FST和FMOD基因在绵羊胎儿妊娠中后期的表达特征比较

本研究以骨骼肌表型差异明显的两个绵羊品种特克塞尔羊和乌珠穆沁羊为试验对象,通过对两个与生长发育相关的基因卵泡抑素(follistatin,FST)与纤维蛋白聚糖基因(fibromodulin,FMOD)在不同日龄、不同骨骼肌部位表达的比较分析,探讨绵羊骨骼肌早期发育规律;同时对绵羊胎儿的85、100、120和135日龄4个阶段,在背最长肌、半腱肌、股四头肌、半膜肌和股二头肌5个部位肌肉FST和FMOD基因的表达量进行了比较研究。实时荧光定量PCR结果表明,随着胎儿日龄的增加,FST基因在半腱肌、股四头肌中表达量有增加的趋势,同时在特克塞尔绵羊中的表达量高于乌珠穆沁羊。在100和120日龄半腱肌、120日龄半膜肌、120日龄股二头肌中FST基因表达量在两个品种间差异显著(P<0.05),在100和135日龄半膜肌、85日龄背最长肌、100日龄股四头肌、100日龄股二头肌中FST基因表达量在两个品种间差异极显著(P<0.01)。FMOD基因在股四头肌中表达量较高,而在其他4种肌肉组织中相对表达量较低,且在乌珠穆沁羊中的表达量高于特克塞尔羊中的表达量。FST和FMOD基因在绵羊早期骨骼肌发育中起着重要作用。     

绵羊胚胎发育中后期骨骼肌BTG2/3的表达动态及其与Myostatin的关系

旨在探究绵羊胚胎中后期骨骼肌BTG2/3(B cell translocation gene 2/3)的动态表达情况,探索肌肉生长抑制素(Myostatin,MSTN)基因对其表达的影响。以特克赛尔(Texel)和乌珠穆沁(Ujimqin)绵羊85、100、120和135d4个不同发育阶段的胎儿背最长肌为研究对象,利用Real-time PCR技术分别检测MSTN及BTG2/3的表达差异;选取100d各胚胎半膜肌、背最长肌、半腱肌和股四头肌检测不同组织BTG2/3的表达差异;构建稳定慢病毒干扰载体pFU-GW-myostatin,分别感染处于增殖和分化阶段的绵羊成肌细胞,检测BTG2/3表达水平变化。结果显示,随着胚胎生长,特克赛尔羊胎儿MSTN表达量持续下降,而乌珠穆沁羊胎儿先下降后上升,特克赛尔羊胎儿BTG2表达量呈先升高后降低趋势,而乌珠穆沁羊表现出先降低后升高再降低的趋势;BTG3的表达量在特克赛尔羊与BTG2表达趋势相同,而乌珠穆沁羊则与BTG2相反;100d胎儿4种不同骨骼肌组织中,特克赛尔羊与乌珠穆沁羊相比,背最长肌和半腱肌中BTG2表达量极显著偏高(P0.01),而半膜肌和背最长肌中BTG3表达量极显著偏低(P0.01);慢病毒干扰载体对成肌细胞具有较高的感染和干扰效率,增殖和分化阶段的成肌细胞被MSTN干扰后,BTG2和BTG3表达量极显著降低(P0.01)。综上表明,BTG2和BTG3对绵羊胎儿中后期骨骼肌发育有重要作用,可能参与myostatin调节通路。本研究对绵羊胚胎发育过程的分子机制研究具有重要意义。     

新疆巴什拜羊肌肉生长抑制素基因DNA甲基化与mRNA表达量分析

为了探究肌肉生长抑制素（myostatin,MSTN）基因在肌肉和脂肪中的DNA甲基化模式及mRNA表达水平,试验以5月龄巴什拜羊羔羊为研究对象,采用亚硫酸氢盐测序法（bisulfite sequencing PCR,BSP）检测MSTN基因启动子区和第1外显子甲基化模式,并通过实时荧光定量PCR检测MSTN基因在巴什拜羊股二头肌、股三头肌、半腱肌、半膜肌、背最长肌和尾脂中的mRNA表达水平。结果显示,肌肉组织甲基化概率高于脂肪组织,其中,股二头肌、股三头肌、半腱肌、半膜肌、背最长肌和尾脂的甲基化概率分别为74.2%、74.2%、83.2%、83.7%、82.1%和25.3%,MSTN基因在股二头肌、股三头肌、半腱肌、半膜肌、背最长肌中的表达水平显著低于尾脂（P < 0.05）,股二头肌、股三头肌、半腱肌、半膜肌和背最长肌之间差异不显著（P > 0.05）,巴什拜羊肌肉和脂肪MSTN基因DNA甲基化水平与MSTN基因表达量呈显著负相关（r=－0.886,P < 0.05）。     

