Abstract Backgrounds Transforming growth factor‐β (TGF‐β) type I receptor (TGFβRI) and type II receptor (TGFβRII) are the members of the TGFβ superfamily, which are potent regulators of cell proliferation and differentiation in many organ systems, and they play key roles in multiple aspects of follicle development. Objectives We aimed to explore the characterization, expression analysis of TGFβRI and TGFβRII genes, and the association with litter size in Tibetan sheep. Methods In this study, we cloned the complete coding sequences of TGFβRI and TGFβRII genes in Tibetan sheep and analyzed their genomic structures. Results The results showed that percentages of sequences homology of the two proteins in Tibetan sheep were the most similar to Ovis aries (100%), followed by Bos mutus (99%). The RT‐qPCR showed that two genes were expressed widely in the different tissues of Tibetan sheep. The TGFβRI expression was the highest in the lung (p < 0.05), followed by the spleen and ovary (p < 0.05). The TGFβRII expression was significantly higher in uterus than that in lung and ovary (p < 0.05). In addition, the χ2 test indicated that all ewes in the population were in Hardy–Weinberg equilibrium, and the population was in medium or low polymorphic information content status. We also found four Single Nucleotide Polymorphism (SNPs), g.9414A > G, g.28881A > G, g.28809T > C, g.10429G > A in sheep TGFβRI gene and g.63940C > T, g.63976C > T, g.64538C > T, g.64504T > A in TGFβRII gene. Three genotypes, except for g.64504T > A, and three haplotypes were identified in each gene. linkage disequilibrium analysis indicated that there was strong linkage disequilibrium in each gene. The association analysis showed that the four SNPs of TGFβRI were associated with litter size (p < 0.05), and g.63940C > T of TGFβRII was confirmed to be associated with litter size (p < 0.05). Conclusions Based on these preliminary results, we can assume that TGFβ receptors (TGFβRI and TGFβRII) may play an important role in sheep reproduction.

Abstract Tibetan sheep is a unique breed living in Qinghai‐Tibet Plateau. Since MSTN/Smad signaling pathway plays a critical role in the regulation of muscle development, we aimed to study the mutton quality, mRNA expression of main transduction genes in the MSTN/Smad signaling pathway, and the effects of those genes on the mutton quality of Tibetan sheep in this study. Six‐month‐old Qinghai‐Tibetan sheep were selected, slaughtered, and their Longissimus lumborum, semitendinosus muscle, arm triceps, and quadriceps femoris muscle were collected. The mutton quality was evaluated, and gene expression and their association with the mutton quality were analyzed using RT‐qPCR. The results showed that the indexes of mutton quality were not significantly different between ewes and rams (p > .05) except for Warner–Bratzler shear force (WBSF) (p < .05). A total of 21 different fatty acids were detected in the muscles of Tibetan sheep, including nine types of SFA, four types of MUFA, and eight types of PUFA. The main transduction genes of the MSTN/Smad signaling pathway were found to be widely expressed in muscle tissues, but no significant differences were observed (p > .05). The correlation analysis of the main genes and mutton quality showed that MSTN was significantly correlated with redness and cooking time; Smad2, Smad3, Smad4, and TGFβRI had significant positive correlations with marbling in arm triceps; Smad3 and TGFβRII had strong negative correlations with pH24 h in Longissimus lumborum; Smad2 was negatively correlated with drip loss in Longissimus lumborum. In short, the expression level of MSTN in muscles was positively correlated with Smad2, Smad3, and Smad4 genes and negatively correlated with TGFβRII genes. Thus, the results of this study provide a theoretical basis for the regulation mechanism of the MSTN/Smad pathway on mutton quality.

