Sertoli cells of testis, and the immunohistochemical average optional density in the S group was significantly greater than the L group (P < 0.05). RT-qPCR and Western blot analysis showed that PLA2 in the S group was greater than that in the L group (P < 0.05). Docosahexaenoic acid, ω-3 polyunsaturated fatty acid (PUFA), and total PUFA content in the testis of the L group were significantly less than those of the S and M groups (P < 0.01). This study showed that PLA2 content in the S group was greater than that in the L group.

Molecular characterization of SPATA6 and association of its SNPs with testicular size in sheep.

The testis is an important organ for maintaining fertility in males, and testis size is positively correlated with ejaculate volume, sperm motility, thus fertility. Spermatogenesis-associated 6 (SPATA6) is an evolutionarily conserved testis-specific gene reported in many species. However, the effect of SPATA6 expression levels on testicular development and the effect of single nucleotide polymorphisms (SNPs) on testis and epididymis phenotype in sheep have not been studied. The purpose of the research was to investigate the expression profile of SPATA6 and its effect on testicular development and to confirm the effect of SNPs on the testis and epididymis phenotype. In this study, we detected a 1245bp coding sequence (CDS) of SPATA6 and encoded 414 amino acids. The expression levels of SPATA6 were significantly higher in the testis than in other tissues and gradually increased with testis development. Moreover, the expression level in the large testis was significantly higher than that in the small testis at six months. A total of 11 SNPs were detected in the coding region of SPATA6 by cDNA-pooling sequencing and improved multiplex ligation detection reaction (iMLDR) methods. Correlation analysis showed that SNP2 (c. 3631C > G) significantly affected left epididymis weight (LEW) and right epididymis weight (REW), and SNP10 (c. 937 A > G) significantly affected REW. And the combined genotype of SNP1 (c. 4245 G > A) and SNP2 significantly affected REW. The current study concluded that SPATA6 plays an important role in testicular development and the SNPs significantly associated with the epididymis phenotype can provide molecular markers for the early selection of high-fertility Hu sheep.

Supplementation of chestnut tannins in diets can improve meat quality and antioxidative capability in Hu lambs.

Chestnut tannins (CNT), as a source of hydrolyzable tannins, positively affect the antioxidant status of livestock. In the current study, 90 male Hu lambs were used to investigate the effect of dietary CNT intake on growth performance, nutrient digestibility, meat quality and oxidative stability, rumen microbial, and the transcriptomes of muscle and liver. A completely randomized design with three CNT intake levels (0, 0.3%, and 0.6%) was used. Rumen microbial and nutrient digestibility were not significantly altered by CNT intake. Diets with 0.3% CNT intake significantly reduced the shear force, yellowness at 24 h, and C20:2 polyunsaturated fatty acids of lamb meat and malondialdehyde in serum and longissimus thoracis (LT) muscle. Meanwhile, the 0.3% CNT diet significantly increased average daily gain during the 1- 21 days and 64- 90 days, dry matter intake during the 1- 21 days, the slaughter weight, and liver index of lambs. The 0.3% CNT diet significantly increased C26:0 saturated fatty acids, total antioxidant capacity, glutathione peroxidase, superoxide dismutase, and catalase in LT muscle. The meat shelf life of 0.3% CNT and 0.6% CNT groups was prolonged by 8.7 h and 5.4 h, respectively. Transcriptomic analysis revealed that CNT supplementation can induce the expression of antioxidant enzyme gene (CAT, SOD1), and the differentially expressed genes were mainly involved in antioxidant activity, transferase activity, and adenosine triphosphate binding. These results suggest that 0.3% CNT intake can relieve the oxidative stress of lambs, and improve the stability of meat color and meat tenderness, due to the enhanced antioxidative capacity.

Heritability and recursive influence of host genetics on the rumen microbiota drive body weight variance in male Hu sheep lambs.