影响羊肉质的几个主要因素

1羊的品种 1.1基因效应对绵羊胴体组成和肉质有影响的主基因是Callipyge基因,Callipyge位点已定位于绵羊第18号染色体上,表型具有非孟德尔遗传方式特征.Callipyge绵羊的瘦肉率和屠宰率都比正常绵羊高;Callipyge绵羊肌肉的数量比正常羊要多42%;Callipyge绵羊的背最长肌、股二头肌、半膜肌、半腱肌比正常绵羊分别重32.1%、42.1%、38.3%、26.4%;Callipyge绵羊背最长肌的嫩度比正常羊低,Callipyge Dorset公羊后裔肉的嫩度没有正常羊的高,研究认为,肌原纤维的断裂减少是主要原因.     

Perilipin基因在羊不同肌肉组织中的表达研究

旨在了解脂滴包被蛋白基因(perilipin)在羊不同肌肉组织中的表达特征,为阐明脂滴包被蛋白(perilipin)对脂肪沉积的调控作用提供科学依据。以苏尼特羊、小尾寒羊为研究对象,选取背最长肌、肋间肌、股二头肌、股四头肌为试验材料,通过实时荧光定量PCR(RT-qPCR)研究perilipin基因的表达情况。结果表明,perilipin基因在苏尼特羊和小尾寒羊4种肌肉组织中均有表达;perilipin基因在苏尼特羊与小尾寒羊肌肉组织中的总体表达量没有显著差异;除肋间肌外,苏尼特羊的背最长肌、股二头肌perilipin基因的相对表达量均高于小尾寒羊,股四头肌二者基本一致;苏尼特羊背最长肌相对表达量最高,股四头肌次之,肋间肌相对表达量最小,背最长肌与肋间肌差异显著(P0.05);小尾寒羊肋间肌相对表达量最高,背最长肌和股四头肌表达量基本一致,股二头肌相对表达量最小,肋间肌与股二头肌差异显著(P0.05)。该研究为阐明perilipin基因在脂肪沉积和分解中的重要作用及羊肉肌内脂肪沉积调控机理提供了理论依据。     

Myf5基因在不同时期绵羊胎儿半腱肌和背最长肌中的表达及生物信息学分析

为了研究MRFs家族中Myf5基因在不同发育时期绵羊胎儿骨骼肌中的表达规律及分子特性,实验采取妊娠30 d(E30)、50 d(E50)和70 d(E70)3个时期绵羊胎儿的半腱肌和背最长肌,用HE染色法观察了其显微结构,通过实时荧光定量PCR技术检测了Myf5基因的表达变化,同时运用生物信息学软件对绵羊Myf5蛋白的...     

摘除卵巢对母羊不同组织中MSTN、MYOG基因表达量的影响

试验通过研究卵巢摘除对布尔山羊杂种母羊羔肝脏、背最长肌、股二头肌中肌细胞生长抑制素(MSTN)和肌细胞生成素(MYOG)基因表达量的影响,旨在探讨去卵巢是否可以促进羔羊生长发育。将体重相近、长势良好的5月龄布尔山羊杂种母羊羔,随机分为处理组(n=20)和对照组(n=20)。试验初期摘除处理组母羊羔卵巢,对照组不摘除。饲养60d后,利用实时定量PCR法检测肝脏、背最长肌、股二头肌中MSTN和MYOG基因mRNA相对表达量。结果表明,卵巢摘除后,母羊股二头肌中MSTN基因mRNA相对表达量是对照组的8.73倍,差异极显著(P0.01),但背最长肌和肝脏中MSTN基因mRNA水平低于对照组(P0.05);处理组背最长肌和股二头肌中MYOG基因表达量极显著高于对照组的5.12倍和6.51倍(P0.01),肝脏中MYOG基因表达量为对照组的2.23倍(0.01P0.05)。可见,卵巢摘除可以上调背最长肌、股二头肌和肝脏中MYOG基因的表达以及股二头肌中MSTN基因的表达。     