繁殖性状是畜禽的重要生产性能之一,直接影响养殖成本与经济效益.与繁殖相关的性状通常受基因调控,其中Smads基因对繁殖性状起着重要的调控作用.Smads基因编码的Smads蛋白是TGF-β超家族中重要的调节因子,也是TGF-β/Smad信号通路中将信号传递至细胞核内的关键信号分子.Smads基因参与调节颗粒细胞的增殖、凋亡等过程,并且调控畜禽卵巢的功能、卵泡的生长发育及激素的合成分泌,从而影响畜禽的繁殖性状.文章介绍了 Smads基因的结构和生物学作用,并综述了 Smads基因对畜禽繁殖性状调控作用的研究进展,以期为进一步研究Smads基因对繁殖性状的调控机制提供参考.

[目的]对藏羊BMP受体11(Bone morphogenetic protein receptor,type Ⅱ,BMPRⅡ)基因进行克隆和生物信息学分析,并检测其在藏羊不同组织中的表达差异,为研究BMPRⅡ基因在藏羊中的生物学功能提供参考.[方法]本试验随机选取年龄和体重相近的6只藏羊进行屠宰,采集各组织样品,提取总RNA并进行质量检测,利用RT-qPCR检测BMPRⅡ基因在藏羊下丘脑、垂体、心脏、肝脏、脾脏、肺脏、肾脏、卵巢、子宫、输卵管、瘤胃、十二指肠和背最长肌等13个组织中的表达量,分析BMPRⅡ基因在藏羊各组织中的特异性表达和分布规律,并对藏羊BMPRⅡ基因编码区序列进行克隆和测序,应用生物学软件对其基因编码产物进行生物信息学分析.[结果]BMPRⅡ基因在上述13个组织中均有表达,在肝脏中的表达量高于子宫、卵巢和脾脏,未达到显著水平(P＞0.05),但显著高于其他组织(P＜0.05),在背最长肌中表达量最低.藏羊BMPRⅡ基因CDS区序列长为3117 bp,编码1038个氨基酸.藏羊BMPRⅡ氨基酸序列与绵羊、牛和牦牛的关系最近,与斑马和鸡的亲缘关系最远.藏羊BMPRⅡ蛋白属于亲水性分泌蛋白,且具有不稳定性,含1个跨膜结构和信号肽,存在127个潜在的磷酸化位点、14个潜在的N-糖基化位点,主要分布在细胞外;藏羊BMPRⅡ蛋白二级结构以无规卷曲为主,其次为α螺旋、延伸链和β转角,三级结构预测结果与二级结构一致.[结论]BMPRⅡ基因在藏羊各项生理活动中发挥着重要作用,本研究结果可为进一步研究BMPRⅡ基因对藏羊繁殖性能调控作用奠定基础.

繁殖性状是衡量绵羊经济价值的重要指标之一,直接影响其养殖效益.我国除了小尾寒羊和湖羊为多胎品种外,大多数地方品种属单胎品种,繁殖率低,且繁殖性状遗传力低、繁殖周期长、选育进展缓慢,制约了绵羊产业的快速发展.近年来,随着生物技术的快速发展和绵羊重要繁殖基因研究的不断深入,分子标记辅助选择育种技术已得到广泛应用,在分子水平上进行早期选择育种,可大大缩短育种年限,加快育种进程.文章综述了褪黑素受体 1A基因(MTNR1A)、绵羊多胎基因(FecB)、骨形态发生蛋白 15 基因(BMP15)、ESR2 基因以及多个分子的调控机制,以期为绵羊育种实践提供参考.