BACKGROUND: Heritable rumen microbiota is an important modulator of ruminant growth performance. However, no information exists to date on host genetics-rumen microbiota interactions and their association with phenotype in sheep. To solve this, we curated and analyzed whole-genome resequencing genotypes, 16S rumen-microbiota data, and longitudinal body weight (BW) phenotypes from 1150 sheep. RESULTS: A variance component model indicated significant heritability of rumen microbial community diversity. Genome-wide association studies (GWAS) using microbial features as traits identified 411 loci-taxon significant associations (P < 10-8). We found a heritability of 39% for 180-day-old BW, while also the rumen microbiota likely played a significant role, explaining that 20% of the phenotypic variation. Microbiota-wide association studies (MWAS) and GWAS identified four marker genera (Bonferroni corrected P < 0.05) and five novel genetic variants (P < 10-8) that were significantly associated with BW. Integrative analysis identified the mediating role of marker genera in genotype influencing phenotype and unravelled that the same genetic markers have direct and indirect effects on sheep weight. CONCLUSIONS: This study reveals a reciprocal interplay among host genetic variations, the rumen microbiota and the body weight traits of sheep. The information obtained provide insights into the diverse microbiota characteristics of rumen and may help in designing precision microbiota management strategies for controlling and manipulating sheep rumen microbiota to increase productivity. Video Abstract.

iTRAQ-based proteomic analysis provides novel insight into the postnatal testicular development of Hu sheep.

Testicular development is an intricate and coordinated process in which thousands of proteins are involved in the regulation of somatic cells development and spermatogenesis. However, knowledge about the proteomic changes during postnatal testicular development in Hu sheep is still elusive. The study was conducted to characterize the protein profiles at four key stages during postnatal testicular development, including infant (0-month-old, M0), puberty (3-month-old, M3), sexual maturity (6-month-old, M6) and body maturity (12-month-old, M12), and between the large- and small-testis groups at 6 months in Hu sheep. Consequently, 5252 proteins were identified using isobaric tags for relative and absolute quantification (iTRAQ) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods, and 465, 1261, 231 and 1080 differentially abundant proteins (DAPs) were found between M0_vs_M3, M3_vs_M6L, M6L_vs_M12, and M6L_vs_M6S, respectively. GO and KEGG analysis revealed that the majority of DAPs were involved in cellular process, metabolic process and immune system-related pathways. Furthermore, a protein-protein interaction network was constructed using 86 fertility-related DAPs, and five proteins with the highest degree were represented as hub proteins, including CTNNB1, ADAM2, ACR, HSPA2 and GRB2. This study provided new insights into the regulation mechanisms of postnatal testicular development and identified several potential biomarkers for selecting the high-fertility rams. SIGNIFICANCE OF THE STUDY: Testicular development is an intricate developmental process in which thousands of proteins are involved in regulating the somatic cells development and spermatogenesis. However, knowledge about the proteome changes during postnatal testicular development in Hu sheep is still elusive. This study provides comprehensive insights into the dynamic changes in the sheep testis proteome during postnatal testicular development. Additionally, testis size is positively correlated with semen quality and ejaculation volume, also for the merits of easy measurement, high heritability and selection efficiency, is an important indicator to select candidate rams with high fertility. The functional analyses of the acquired candidate proteins may help us gain a better understanding of the molecular regulatory mechanisms of testicular development.

A sheep pangenome reveals the spectrum of structural variations and their effects on tail phenotypes.

Structural variations (SVs) are a major contributor to genetic diversity and phenotypic variations, but their prevalence and functions in domestic animals are largely unexplored. Here we generated high-quality genome assemblies for 15 individuals from genetically diverse sheep breeds using Pacific Biosciences (PacBio) high-fidelity sequencing, discovering 130.3 Mb nonreference sequences, from which 588 genes were annotated. A total of 149,158 biallelic insertions/deletions, 6531 divergent alleles, and 14,707 multiallelic variations with precise breakpoints were discovered. The SV spectrum is characterized by an excess of derived insertions compared to deletions (94,422 vs. 33,571), suggesting recent active LINE expansions in sheep. Nearly half of the SVs display low to moderate linkage disequilibrium with surrounding single-nucleotide polymorphisms (SNPs) and most SVs cannot be tagged by SNP probes from the widely used ovine 50K SNP chip. We identified 865 population-stratified SVs including 122 SVs possibly derived in the domestication process among 690 individuals from sheep breeds worldwide. A novel 168-bp insertion in the 5' untranslated region (5' UTR) of HOXB13 is found at high frequency in long-tailed sheep. Further genome-wide association study and gene expression analyses suggest that this mutation is causative for the long-tail trait. In summary, we have developed a panel of high-quality de novo assemblies and present a catalog of structural variations in sheep. Our data capture abundant candidate functional variations that were previously unexplored and provide a fundamental resource for understanding trait biology in sheep.