Texel和乌珠穆沁绵羊妊娠中后期胎儿背最长肌中 AKT基因的发育表达模式

选择绵羊胎儿期骨骼肌中表达稳定性高的内参基因,探讨AKT基因在绵羊妊娠中后期背最长肌中的发育性表达模式。以Texel和乌珠穆沁绵羊为研究对象,采用荧光定量PCR技术检测SDHC、YWHAB、RPLPO、RPS18、B2M 5个内参基因的mRNA表达水平,同时检测AKT基因mRNA丰度在其胎儿期70、85、100、120和135 d间的表达变化,并分析其在品种间和不同生长阶段间的表达差异及其对肌肉生长发育的影响。5个内参基因的定量结果经GeNorm程序统计学分析处理,表达稳定性由高到低排序为RPS18＞RPLPO＞SDHC＞B2M＞YWHAB,确定用RPLPO、RPS18和SDHC 3个基因校正绵羊胎儿期不同生长阶段的目的基因表达量。在5个发育阶段中,两个品种绵羊胎儿AKT基因表达模式有一定差异,AKT表达量在Texel绵羊85和120 d时均有明显的增加,而在乌珠穆沁绵羊的100 d时出现1个表达峰,这与试验对两品种绵羊组织学分析的结果是一致的,进一步印证了Texel绵羊在85和120 d以及乌珠穆沁绵羊在100 d时存在肌发生波的推测。以上结果初步揭示绵羊妊娠中后期生长发育过程中AKT基因表达的发育性变化模式和品种差异,为深入研究AKT基因的作用机制提供了理论基础。     

不同群体肉羊背最长肌中H-FABP mRNA表达的比较研究

心脏型脂肪酸结合蛋白(H-FABP)基因是影响肌间脂肪(IMF)含量进而调控肉品质的的主要候选基因。试验以乌珠穆沁羊以及以南非肉用美利奴羊、东北细毛羊、小尾寒羊、杜泊羊为亲本获得的3种杂交羊(杜寒杂DS组,南寒杂SS组,东南杂NS组)为研究对象,采用实时荧光定量PCR技术测定了背最长肌中H-FABP mRNA的相对表达量,比较该基因在不同群体中的表达差异。结果表明:H-FABP在不同品种肉羊背最长肌中均有表达,但不同群体间差异不显著(P>0.05)。以乌珠穆沁羊H-FABP mRNA表达量为参照,H-FABP mRNA在杜寒杂交羊中的相对表达量约为乌珠穆沁羊的3倍,南寒杂交羊中的相对表达量约为乌珠穆沁羊的0.5倍,东南杂交羊中的相对表达量约为乌珠穆沁羊的1.4倍。H-FABP可能是调控纯种地方绵羊及杂交肉羊肌肉肉质性状的候选基因。     

贵州黑山羊不同肌肉组织FASN基因表达与肌内脂肪含量相关性分析

为探究FASN基因在贵州黑山羊不同肌肉组织中调控脂肪沉积的作用，采集12月龄公、母贵州黑山羊胸肌、背最长肌、肱三头肌、臂股二头肌，运用实时荧光定量PCR法（qPCR）检测FASN基因表达量，酸水解法测定肌内脂肪（IMF）含量，并通过Pearson分析FASN基因表达量与IMF含量相关性。结果显示，FASN基因在公、母羊不同组织中表达量均为臂股二头肌>胸肌>肱三头肌>背最长肌，其中公羊臂股二头肌显著高于背最长肌，母羊臂股二头肌极显著高于背最长肌；相同组织比较，母羊胸肌、肱三头肌、臂股二头肌表达量均高于公羊，背最长肌表达量低于公羊，但均未达到差异显著水平。IMF含量测定发现，公、母羊4个组织中IMF含量均为臂股二头肌>胸肌>肱三头肌≥背最长肌，同性别各组织之间比较差异不显著；相同组织比较，母羊4个组织中IMF含量均高于公羊。FASN基因表达量与IMF含量相关性分析表明，公、母羊FASN基因表达量与胸肌IMF含量呈正相关，与背最长肌、肱三头肌、臂股二头肌IMF含量呈负相关。     