为探究藏羊(Ovis aries)BMPR-ⅠB基因的序列特征、表达特征及其多态性对产羔性状的影响,本研究利用RT-qPCR、克隆测序、生物信息学方法和iMLDR技术等检测藏羊BMPR-ⅠB基因的组织表达特性,克隆了藏羊BMPR-ⅠB基因的编码区,分析了 BMPR-ⅠB理化性质和蛋白质结构,并检测了其在臧羊群体中的多态性.RT-qPCR结果表明,BMPR-ⅠB基因在下丘脑、垂体、心脏、肝脏、脾脏、肺脏、肾脏、卵巢、输卵管、子宫、瘤胃、十二指肠、臂三头肌、半腱肌、股四头肌和背最长肌等16个组织中均有表达,其中在卵巢、输卵管和子宫中的表达量高于其他组织的,在肌肉组织中的表达量最低.生物信息学分析结果表明,藏羊BMPR-ⅠB基因包含1个长为1 509 bp的开放阅读框,编码502个氨基酸,其编码区氨基酸序列与绵羊的关系最近;BMPR-ⅠB蛋白的分子式为C2511H3986N694O747S33,分子量为56 907.45 Da,总原子数为7 971,理论等电点为7.78,T1/2为30 h,不稳定指数为54.07,亲水性均值(grand average of hydropa-thicity,GRAVY)为-0.357;BMPR-ⅠB蛋白含信号肽,1个跨膜区,属于跨膜蛋白;存在87个潜在的磷酸化位点、2个N-糖基化修饰位点;α-螺旋、延伸链、β-转角和无规卷曲所占的比例分别为34.86％、13.15％、3.39％和48.61％,属于混合性蛋白.基因分型结果表明,藏羊BMPR-ⅠB基因编码区FecB位点存在A＞G突变,且存在3种基因型,分别为野生++型、杂合突变B+型和纯合突变BB型,多态信息含量为0.07,属于低度多态,且该位点处于Hardy-Weinberg平衡状态.藏羊平均产羔数在野生++型和杂合突变+型之间差异显著(P＜0.05).本研究结果为进一步研究BMPR-ⅠB基因对藏羊繁殖性能的调控作用提供参考数据.

肌肉生长抑制素(myostain,MSTN)作为肌肉生长抑制因子,属于TGF-β超家族成员,MSTN突变会引起肌肉过度增长,出现"双肌"表型,该基因对于提高动物肌肉产量及瘦肉率有重要意义.为了分析藏羊(Ovis aries)MSTN基因与产肉性状的关联性,本研究随机选取体重相近、健康无病的6只6月龄藏羊,屠宰后测定了藏羊的屠宰性能、肉品质和肉质营养成分;利用荧光定量PCR检测藏羊MSTN基因在臂三头肌、股四头肌、背最长肌和半腱肌中mRNA的表达量,并对MSTN基因表达量与产肉性状进行关联性分析.结果表明:藏羊所有屠宰性能指标在公羊与母羊之间无显著差异,公羊和母羊的屠宰率分别为51.32％和52.01％,胴体重分别为22.08和20.03 kg.肉品质分析结果表明,除了剪切力在公、母羊之间有显著差异(P<0.05)外,其余指标在公、母羊间差异均不显著.肉质营养成分分析结果中,各指标在公、母羊间差异不显著.其中公羊与母羊的蛋白质含量分别为23.14％和22.17％,粗脂肪含量分别为2.46％和2.87％.MSTN基因在藏羊4种肌肉中均有表达,在各组织间表达未达显著差异水平.对MSTN基因的表达量与屠宰性能和肉品质中各指标的相关性分析结果显示,藏羊MSTN基因在臂三头肌中的表达量与宰前活重呈显著正相关(P<0.05,1.000);在背最长肌中的表达量与红度值呈显著正相关(P<0.05,0.831),与熟肉率呈极显著负相关(P<0.01,-0.958);股四头肌中的表达量与熟肉率呈极显著负相关(P<0.01,-1.000);半腱肌中的表达量与各指标相关性均不显著.可见,MSTN基因与藏羊产肉性状存在相关性,本研究可为MSTN基因对藏羊肉品质调控机理的研究提供理论依据.