Whole genome sequencing of simmental cattle for SNP and CNV discovery.

BACKGROUD: The single nucleotide polymorphisms (SNPs) and copy number variations (CNVs) are two major genomic variants, which play crucial roles in evolutionary and phenotypic diversity. RESULTS: In this study, we performed a comprehensive analysis to explore the genetic variations (SNPs and CNVs) of high sperm motility (HSM) and poor sperm motility (PSM) Simmental bulls using the high-coverage (25×) short-read next generation sequencing and single-molecule long reads sequencing data. A total of ~ 15 million SNPs and 2,944 CNV regions (CNVRs) were detected in Simmental bulls, and a set of positive selected genes (PSGs) and CNVRs were found to be overlapped with quantitative trait loci (QTLs) involving immunity, muscle development, reproduction, etc. In addition, we detected two new variants in LEPR, which may be related to the artificial breeding to improve important economic traits. Moreover, a set of genes and pathways functionally related to male fertility were identified. Remarkably, a CNV on SPAG16 (chr2:101,427,468 - 101,429,883) was completely deleted in all poor sperm motility (PSM) bulls and half of the bulls in high sperm motility (HSM), which may play a crucial role in the bull-fertility. CONCLUSIONS: In conclusion, this study provides a valuable genetic variation resource for the cattle breeding and selection programs.

Characterization of meat quality traits, fatty acids and volatile compounds in Hu and Tan sheep.

Sheep breed has a major influence on characteristics of meat quality and intramuscular fat (IMF), however, studies into the relationship between sheep breed and meat quality traits rarely consider the large variation in IMF within breed. In this study, groups of 176 Hu and 76 Tan male sheep were established, weaned at 56 days old, with similar weights, and representative samples were selected based on the distribution of IMF in each population, to investigate variations in meat quality, IMF and volatile compound profiles between breeds. Significant differences were observed in drip loss, shear force, cooking loss, and color coordinates between Hu and Tan sheep (p < 0.01). The IMF content and the predominate unsaturated fatty acids, oleic and cis, cis-linoleic acids, were similar. Eighteen out of 53 volatile compounds were identified as important odor contributors. Of these 18 odor-active volatile compounds, no significant concentration differences were detected between breeds. In another 35 volatile compounds, γ-nonalactone was lower in Tan sheep relative to Hu sheep (p < 0.05). In summary, Tan sheep exhibited lower drip loss, higher shear force values, and redder color, had less saturated fatty acids, and contained less γ-nonalactone against Hu sheep. These findings improve understanding of aroma differences between Hu and Tan sheep meat. Graphical Abstract.

Effect of corn straw or corncobs in total mixed ration during peri-puberty on testis development in Hu lambs.

