Characterizing the impact of genomic variants on genome function and ultimately complex traits in livestock is essential for the development of sustainable precision agriculture and comparative genomics. Here, as part of the Farm animal Genotype-Tissue Expression (FarmGTEx) project, we present the pilot phase of the SheepGTEx resource through analyzing 6,761 RNA-sequencing samples of 51 primary tissues in a multi-breed population of sheep. We identify millions of regulatory variants associated with seven types of molecular phenotypes, and fine-map 322,467 primary and 113,968 non-primary effects, revealing a high degree of regulatory allelic heterogeneity. We systematically characterize the pleiotropic effects of these variants on molecular phenotypes, assess their context-specific regulatory patterns across tissues, breeds, sexes, and developmental stages, as well as explore their evolutionary constraints across mammals. Finally, we demonstrate the substantial potential of the SheepGTEx resource (https://sheepgtex.farmgtex.org), by providing examples of regulatory mechanisms underpinning 34 complex traits, population divergence between European and Asian breeds, and adaptive evolution in sheep over the past ten millennia.

The long non-coding RNAs (lncRNAs) can bind to transcription factors or RNA-binding proteins to play important regulatory roles in muscle growth and development. This study investigated the functional role of the LNC_004268 in sheep myoblast proliferation and differentiation, as well as its interaction with the RNA-binding protein hnRNPK and the downstream target gene CNOT2. LNC_004268 shows higher expression in the longissimus dorsi muscle of Small-tailed Han sheep (STH) compared to Sunite sheep (SNT) and is primarily localized in the nucleus. Functionally, LNC_004268 inhibits myoblast proliferation while promoting differentiation and myotube formation. Mechanistically, LNC_004268 binds hnRNPK, stabilizing CNOT2 mRNA. Actinomycin D (ACD) assays confirmed that LNC_004268 or hnRNPK overexpression delays CNOT2 mRNA decay, with combined expression further enhancing stability. Notably, CNOT2 functionally recapitulates LNC_004268's effects on myogenesis. Collectively, our study demonstrates that LNC_004268 regulates myoblast proliferation and differentiation by stabilizing CNOT2 mRNA via hnRNPK interaction, advancing understanding of lncRNA mediated muscle development regulation.

在相同饲养管理条件下,以小尾寒羊和甘肃高山细毛羊母羊作为研究对象,测定产后30 d的泌乳量,分析常乳和初乳中6种乳成分的含量,以及饱和脂肪酸(Saturated fatty acid,SFA)、单不饱和脂肪酸(Mo-nounsaturated fatty acid,MUFA)和多不饱和脂肪酸的含量(Polyunsaturated fatty acids,PUFA).结果表明,小尾寒羊产后30 d的泌乳量为40.7 kg,极显著高于甘肃高山细毛羊的25.6 kg(P＜0.01).在初乳和常乳中,小尾寒羊的乳蛋白质、酪蛋白和可溶固形物含量都高于甘肃高山细毛羊(P＜0.05).在2个品种中,初乳中的乳蛋白质、酪蛋白和可溶固形物含量也高于常乳(P＜0.05).在小尾寒羊初乳、小尾寒羊常乳、甘肃高山细毛羊初乳和甘肃高山细毛羊常乳中分别检测到16、17、13和14种SFA、6种MUFA,以及8、8、7和6种PUFA,棕榈酸、油酸、肉豆蔻酸、硬脂酸是其中的主要成分.上述4种乳样中的SFA/MUFA/PUFA值分别为 13.66/6.89/1.00、16.08/9.16/1.00、10.16/7.89/1.00 和 22.28/8.61/1.00.在不同品种以及泌乳期之间,乳中部分脂肪酸的含量表现出明显差异.说明,小尾寒羊泌乳能力更强,乳中乳蛋白、酪蛋白、可溶固形物及部分有益脂肪酸的含量更高,更有利于羔羊的存活和生长发育.同时,在2个品种中,初乳中均有更高含量的乳蛋白质、酪蛋白质、可溶固形物以及部分有益的MUFA和PUFA.