The Expression Comparison Analysis of FST and FMOD Genes in Sheep Fetus during Mid and Late Pregnancy

In order to investigate skeletal muscle development mechanism,the expression of follistatin (FST) and fibromodulin (FMOD) gene were comparatively analyzed in fetus different skeletal muscles and different stages between Ujumqin sheep and Texel sheep during early stage.Two gene expression were analyzed including longissimusdorsi,semimembranosus,semitendinosus quadricepsfemoris and biceps femoris muscle were compared by variance analysis methods of four stages (85,100,120 and 135 d).The results of quantitative Real-time PCR showed that the expression of FST gene showed a tendency to increase with the addition of stage,and the expression of FST gene in Texel sheep was higher than that in Ujumqin sheep in semitendinosus (100,120 d),semimembranosus (120 d),and biceps femoris (120 d) (P<0.05).The expression of FST gene in Texel sheep was extremly higher than that of Ujumqin sheep in semitendinosus (100,135 d),longissimusdorsi (85 d),quadricepsfemoris (100 d) and biceps femoris (100 d) (P<0.01).As a whole,the expression of FMOD gene was the highest in quadriceps femoris,and was lower in other four muscles,it's expression in Ujumqin sheep was higher than that in Texel sheep.It was inferred that FST and FMOD genes might be related to muscle development during early stage.     

Analysis of DNA Methylation and mRNA Expression Level of MSTN Gene in Xinjiang Bashiby Sheep

In order to investigate DNA methylation and expression levels of myostatin (MSTN) gene in mscule and fat, 5 months of age of Bashiby sheep were selected, the promoter region and exon 1 methylation levels of MSTN gene was analyzed using bisulfite sequencing PCR (BSP). Real-time PCR was used to detect the expression level of MSTN gene in biceps femoris, femoral triceps, semitendinosus, semimembranosus, longissimus dorsi muscle and tail fat of Bashiby sheep. The results showed that the methylation probability of muscle was higher than fat, the methylation probability of biceps femoris, femoral triceps, semitendinosus, semimembranosus, longissimus dorsi muscle and tail fat were 74.2%, 74.2%, 83.2%, 83.7%, 82.1% and 25.3%, respectively. The expression levels of MSTN gene in biceps femoris, femoral triceps, semitendinosus, semimembranosus, longissimus dorsi muscle were significantly lower than tail fat in Bashiby sheep (P < 0.05), but there were no significant difference among biceps femoris, femoral triceps, semitendinosus, semimembranosus and longissimus dorsi muscle (P > 0.05).The DNA methylation was negatively correlated with the expression levels in muscle and fat of Bashiby sheep (r=-0.886, P < 0.05).     

藏羊MSTN基因克隆、生物信息学及组织表达分析

【目的】 对藏羊肌肉生长抑制素（myostatin，MSTN）基因进行克隆和生物信息学分析，检测其在藏羊不同组织中的表达，为探究MSTN基因在藏羊中的生物学功能提供参考。【方法】 以藏羊背最长肌组织cDNA为模板，克隆藏羊MSTN基因完整CDS区序列并测序，用SeqMan程序对测序结果进行拼接，并用BLAST在线程序对组装后的序列进行分析鉴定。用生物信息学软件进行相似性比对、系统进化树构建及生物信息学分析，用实时荧光定量PCR检测MSTN基因在藏羊不同组织中的表达量。【结果】 藏羊MSTN基因CDS全长为1 128 bp，编码375个氨基酸。藏羊MSTN基因氨基酸序列与绵羊、牦牛、牛、猪、恒河猴、人、黑猩猩、犬、鸡及斑马的相似性依次为100.0%、93.4%、93.4%、95.2%、94.4%、94.2%、94.4%、93.1%、87.8%和87.5%；系统进化树分析结果表明，藏羊与绵羊的亲缘关系最近，与斑马和鸡的亲缘关系最远。藏羊MSTN蛋白属于亲水性分泌蛋白，且具有不稳定性，不含跨膜结构，含1个信号肽，存在31个潜在的磷酸化位点、2个N-糖基化修饰位点，主要分布在线粒体和细胞质中；MSTN蛋白二级结构以无规卷曲为主，其次是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。实时荧光定量PCR结果显示，MSTN基因在藏羊不同组织中均有表达，其中在臂三头肌和半腱肌的表达量显著高于肺脏、下丘脑、心脏、肝脏、十二指肠、瘤胃和肾脏（P<0.05）。【结论】 成功克隆了藏羊MSTN基因；该基因在藏羊肌肉中的表达量高于内脏组织。该结果为进一步研究MSTN基因对藏羊肌肉生长发育的调控机制奠定了基础。     