为研究骨形态发生蛋白受体-ⅠA(bone morphogenetic protein receptorⅠA,BMPR-ⅠA)基因多态性对藏羊(Tibetan sheep)繁殖性能的遗传效应,以403只产单羔藏羊母羊和30只产双羔藏羊母羊为研究对象,采用多重高温连接酶检测反应技术(improved multiple ligase detection reac-tion,iMLDR)对BMPR-ⅠA基因两个SNPs在藏羊群体中的多态性进行检测及其与产羔数进行关联分析.结果 显示:在BMPR-ⅠA基因第8和第9外显子上检测到g.136901 G＞A和g.139032G＞A两个同义突变位点.在g.136901G＞A和g.139032G＞A突变位点上均检测到GG、GA和AA三种基因型,其优势基因型均为GG,优势等位基因均为G;藏羊群体处于低度多态(PIC＜0.25).x2适合性检验结果显示,g.136901G＞A和g.139032G＞A两个突变位点均处于哈迪-温伯格平衡状态.连锁不平衡分析发现,两个位点之间连锁不紧密.关联性分析结果发现,g.136901G＞A和g.139032G＞A两个突变位点与藏羊平均产羔数均无显著相关性(P＞0.05),在g.136901G＞A突变位点上,突变型平均产羔数高于野生型,但差异不显著(P＞0.05).上述结果初步表明,BMPR-ⅠA基因g.136901G＞A突变位点对藏羊产羔数有一定潜在影响.

[目的]研究骨形态发生蛋白受体Ⅱ(bone morphogenetic protein receptorⅡ,BM PRⅡ)基因多态性及其单倍型与藏羊产羔性状的相关性.[方法]以433只藏羊母羊为研究对象,采用改良多重高温连接酶检测反应技术(improved multiple ligase detection reaction,iMLDR)对BMPRⅡ基因9个单核苷酸多态性(single nucleotide polymorphism,SNP)位点在藏羊群体中的多态性进行检测,使用Haploview 4.2软件分析其连锁不平衡性并构建单倍型,应用基因关联分析探究BMPRⅡ基因多态性及单倍型与藏羊产羔性状的关联.[结果]藏羊BMPRⅡ基因外显子12中存在6个SNPs,分别为:g.90192 T＞C、g.142532 C＞T、g.142614 A＞G、g.142751T＞C、g.143138 A＞G和g.143189 G＞A,外显子3、9和13中各存在1个SNP,分别为:g.143570 C＞G、g.124843 A＞C和g.145233 A＞G,这9个SNPs均为同义突变.9个SNPs中,除g.90192 T＞C外,均存在3种基因型.多态信息含量分析显示,群体在g.90192 T＞C、g.142614 A＞G和g.145233 A＞G位点处于低度多态(PIC＜0.25),在g.124843 A＞C、g.142532 C＞T、g.142751T＞C、g.143138 A＞G、g.143189 G＞A和g.143570 C＞G位点均处于中度多态(0.25＜ PIC＜0.5).x2适合性检验结果显示,所有突变位点均未偏离哈代-温伯格平衡状态.相关性分析显示,不同位点不同基因型与藏羊产羔性状均无显著相关(P＞0.05),但g.142532 C＞T位点CT基因型平均产羔数高于CC和TT基因型,g.142751T＞C位点CC基因型平均产羔数高于TT和TC基因型,g.143189 G＞A位点GA基因型平均产羔数高于GG和AA基因型,g.143570 C＞G位点CG基因型平均产羔数高于CC和GG基因型,g.145233 A＞G位点GG基因型平均产羔数高于AA和AG基因型,差异均不显著(P＞0.05).BMPRⅡ基因中除g.90192 T＞C位点外的8个SNPs位点在藏羊群体中共形成14种单倍型(H1～H14),单倍型与藏羊产羔数间均无显著相关(P＞0.05).[结论]BMPRⅡ基因g.142532 C＞T、g.142751T＞C、g.143189 G＞A、g.143570 C＞G和g.145233 A＞G位点对藏羊产羔数有一定潜在的影响.