[目的]研究角蛋白关联蛋白1-4基因(KRTAP1-4)多态性对陇东绒山羊羊绒性状的影响.[方法]以382只1岁龄陇东绒山羊为研究对象,测定其产绒量、绒层高度和绒纤维的平均直径.采集试羊皮肤、肾脏、脾脏、肝脏、肺脏、心脏组织及血样,采用逆转录聚合酶链反应(RT-PCR)检测KRTAP1-4基因在6个组织中的表达谱;采用聚合酶链反应-单链构象多态性(PCR-SSCP)检测KRTAP1-4基因的多态性,分析3个羊绒表型性状间的Pearson相关性及KRTAP14基因型与羊绒性状间的相关性.[结果]RT PCR发现,KRTAP14基因只在陇东绒山羊的皮肤中表达.在陇东绒山羊KRTAP1-4基因上检测到A、B、C、D4个等位基因和AA、AB、AC、AD、BB 5种基因型.测序结果表明,山羊KRTAP14基因上有1个插入/缺失位点(c.223_252insTGCCAACCGATCTCCATCCAGAC-CAGCTGC)、2个同义突变的单核苷酸多态性(c.333T＞C和c.573C＞T),以及1个Chi序列(c.14-c.21,5'-CCACCAGC-3')和2个Chi-like序列(c.46-c.53,5'-ACTGGTGG-3';c.392一c.399,5'-GCACCAGC-3').Pearson相关性系数显示,产绒量与平均纤维直径(r=0.324,P＜0.001)及绒层高度(r=0.465,P＜0.001)呈中等正相关,平均纤维直径与绒层高度呈弱的正相关(r=0.201,P＜0.001).相关性分析结果表明,AA、AB和AC型山羊个体的产绒量和绒层高度极显著低于BB型个体(P＜0.01).[结论]KRTAP1-4基因可以作为产绒量和绒层高度的分子标记用于陇东绒山羊的育种实践中.

Keratin-associated proteins (KAPs) are components of cashmere fibres. The gene encoding the KAP1-3 protein (KRTAP1-3) has been described in goats, but little is known about sequence variation in this gene and if it affects cashmere fibre traits. In this study, we used a polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) technique to screen for nucleotide sequence variation in caprine KRTAP1-3 in 327 Longdong cashmere goats, then analysed association between the genetic variation that was revealed and some cashmere fibre traits. Six PCR-SSCP patterns representing six different variant sequences of KRTAP1-3 (named A to F) were revealed. Among these variant sequences, seven single nucleotide polymorphisms (SNPs) were detected, with two of them being non-synonymous. Goats with genotype AC had higher mean fibre diameter (MFD) than those with genotype AB (P < 0.001), while goats with genotype AB had higher MFD than those with AA (P < 0.001). The presence of C (P < 0.001) and B (P = 0.006) in a genotype was associated with increased MFD, and together this suggests that variation in caprine KRTAP1-3 affects the key fibre trait of MFD.

<span id="ChDivSummary" name="ChDivSummary" class="abstract-text">为鉴定子午岭黑山羊KRTAP36-1基因,并分析其与羊绒性状的相关性,以绵羊KRTAP36-1基因编码区序列为模板,利用GenBank的BLAST功能,在山羊染色体上进行同源性搜索,用聚合酶链反应-单链构象多态性（Polymerase chain reaction-single strand conformation polymorphism,PCR-SSCP）和测序方法,检测342只子午岭黑山羊上的核苷酸序列变异,最后用一般线性混合效应模型分析序列变异与羊绒性状的相关性。结果表明,在342只子午岭黑山羊上检测到3条不同的核苷酸序列（命名为A、B和C）,进化树发现这3条序列与绵羊KRTAP36-1序列有最高的同源性,表明这些序列来自于山羊KRTAP36-1基因。测定结果表明,山羊KRTAP36-1基因有2个SNPs,其中一个（c.119A/G）为错义突变,导致p.Tyr40Cys的氨基酸改变。相关性分析表明,KRTAP36-1核苷酸序列A的存在与子午岭黑山羊较小的产绒量相关（P&lt;0.001）,B的存在与较大的产绒量相关（P=0.004）,BB型个体的产绒量高于AB型和AA型个体（P&lt;0.001）。KRTAP36-1的核苷酸序列变异对子午岭黑山羊的羊绒平均纤维直径和绒层高度无影响（P&gt;0.05）。综上,KRTAP36-1基因可以作为子午岭黑山羊羊绒性状的分子标记,用于提高该群体的产绒量。</span>