Variation in proteolysis, sarcomere length, collagen content, and tenderness among major pork muscles

The objectives of this experiment were to determine the extent of variation in proteolysis, sarcomere length, and collagen content among pork muscles and the association of those factors with tenderness variation among muscles at 1 d postmortem. Twenty-three white composite barrows were slaughtered and carcasses (66 kg) were chilled at 0 degrees C for 24 h. At 1 d postmortem, the longissimus lumborum, biceps femoris, semimembranosus, semitendinosus, and triceps brachii, long head were dissected from one side of each carcass and frozen. Trained sensory panelists evaluated tenderness, amount of connective tissue, juiciness, and pork flavor intensity of grilled (70 degrees C) chops on 8-point scales. Raw chops were used for total collagen content, sarcomere length, and the extent of desmin proteolysis. Tenderness ratings were highest (P < .05) for semitendinosus (7.2) and triceps brachii (7.1), followed by longissimus lumborum (6.4) and semimembranosus (5.7) and were lowest (P < .05) for biceps femorus (4.0). The simple correlations between longissimus lumborum tenderness and the tenderness of other muscles were .54 (semimembranosus), .34 (semitendinosus), .36 (triceps branchii), and .17 (biceps femorus). Total collagen was highest (P < .05) for biceps femorus (7.1 mg/g muscle), followed by triceps branchii (6.0 mg/g) and semitendinosus (5.3 mg/g), and lowest for semimembranosus (4.5 mg/g) and longissimus lumborum (4.1 mg/g). Sarcomere length was longest (P < .05) for semitendinosus (2.5 microm) and triceps branchii (2.4 microm), followed by semimembranosus (1.8 microm), longissimus lumborum (1.8 microm), and biceps femorus (1.7 microm). Proteolysis of desmin was greatest (P < .05) in longissimus lumborum (39.3%), followed by semimembranosus (21.0%) and biceps femoris (18.5%), then semitendinosus (.2%) and triceps brachii (.2%). Multiple linear regression using total collagen, sarcomere length, and proteolysis accounted for 57% of the variation in tenderness rating among all samples. Piecewise linear regression was used to account for the interaction of sarcomere length with proteolysis and collagen. This analysis accounted for 72% of the variation in tenderness rating. Variation in collagen, proteolysis, and sarcomere length and the degree of their interaction with one another determine the tenderness of individual muscles.     

Influence of zeranol implants and electrical stimulation on the palatability traits of five muscles in Angus and Limousin bulls and steers

A split-plot design was used to study effects of male status (bull vs steer), zeranol implants, breed types (Angus vs Limousin) and electrical stimulation on palatability traits of five major muscles when all animals were fed to a similar fat-thickness endpoint. Warner-Bratzler shear (WBS) determinations and taste-panel evaluations were performed on the longissimus, adductor, semimembranosus, semitendinosus and biceps femoris muscles removed 24 h postmortem. Breed was the most consistent main effect difference, with Angus cattle having lower WBS values than Limousin for the longissimus and semitendinosus muscles. Angus cattle also were favored in many sensory traits. Few significant differences were noted between bulls and steers in taste-panel traits. Differences due to electrical stimulation were most evident in the longissimus muscle, with higher panel-evaluation scores, an enhancement of marbling and lean color and a lower incidence of heat ring. The implant tended to have negative effects on palatability in this trial. An overall ranking of the five muscles based on WBS from most tender to least tender was: adductor, semimembranosus, longissimus, semitendinosus and biceps femoris. Correlations of palatability traits of one muscle to other muscles within an experimental unit were found to be low.     