Corn straw and corncobs contain large amounts of crude fibers and are widely used in mutton sheep husbandry in northwest China. The aim of this study was to determine whether feeding with corn straw or corncobs affects lamb testis development. A total of 50 healthy Hu lamb at two-month-old (average body weight of 22.3 ± 0.1 kg) were randomly and equally divided into two groups, and the lambs were equally allocated to five pens in each group. The corn straw group (CS) received a diet containing 20% corn straw, whereas the corncobs group (CC) received a diet containing 20% corncobs. After a 77-day feeding trial, the lambs, except the heaviest and lightest in each pen, were humanely slaughtered and investigated. Results revealed no differences in body weight (40.38 ± 0.45 kg vs. 39.08 ± 0.52 kg) between the CS and CC groups. Feeding diet containing corn straw significantly (P < 0.05) increased testis weight (243.24 ± 18.78 g vs. 167.00 ± 15.20 g), testis index (0.60 ± 0.05 vs. 0.43 ± 0.04), testis volume (247.08 ± 19.99 mL vs. 162.31 ± 14.15 mL), diameter of seminiferous tubule (213.90 ± 4.91 µm vs. 173.11 ± 5.93 µm), and the number of sperm in the epididymis (49.91 ± 13.53 × 108/g vs. 19.34 ± 6.79 × 108/g) compared with those in the CC group. The RNA sequencing results showed 286 differentially expressed genes, and 116 upregulated and 170 downregulated genes were found in the CS group compared with the CC group. The genes affecting immune functions and fertility were screened out. Corn straw decreased the mtDNA relative copy number in the testis (P < 0.05). These results suggest that compared with corncobs, feeding corn straw in the early reproductive development stage of lambs increased the testis weight, diameter of seminiferous tubule and the number of cauda sperm.

Genomic analysis reveals a KIT-related chromosomal translocation associated with the white coat phenotype in yak.

White coat pigmentation is a striking phenotype of many domesticated species and has various genetic controls. The Tianzhu White yak, an indigenous breed with a complete white coat, has fascinated Tibetans for centuries. However, the genetic basis of this trait remains unknown. Here, we conducted population genomics analysis and genome-wide association study based on the whole-genome sequencing data of 38 white and 59 non-white-coated yak. The results revealed the presence of KIT-linked Cs alleles characterized by the translocations between chromosomes 6 and 29 in all-white yak. Furthermore, structural variations showed additional duplications of the Cs alleles in white yak compared with colour-sidedness cattle. Interestingly, the Cs alleles associated with the white coat phenotype in yak were found to have introgressed from taurine cattle. Our findings unveil the shared genetic control of the white coat phenotype and its evolution in closely related bovine species.

Association of SNPs within PTPN3 gene with wool production and growth traits in a dual-purpose sheep population.

Protein tyrosine phosphatase non-receptor type 3 (PTPN3), a member of the membrane-associated non-receptor protein tyrosine phosphatase (PTP) family, plays significant roles in the cytoplasm and affects the development and growth of skin and hair. A recent study identified the PTPN3 as the potential gene related to sheep wool quality. To detect single-nucleotide polymorphisms (SNPs) of PTPN3 and elucidate its association with wool production and growth traits in fine wool sheep a total of 644 healthy SG (South African mutton merino♂ × Gansu alpine fine-wool sheep♀, SG) and SSG (South African mutton merino♂ × SG♀, SSG) hybrid sheep were selected. Pooled-DNA sequencing and SNPscan methods were used to scan and genotype SNPs within PTPN3. Association analyses between SNPs and wool production and growth traits were implemented. Consequently, the results revealed that PTPN3 has six SNPs (two missense mutations, one synonymous mutation, and three intron mutations), of which four loci (SNP2, SNP3, SNP4, and SNP5) were significantly positively correlated with growth and wool traits (p < 0.05). SNP4 was significantly (p < 0.05) linked with thigh wool length, and SNP6 was significantly (p < 0.05) associated with abdomen wool length. Moreover, one strongly linked SNP block was identified to be correlated with wool production and growth traits (body weight and body size). The significant SNPs founded by this study could serve as useful genetic markers for breeding fine-wool sheep.®.

Variants of ADPGK gene and its effect on the male reproductive organ parameters and sperm count in Hu sheep.