角蛋白关联蛋白(KAPs)是羊绒纤维的主要结构成分,但尚未在山羊基因组中鉴定出KRTAP2-1基因,本研究以人类KRTAP2-1基因为模板,在山羊基因组中进行同源性搜索,在19号染色体上发现了一个399 bp的开放阅读框.采用聚合酶链反应-单链构象多态性(PCR-SSCP)方法,在249只陇东山羊上检测到7条不同的序列(A-G).进化树结果表明,这7条序列与人类和绵羊的KAP2-1序列有较高的同源性,表明该序列是山羊KRTAP2-1的7个等位基因.测序结果表明,KRTAP2-1基因中存在12个单核苷酸多态性(SNPs)(5个SNPs位于编码区,7个SNPs位于非编码区),以及c.-61_-63delCTC和c.93_95delCCG两个碱基缺失,其中c.93_95delCCG引起了第32位精氨酸的移码缺失.山羊KAP2-1蛋白含有丰富的半胱氨酸、脯氨酸、苏氨酸和丝氨酸.相关性分析表明,等位基因C的存在与较小的羊绒纤维直径相关(存在:13.4±0.05 μm;缺失:13.6±0.03 μm;P=0.010).本研究表明,山羊KRTAP2-1基因有丰富的多态性,可作为陇东绒山羊羊绒纤维直径选育的分子标记用于生产实践.

MicroRNAs (miRNAs) have been found to be involved in lipid deposition and metabolism. However, there have been no reports on the roles of miR-148a in the proliferation and adipogenesis of preadipocytes in sheep. In this study, the expression of miR-148a was profiled in the eight tissues of Tibetan ewes and differentiated preadipocytes, and the role of miR-148a in differentiation and proliferation of ovine preadipocytes was investigated using Oil Red O staining, CCK-8, EdU staining, cell cycle detection, and RT-qPCR. The effect of PTEN on the differentiation of ovine preadipocytes was also investigated. The miR-148a was widely expressed in the eight tissues investigated and had significantly increased expression in liver, spleen and subcutaneous adipose tissues, and the heart. The expression of miR-148a continued to increase with the differentiation of ovine preadipocytes. The over-expression of miR-148a significantly promoted differentiation but inhibited the proliferation of ovine preadipocytes. The inhibition of miR-148a had the opposite effect on the differentiation and proliferation of ovine preadipocytes with over-expressed miR-148a. The results from the dual luciferase reporter assays showed that miR-148a mimic significantly decreased the luciferase activity of PTEN-3′UTR dual luciferase reporter vector, suggesting that PTEN is a target gene of miR-148a. In over-expressed-PTEN preadipocytes, the number of lipid droplets remarkably decreased, and the expression levels of adipogenesis marker genes PPARγ, FASN, FATP4, GLUT4, C/EBPβ and LPL were also significantly down-regulated. These results suggest that miR-148a accelerated the adipogenic differentiation of ovine preadipocytes by inhibiting PTEN expression, and also inhibited the proliferation of ovine preadipocytes.

Circular RNAs are a class of noncoding RNA with a widespread occurrence in eukaryote tissues, and with some having been demonstrated to have clear biological function. In sheep, little is known about the role of circular RNAs in mammary gland tissue, and therefore an RNA sequencing approach was used to compare mammary gland tissue expression of circular RNAs in 9 Small Tail Han sheep at peak lactation, and subsequently when they were not lactating. These 9 sheep had their RNA pooled for analysis into 3 libraries from peak lactation and 3 from the nonlactating period. A total of 3,278 and 1,756 circular RNAs were identified in the peak lactation and nonlactating mammary gland tissues, respectively, and the expression and identity of 9 of them was confirmed using reverse transcriptase-polymerase chain reaction analysis and DNA sequencing. The type, chromosomal location and length of the circular RNAs identified were ascertained. Forty upregulated and one downregulated circular RNAs were characterized in the mammary gland tissue at peak lactation compared with the nonlactating mammary gland tissue. Gene ontology enrichment analysis revealed that the parental genes of these differentially expressed circular RNAs were related to molecular function, binding, protein binding, ATP binding, and ion binding. Five differentially expression circular RNAs were selected for further analysis to predict their target microRNAs, and some microRNAs reportedly associated with the development of the mammary gland were found in the constructed circular RNA-microRNA network. This study reveals the expression profiles and characterization of circular RNAs at 2 key stages of mammary gland activity, thereby providing an improved understanding of the roles of circular RNAs in the mammary gland of sheep.