SMAD1, SMAD2, and SMAD3 are important transcription factors downstream of the TGF-β/SMAD signaling pathway that mediates several physiological processes. In the current study, we used cloning sequencing, RT-qPCR, bioinformatics methods and iMLDR technology to clone the coding region of Tibetan sheep genes, analyze the protein structure and detect the tissue expression characteristics of Tibetan sheep genes, and detect the polymorphisms of 433 Tibetan sheep and analyze their correlation with litter size. The results showed that the ORFs of the SMAD1, SMAD2 and SMAD3 genes were 1398 bp, 1404 bp and 1278 bp, respectively, and encoded 465, 467 and 425 amino acids, respectively. The SMAD1, SMAD2, and SMAD3 proteins were all unstable hydrophilic mixed proteins. SMAD1, SMAD2 and SMAD3 were widely expressed in Tibetan sheep tissues, and all were highly expressed in the uterus, spleen, ovary and lung tissues. Litter sizes of the genotype CC in the SMAD1 gene g.10729C>T locus were significantly higher than that of CT (p < 0.05). In the SMAD3 gene g.21447C>T locus, the genotype TT individuals showed a higher litter size than the CC and CT genotype individuals (p < 0.05). These results preliminarily demonstrated that SMAD1, SMAD2 and SMAD3 were the major candidate genes that affected litter size traits in Tibetan sheep and could be used as a molecular genetic marker for early auxiliary selection for improving reproductive traits during sheep breeding.

SMAD4, SMAD5 and SMAD7 belonging to the transforming growth factor β (TGF-β) superfamily are indispensable for oocyte formation and development, ovarian organogenesis and folliculogenesis. However, only a few studies have investigated the characteristics of SMAD4, SMAD5 and SMAD7 in Tibetan sheep and the effect of their polymorphism on litter size. In this study, we examined the expression of SMAD4, SMAD5 and SMAD7 in 13 tissues of Tibetan sheep by reverse transcription-quantitative polymerase chain reaction. Further, cDNA of these genes was cloned, sequenced and subjected to bioinformatics analysis. DNA sequencing was also used to detect single nucleotide polymorphisms (SNPs). However, iM-LDRTM technology was used for SNP genotyping. Associations between polymorphisms and litter size were analyzed using data from genotyping of 433 Tibetan sheep. The results showed that the expression of SMAD4, SMAD5 and SMAD7 genes was ubiquitous in the tissues of Tibetan sheep, such as the ovary, uterus and oviduct, hypothalamus, hypophysis, heart, liver, spleen, lung, kidney, rumen, duodenum and longissimus dorsi. However, the expression was unbalanced and upregulated in the spleen, lung, ovary and uterus and downregulated in the longissimus dorsi. The bioinformatics analysis showed that SMAD4, SMAD5 and SMAD7 in Tibetan sheep encoded proteins of 533, 465 and 427 amino acids, respectively. Sequence homology analysis of the three proteins among other animals showed that the sequences of SMAD4, SMAD5 and SMAD7 of Tibetan sheep were similar to those in sheep, yak, cattle, dog, human, pig, chimpanzee, rhesus monkey and house mouse. Two synonymous mutations, g.51537A>G and g.319C>T, were detected in SMAD5 and SMAD7, respectively. The associations of these SNPs and litter size were determined, and it was found that both g.51537A>G and g.319C>T have no significant effect on the litter size of Tibetan sheep. The results provided novel insights into the molecular characterization, expression profiles and polymorphisms of SMAD4, SMAD5 and SMAD7 in Tibetan sheep, but our results do not support associations between these genes and the litter size of Tibetan sheep.

为了对藏羊羔羊生长发育过程中体尺性状进行综合评价,本研究采用相关性、聚类及主成分分析,对80只不同月龄阶段(0、2、4、6月龄)的藏羊羔羊体重、体高、体长、胸围、胸深、胸宽和管围7个体尺指标进行分析,并综合评价藏羊羔羊生长发育情况.结果表明:80只藏羊羔羊体尺性状变异系数在0、4、6月龄时均较小,性状稳定;在2月龄时,体重、胸围和胸深3个体尺性状的变异系数均较大,整体选育的潜力较大.相关性分析表明,在不同月龄阶段,藏羊羔羊的体重和体尺性状之间存在着不同程度的正相关,且体重与体高、体长和胸围之间的相关系数较大.聚类分析发现,在藏羊羔羊0、2、4、6月龄,所有体尺性状分为两类,其中体高、体长和胸围聚为一类,体重、管围、胸宽和胸深聚为一类.主成分分析表明,第1主成分贡献较大因子为体重,其次是体长和胸围.本研究结果可为藏羊羔羊的选育提供参考依据.