ADP-dependent glucokinase (ADPGK) plays an important role instead of hexokinase in regulating energy metabolism via the Embden-Meyerhof-Parnas Pathway. And energy provided via glycolysis promotes testis development and spermatogenesis. In this study, 466 Hu sheep were screened for mutations in the ADPGK gene to examine the association of the ADPGK gene polymorphisms with the testis traits and spermatogenesis. The NC_056060.1: g.31295 C > T SNP was found in the 3'-UTR region, resulting in two genotypes CC and TC type with genotypic frequencies of 0.66 and 0.34, respectively. This mutation was significantly associated with testis weight, testis long circumference, testis short girth, epididymis weight, and sperm concentration (p < 0.05). Moreover, TC genotype individuals had an increased tendency in the expression of the ADPGK gene and had significant reproductive performance advantages compared with CC genotype individuals in the study. And compared with the small testes (<50 g), the ADPGK gene expression of big testes (>160 g) increased significantly. This indicates an association between the ADPGK gene and reproductive organ parameters and sperm count in selected Hu sheep breed, and this SNP may serve as an effective DNA molecular marker for marker-assisted selection in Hu sheep breeding programs.

Identification of Y-SNPs within ovine MSY region and their association with testicular size.

The screening of genomic variations within the male-specific region of the mammalian Y chromosome (MSY) is one of the most effective ways to investigate paternal evolutionary history and identify molecular markers related to male fertility. The current study was to identify single nucleotide polymorphisms (SNPs) within single-copy genes of the ovine MSY, and confirm whether they are associated with testicular size. A total of 21 Y-specific gene fragments were successfully amplified to screen Y-SNPs in 956 rams across nine sheep breeds. Three Y-SNPs, including SRY16: g.88 A > G in South African Mutton Merino sheep, ZFY16: g.146 C > T in Suffolk and South African Mutton Merino sheep, and EIF2S3Y2: g.77 C > G in Hu and Tan sheep, were identified using DNA-pooled sequencing and PCR restriction fragment length polymorphism (PCR-RFLP) methods. The investigation of the global distribution for three Y-SNPs showed that the C allele of ZFY16: g.146 C > T co-segregated with haplogroup y-HC, and the C/G allele of EIF2S3Y2: g.77 C > G co-segregated with haplogroup y-HA/y-HB1 in Hu sheep according to data mining from a previous study. In addition, association analysis revealed that ZFY16: g.146 C > T had a significant effect on yearling scrotal circumference in Suffolk sheep, and EIF2S3Y2: g.77 C > G was significantly associated with testicular and epididymis weight in Hu sheep (P ≤ 0.05). The current study concluded that Y-SNPs were associated with testicular size in specific sheep, which provides valuable candidate makers for selecting elite rams at an early age.

Characterization of fatty acid compositions in longissimus thoracis muscle and identification of candidate gene and SNPs related to polyunsaturated fatty acid in Hu sheep.

Fatty acid (FA) composition contributes greatly to the quality and nutritional value of lamb meat. In the present study, FA was measured in longissimus thoracis (LT) muscles of 1,085 Hu sheep using gas chromatography. Comparative transcriptomic analysis was conducted in LT muscles to identify differentially expressed genes (DEGs) between six individuals with high polyunsaturated fatty acids (H-PUFA, 15.27% ± 0.42%) and six with low PUFA (L-PUFA, 5.22% ± 0.25%). Subsequently, the single nucleotide polymorphisms (SNPs) in a candidate gene PLIN2 were correlated with FA traits. The results showed a total of 29 FA compositions and 8 FA groups were identified, with the highest content of monounsaturated fatty acids (MUFA, 46.54%, mainly C18:1n9c), followed by saturated fatty acids (SFA, 44.32%, mainly C16:0), and PUFA (8.72%, mainly C18:2n6c), and significant correlations were observed among the most of FA traits. Transcriptomic analyses identified 110 upregulated and 302 downregulated DEGs between H-PUFA and L-PUFA groups. The functional enrichment analysis revealed three significant Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways and 17 gene ontology (GO) terms, in which regulation of lipolysis in adipocytes, the AMPK signaling pathway, and the PPAR signaling pathway may play important roles in FA metabolism and biosynthesis. In addition, weighted gene co-expression network analysis (WGCNA) identified 37 module genes associated with PUFA-related traits. In general, PLIN1, LIPE, FABP4, LEP, ACACA, ADIPOQ, SCD, PCK2, FASN, PLIN2, LPL, FABP3, THRSP, and ACADVL may have a great impact on PUFA metabolism and lipid deposition. Four SNPs within PLIN2 were significantly associated with FA. Of those, SNP1 (g.287 G>A) was significantly associated with C18:1n9c and MUFA, and SNP4 (g.7807 T>C) was significantly correlated with PUFA (C18:3n3). In addition, the combined genotype of SNP1 (g.287 G>A), SNP3 (g.7664 T>C), and SNP4 (g.7807 T>C) were significantly correlated with C16:1, C17:0, C18:1C6, PUFA (C18:3n3, C22:6n3), and n-6/n-3 PUFA. These results contribute to the knowledge of the biological mechanisms and genetic markers involved in the composition of FA in Hu sheep.