1990年,甘肃省庆阳市开始引入辽宁绒山羊对子午岭黑山羊进行杂交改良,但目前尚不清楚2个品种在脂肪酸含量、肌肉营养成分等方面的差异,影响了杂交改良效果.试验旨在分析两个绒山羊品种的产肉性能、肉品质、肌肉营养成分和脂肪酸含量差异,为绒山羊的杂交改良提供理论依据.本研究选取相同饲养管理条件下、9月龄的子午岭黑山羊和辽宁绒山羊公羊各5只,测定其屠宰性能以及背最长肌、前腿肌和后腿肌处的肉品质、脂肪酸含量和肌肉营养成分.结果表明:子午岭黑山羊的胴体重、屠宰率、净肉重、净肉率、眼肌面积、GR值、剪切力和滴水损失低于辽宁绒山羊(P<0.05),但其肌肉的平均亮度值、色度值、pH1和pH24高于辽宁绒山羊(P<0.01).营养成分测定结果表明,子午岭黑山羊肌肉的水分和粗灰分含量高于辽宁绒山羊(P<0.05),但肌内脂肪和粗蛋白含量低于辽宁绒山羊.在2个山羊品种的肌肉中均检测到11种饱和脂肪酸(SFA,以棕榈酸和硬脂酸为主)、10种多不饱和脂肪酸(PUFA,以亚油酸和顺-11,14-二十碳二烯酸为主)和6种单不饱和脂肪酸(MUFA,以油酸为主),子午岭黑山羊肉中的SFA、PUFA、n-3 PUFA、n-6 PUFA含量和PUFA/SFA值均高于辽宁绒山羊(P<0.01),但MUFA含量低于辽宁绒山羊(P<0.01).结果表明,辽宁绒山羊有更高的产肉力,但子午岭黑山羊肌肉品质和营养成分更佳,脂肪酸组成和含量更符合人类健康膳食标准.

Carcass weight, meat quality and muscle components are important traits economically and they underpin most of the commercial return to goat producers. In this study, the Longissimus dorsi muscle tissues were collected from five Liaoning cashmere (LC) goats and five Ziwuling black (ZB) goats with phenotypic difference in carcass weight, some meat quality traits and muscle components. The histological quantitative of collagen fibers and the transcriptome profiles in the Longissimus dorsi muscle tissues were investigated using Masson-trichrome staining and RNA-Seq, respectively. The percentage of total collagen fibers in the Longissimus dorsi muscle tissues from ZB goats was less than those from LC goats, suggesting that these ZB goats had more tender meat. An average of 15,919 and 15,582 genes were found to be expressed in Longissimus dorsi muscle tissues from LC and ZB goats, respectively. Compared to LC goats, the expression levels of 78 genes were up-regulated in ZB goats, while 133 genes were down-regulated. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses revealed that the differentially expressed genes (DEGs) were significantly enriched in GO terms related to the muscle growth and development and the deposition of intramuscular fat and lipid metabolism, hippo signaling pathway and Jak-STAT signaling pathway. The results provide an improved understanding of the genetic mechanisms regulating meat production performance in goats, and will help us improve the accuracy of selection for meat traits in goats using marker-assisted selection based on these differentially expressed genes obtained.

Keratin-associated proteins are important components of wool fibers. The gene encoding the high-sulfur keratin-associated protein 2-1 has been described in humans, but it has not been described in sheep. A basic local alignment search tool nucleotide search of the Ovine Genome Assembly version 4.0 using a human keratin-associated protein 2-1 gene sequence revealed a 399-base pair open reading frame, which was clustered among nine previously identified keratin-associated protein genes on chromosome 11. Polymerase chain reaction–single strand conformation polymorphism analysis revealed four different banding patterns, with these representing four different sequences (A–D) in Chinese sheep breeds. These sequences had the highest similarity to human keratin-associated protein 2-1 gene, suggesting that they represent variants of ovine keratin-associated protein 2-1 gene. Nine single nucleotide variations were detected in the gene, including one non-synonymous nucleotide substitution. Differences in variant frequencies between fine-wool sheep breeds and coarse-wool sheep breeds were detected. The gene was found to be expressed in various tissues, with the highest expression level in skin, and moderate expression levels in heart and lung tissue. These results reveal that the ovine keratin-associated protein 2-1 gene is variable and suggest the gene might affect variation in mean fiber diameter.