Increasing tenderness of beef round and sirloin muscles through prerigor skeletal separations

Crossbred steers (n = 30) were used to explore and compare tenderness improvements in beef round and sirloin muscles resulting from various methods of prerigor skeletal separations. Animals were slaughtered according to industry procedures, and at 60 min postmortem one of six treatments was applied to each side: A) control, B) saw pelvis at the sirloin-round junction, C) separate the pelvic-femur joint, D) saw femur at mid-point, E) combination of B and C, and F) combination of B and D. After 48 h, the following muscles were excised from each side: semimembranosus, biceps femoris, semitendinosus, and adductor from the round; vastus lateralis and rectus femoris from the knuckle; and gluteus medius, biceps femoris and psoas major from the sirloin. Following a 10-d aging period, samples were removed from each muscle to determine the effect of treatment on sarcomere length and Warner-Bratzler shear force. Most skeletal separation treatments resulted in longer sarcomeres than controls for semimembranosus, adductor, semitendinosus, and gluteus medius muscles. All skeletal separation treatments yielded shorter sarcomeres for the psoas major as compared with controls. Warner-Bratzler shear force differed among treatments for rectus femoris, semitendinosus, and psoas major. For rectus femoris, treatments C, D, E, and F resulted in lower (P < 0.05) shear values than for controls. Treatments B, D, and F increased shear force of the semitendinosus relative to controls (P < 0.05) within muscle. Treatment F resulted in higher shear force values for the PM than controls (P < 0.05). Correlations between sarcomere length and shear force were found to be low and quite variable among muscles. In general, treatments increased sarcomere length of several muscles from the sirloin/round region, but had mixed effects on shear force values.     

Effects of postmortem aging and USDA quality grade on Warner-Bratzler shear force values of seventeen individual beef muscles

Forty USDA Select and 40 upper two-thirds USDA Choice beef carcasses were used to determine the effects of postmortem aging on tenderness of 17 individual beef muscles. Biceps femoris-long head, complexus, gluteus medius, infraspinatus, longissimus dorsi, psoas major, rectus femoris, semimembranosus, semitendinosus, serratus ventralis, spinalis dorsi, supraspinatus, tensor fasciae latae, teres major, triceps brachii-long head, vastus lateralis, and vastus medialis muscles were removed from each carcass. Seven steaks (2.54-cm thick) were cut from every muscle, and each steak was assigned to one of the following postmortem aging periods: 2, 4, 6, 10, 14, 21, or 28 d postmortem. After completion of the designated aging period, steaks were removed from storage (2 degrees C, never frozen), cooked to a peak internal temperature of 71 degrees C, and evaluated using Warner-Bratzler shear force (WBSF). Analysis of WBSF revealed a 3-way interaction (P = 0.004) among individual muscle, USDA quality grade, and postmortem aging period. With the exception of the Select teres major, WBSF of all muscles (both quality grades) decreased with increasing time of postmortem storage. Nonlinear regression was used to characterize the extent (aging response) and rate of decrease in WBSF from 2 through 28 d postmortem for each muscle within each quality grade. In general, WBSF of upper two-thirds Choice muscles decreased more rapidly from 2 to 10 d postmortem than did corresponding Select muscles. Muscles that had greater aging responses generally had greater 2-d WBSF values. The upper two-thirds Choice psoas major, serratus ventralis, and vastus lateralis muscles required similar aging times to complete a majority of the aging response (< or =0.1 kg of aging response remaining) compared with analogous Select muscles. The upper two-thirds Choice complexus, gluteus medius, semitendinosus, triceps brachii-long head, and vastus medialis muscles required 4 to 6 d less time to complete a majority of the aging response than did comparable Select muscles. Aging times for Select biceps femoris-long head, infraspinatus, longissimus dorsi, rectus femoris, semimembranosus, spinalis dorsi, supraspinatus, and tensor fasciae latae muscles were > or =7 d longer than those for corresponding upper two-thirds Choice muscles. Results from this study suggest that muscle-to-muscle tenderness differences depend on quality grade and aging time and that postmortem aging should be managed with respect to individual muscle and USDA quality grade.