In this study, 29 fatty acids (FAs) compositions and 8 FA groups were identified in 1,085 longissimus thoracis muscles of Hu sheep, and a significant correlation was observed among most FA traits. Transcriptomic analyses identified 412 differentially expressed genes (DEGs) between high polyunsaturated fatty acid (H-PUFA) and low PUFA (L-PUFA) groups. The functional enrichment analysis revealed that regulation of lipolysis in adipocytes, the AMPK signaling pathway, and the PPAR signaling pathway may play important roles in FA metabolism and biosynthesis. In general, PLIN1, LIPE, FABP4, LEP, ACACA, ADIPOQ, SCD, PCK2, FASN, PLIN2, LPL, FABP3, THRSP, and ACADVL may affect PUFA metabolism and lipid deposition. Four single nucleotide polymorphisms (SNPs) within PLIN2 were significantly associated with FA, especially SNP4 (g.7807 T>C) significantly correlated with C18:3n3, the combined genotype significantly associated with C18:3n3, C22:6n3, and n-6/n-3 PUFA. These results contribute to the knowledge of biological mechanisms and genetic markers involved in the FA composition of Hu sheep.

Effects of pre-partum dietary crude protein level on colostrum fat globule membrane proteins and the performance of Hu ewes and their offspring.

Dietary proteins play important roles in the growth and reproduction of sheep, and the ewe's demand for proteins increases dramatically during late pregnancy. This research aimed to investigate the effect of dietary crude protein (CP) levels during late pregnancy on colostrum fat globule membrane (MFGM) protein and the growth performance of Hu sheep and their offspring, and provide a reference for the protein intake of ewes during late pregnancy. A total of 108 multiparous Hu sheep (45.6 ± 1.18 kg) were selected for this study, then 60 pregnant ewes confirmed by B-scan ultrasonography were randomly divided into three treatments (20 ewes/treatment) and fed by total mixed ration pellet with CP levels at 9.00% (LP), 12.0% (MP), and 15.0% (HP) during late pregnancy, respectively. The weight and dry matter intake of ewes during late pregnancy were recorded to calculate the average daily gain (ADG) and feed conversion ratio (FCR). Twin lambs were weighed on days 0, 7, 14, 30, 60, and 180 after birth to calculate ADG. Meanwhile, the colostrum of ewes was collected within 12 h after delivery. The colostrum MFGM proteins were identified and quantified by the isobaric tag for relative and absolute quantification (iTRAQ) coupled with liquid chromatography-tandem mass spectrometry methods. In addition, biological functions of differentially expressed proteins (DEPs) were annotated by Gene Ontology annotation and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis. The results revealed that a 15.0% CP level had significant effects on the BW of lambs on days 0, 7, and 30 (P < 0.05). Notably, a total of 1,529 MFGM proteins were identified and 286 DEPs were found among three treatments. Functional analysis showed that DEPs were mainly involved in cell growth, differentiation, and tissue repair, and involved in metabolic pathways, such as the porphyrin and chlorophyll metabolism pathways. In this study, lambs in HP treatment had better growth performance; moreover, dietary 15.0% CP level also affected the colostrum MFGM proteins composition of Hu ewes. These observations can facilitate future studies on the feeding regimen of ewes during late pregnancy.

Composition of Fatty Acids and Localization of SREBP1 and ELOVL2 Genes in Cauda Epididymides of Hu Sheep with Different Fertility.