MicroRNAs (miRNAs) are small non-coding RNAs that are involved in mammary gland development and lactation in livestock. Little is known about the roles of miRNAs in ovine mammary gland development, hence in this study the expression profiles of miRNAs of the mammary gland tissues of ewes at peak-lactation and during the non-lactating period were investigated using RNA sequencing. A total of 147 mature miRNAs were expressed in the two periods. Compared with peak-lactation, eight miRNAs in the non-lactating ewe mammary gland were significantly up-regulated, whereas fifteen miRNAs were down-regulated. A KEGG analysis revealed that the target genes of the up-regulated miRNAs were significantly enriched in lysosome, Wnt and MAPK signaling pathways, while the target genes of down-regulated miRNAs were significantly enriched in the PI3K-Akt signaling pathway, protein processing in endoplasmic reticulum and axon guidance. These results suggest that further study of the differentially expressed miRNAs could provide a better understanding of the molecular mechanisms of mammary development and lactation in sheep.

Variation in some caprine keratin-associated protein (KAP) genes has been associated with cashmere fiber traits, but many KAP genes remain unidentified in goats. In this study, we confirm the identification of a KAP27-1 gene (KRTAP27-1) and describe its effect on cashmere traits in 248 Longdong cashmere goats. A polymerase chain reaction–single strand conformation polymorphism (PCR-SSCP) analysis was used to screen for sequence variation in this gene, and three sequence variants (named A to C) were found. These sequences have the highest similarity (77% identity) to a human KRTAP27-1 sequence, while sharing some homology with a predicted caprine KRTAP27-1 sequence ENSCHIG00000023347 in the goat genome construct (ARS1:CM004562.1) at chromosome 1 position 3,966,193–3,973,677 in the forward strand. There were two single nucleotide polymorphisms (SNPs) detected in the coding sequence, including one nonsynonymous SNP (c.413C/T; p.Ala138Val) and one synonymous SNP (c.495C/T). The C variant differed from A and B at c.413C/T, having cytosine in its nucleotide sequence, while the B variant differed from A and C at c.495C/T, having thymine in its nucleotide sequence. Goats of the genotypes AB and BB produced cashmere fibers of higher mean fiber diameter (MFD) than goats of genotype AA, but no difference in MFD was detected between the AB and BB goats. These results suggest that B is associated with increased MFD. Expression of the caprine KRTAP27-1 sequence was predominantly detected in the skin tissue of goats but not or only weakly detected in other tissues, including longissimus dorsi muscle, heart, kidney, liver, lung and spleen.

羊源CXC趋化因子受体2基因(chemokine C-X-C motif receptor 2,CXCR2)主要表达于炎性细胞表面,在局部炎症反应、肿瘤发生等方面起重要调控作用.为研究CXCR2基因在患临床型乳房炎绵羊乳腺组织中的表达差异及可能的生物学功能,本研究选取正常和患临床型乳房炎的绵羊(Ovis aries)各3只,收集乳腺组织,HE染色观察其组织形态学结构差异,运用实时荧光定量PCR (qRT-PCR)和ELISA技术分别从mRNA和蛋白水平检测两者中CX-CR2的表达差异,通过miRbase数据库和miranda软件对绵羊CXCR2基因的靶向miRNAs进行预测,并采用DA-VID软件对CXCR2基因的生物功能进行分析.结果表明,与正常组相比,患病组的乳腺组织中结缔组织明显增生,腺泡腔萎缩,腔内含有脱落的上皮细胞并且有大量白细胞浸润;患病组中CXCR2 mRNA和蛋白的表达量极显著上调(P＜0.01);靶向miRNAs预测显示,绵羊CXCR2基因可能受到oar-miR-3956-3 p、oar-miR-665-5p、oar-miR-432等miRNAs的调控;GO分析发现CXCR2基因参与构成膜的主要成分、CXC趋化因子受体活性、趋化作用等过程;KEGG通路富集发现CXCR2基因参与细胞因子—受体相互作用、趋化因子信号转导通路和内吞作用信号通路.由此推测,在绵羊乳房炎发生过程中,CXCR2基因可能通过与配体结合,以调控炎性细胞的趋化迁移,进而参与免疫应答反应.本研究为更深入地研究和探讨CXCR2基因在乳房炎发生发展过程中的分子生物学功能提供了研究素材.