The epididymis is an organ that transports, matures and stores sperm, and has functions such as secretion and absorption. Polyunsaturated fatty acid (PUFA) compositions in sperm membrane were changed during the process of epididymis maturation and influence the male fertility. This study aimed to investigate differences in crude fat and fatty acid content in cauda epididymis between high and low fertility of Hu sheep. One hundred and seventy-nine Hu ram lambs were fed from 56 days to 6 months under the same environment. After the feeding trial, all rams were slaughtered, and the body weight, testicular weight, epididymal weight and sperm density were measured. Pearson correlation analysis showed significantly moderate positive correlation between epididymal weight and sperm density and testicular weight. Eighteen rams were selected and divided into the high fertility group (H, n = 9) and low fertility group (L, n = 9) according to the epididymal weight, sperm density and histomorphology. The crude fat content, fatty acid profile and genes related to fatty acid metabolism were detected. The crude fat content, total fatty acid, total n-3 PUFA and docosahexaenoic acid (C22:6n-3, DHA) content of cauda epididymis in high fertility group was significantly higher than those in low fertility group (p < 0.05). However, the ratio of n-6/n-3 PUFA was significantly lower than that in group L (p < 0.05). Immunohistochemistry results showed that SREBP1 and ELOVL2 were expressed in pseudostratified columnar ciliated epithelium and smooth muscle cells. The mRNA expression of SREBP1 (p = 0.09) and ELOVL2 (p < 0.05) in the high fertility group were increased. In conclusion, the high expression of SREBP1 and ELOVL2 may contribute to high n-3 PUFA content in cauda epididymis of high-fertility Hu sheep.

Expression profile of FASN gene and association of its polymorphisms with intramuscular fat content in Hu sheep.

The content of intramuscular fat (IMF) is one of the most important factors that has a large impact on meat quality, and it is an effective way to improve IMF according to marker-assisted selection (MAS). Fatty-acid synthase (FASN) is a key gene in meat lipid deposition and fatty acid composition. Thus, this study was conducted to investigate the expression profile of FASN in mRNA and protein levels using real-time quantitative PCR (RT-qPCR) and western-blot methods. In addition, single nucleotide polymorphisms (SNPs) within FASN in 921 Hu rams with IMF content records were investigated using DNA-pooling sequencing and improved multiple ligase detection reaction (iMLDR) methods. Consequently, the highest mRNA expression level of FASN was observed in the perinephric fat, and the lowest in the liver among the 11 tissues analyzed, while no significant difference was found in mRNA and protein expression levels in longissimus dorsi among individuals with different IMF contents. A total of 10 putative SNPs were identified within FASN, and 9 of them can be genotyped by iMLDR method. Notably, two SNPs were significantly associated with IMF content, including NC_040262.1: g.5157 A > G in intron 5 (p = 0.046) and NC_040262.1: g.9413 T > C in intron 16 (p = 0.041), which supply molecular markers for improving meat quality in sheep breeding.

PRAMEY: A Bovid-Specific Y-Chromosome Multicopy Gene Is Highly Related to Postnatal Testicular Growth in Hu Sheep.

PRAMEY (preferentially expressed antigen in melanoma, Y-linked) belongs to the cancer-testis antigens (CTAs) gene family and is predominantly expressed in testis, playing important roles in spermatogenesis and testicular development. This study cloned the full-length cDNA sequence of ovine PRAMEY using the rapid amplification of cDNA ends (RACE) method and analyzed the expression profile and copy number variation (CNV) of PRAMEY using quantitative real-time PCR (qPCR). The results revealed that the PRAMEY cDNA was 2099 bp in length with an open reading frame (ORF) of 1536 bp encoding 511 amino acids. PRAMEY was predominantly expressed in the testis and significantly upregulated during postnatal testicular development. The median copy number (MCN) of PRAMEY was 4, varying from 2 to 25 in 710 rams across eight sheep breeds. There was no significant correlation between the CNV of PRAMEY and testicular size, while a significant positive correlation was observed between the mRNA expression and testicular size in Hu sheep. The current study suggests that the expression levels of PRAMEY were closely associated with testicular size, indicating that PRAMEY may play an important role in testicular growth.