猪肌肉组织Col3a1基因表达的发育性变化及其对肌内胶原特性的影响(英文)

以30~90kg体重莱芜猪和40~100kg体重鲁莱黒猪共84头去势公猪为试验对象(每组6头),采用相对定量RT-PCR方法,以β-actin作为内标,研究肌肉中编码Ⅲ型胶原的Col3a1基因表达的发育性变化及其对肌肉中胶原蛋白含量和性质(溶解度)的影响。结果表明:研究的两个品种猪肌肉中Col3a1基因表达的发育性变化基本一致,即随体重的增加,肌肉中Col3a1 mRNA表达呈逐渐增加趋势,但莱芜猪和鲁莱黑猪分别在70kg和80kg体重组表达量略有下降。总体上莱芜猪肌肉组织Col3a1 mRNA表达丰度显著高于鲁莱黑猪(P<0.05)。相关分析表明,莱芜猪肌肉组织Col3a1 mRNA表达的发育性变化与总胶原和不溶性胶原含量呈极显著正相关(P<0.01),与胶原溶解度呈极显著负相关(P<0.01)。鲁莱黑猪肌肉组织Col3a1 mRNA表达的发育性变化与不溶性胶原和胶原溶解度分别呈显著正相关和负相关(P<0.05)。研究结果提示:猪肌肉组织中Col3a1基因表达具有明显的体重发育和品种特征,其mRNA表达对于肌内胶原的含量和性质有重要影响。     

猪肌肉组织Col3a1基因表达的发育性变化及其对肌内胶原特性的影响

以30—90妇体重莱芜猪和40—100kg体重鲁莱黑猪共84头去势公猪为试验对象（每组6头）,采用相对定量RT-PCR方法,以β-actin作为内标,研究肌肉中编码Ⅲ型胶原的Col3al基因表达的发育性变化及其对肌肉中胶原蛋白含量和性质（溶解度）的影响。结果表明：研究的两个品种猪肌肉中Col3al基因表达的发育性变化基本一致,即随体重的增加,肌肉中Col3al mRNA表达呈逐渐增加趋势,但莱芜猪和鲁莱黑猪分别在70妇和80妇体重组表达量略有下降。总体上莱芜猪肌肉组织Col3al mRNA表达丰度显著高于鲁莱黑猪（P〈0．05）。相关分析表明,莱芜猪肌肉组织Col3al mRNA表达的发育性变化与总胶原和不溶性胶原含量呈极显著正相关（P〈0．01）,与胶原溶解度呈极显著负相关（P〈0．01）。鲁莱黑猪肌肉组织Col3al mRNA表达的发育性变化与不溶性胶原和胶原溶解度分别呈显著正相关和负相关妒〈0．05）。研究结果提示：猪肌肉组织中Col3al基因表达具有明显的体重发育和品种特征,其mRNA表达对于肌内胶原的含量和性质有重要影响。     

藏鸡IRX3基因表达与肌内脂肪的沉积关系

旨在构建藏鸡IRX3基因的组织及时序表达谱,确定该基因与肌内脂肪的沉积关系。利用荧光定量PCR技术检测IRX3基因在不同组织和不同发育阶段肌肉组织中的表达水平,同时采用单因素方差分析方法分析肌内脂肪含量,并分析其时序表达与肌内脂肪含量的相关性。结果表明,IRX3基因在藏鸡各个组织中均有表达,在藏鸡肺组织中相对表达水平较高,极显著高于其他各个组织(P0.01)。IRX3 mRNA在210日龄公藏鸡腿肌和胸肌中相对表达水平均极显著高于其他日龄的表达水平(P0.01),而在1日龄母藏鸡胸肌中相对表达水平极显著高于其他日龄(P0.01)。藏鸡不同发育阶段肌肉组织中IRX3 mRNA表达水平与其IMF含量的相关性分析显示,公藏鸡腿肌IRX3 mRNA表达量与肌内脂肪含量呈正相关(P0.01),而在母藏鸡胸肌IRX3 mRNA表达量与肌内脂肪含量呈负相关(P0.01)。IRX3基因表达与藏鸡肌内脂肪含量存在相关性,推测IRX3可能对肌内脂肪的沉积具有调控作用。     

猪Musclin基因的组织表达特征及其与生脂基因的相关性分析

根据人、大鼠、小鼠Musclin基因序列设计引物,从大白猪肌肉组织提取总mRNA, R T-PCR扩增猪Musclin cDNA;利用半定量(Semi Quantitative, SQ) RT-PCR检测猪Musclin mRNA在不同组织、以及不同生长阶段和不同品种猪脂肪与肌肉组织中的表达量,研究不同品种猪皮下脂肪组织Musclin基因与脂肪酸合成酶(FAS)、过氧化物酶体增殖物激活受体γ (PP ARγ)、甘油三酯水解酶(TGH)基因mRNA表达量的相关性.结果表明:猪Musclin基因片段与其他物种的同源性达59%以上, 该基因在内脏脂肪和皮下脂肪表达丰度最高,脾脏表达量最低(P<0.05);随月龄增长猪肌肉和皮下脂肪组织中Musclin mRNA表达量呈显著下降( P<0.05) ;瘦肉型猪肌肉和皮下组织Musclin mRNA表达显著高于脂肪型猪(P<0.05);皮下脂肪组织中该基因mRNA表达与FAS、PPARγ分别呈显著负相关和正相关(P<0.05),与TGH相关性不显著 (P>0.05).因此,猪Musclin基因除在肌肉中表达外,在脂肪等其他组织也能表达 ;该基因的表达与猪生长阶段、品种有关,且与生脂基因FAS、PPARγ密切相关.推测该基因可能在脂代谢中起到一定的作用 [动物学报 54(3):453-459,2008].     

睾丸注射法制备携带猪PID1基因的转基因兔

目的研究磷酸酪氨酸互作结构域1(PID1)基因与肌内脂肪含量的关系,探究睾丸注射法在转基因动物制备中的可行性。方法将携带猪PID1基因的重组质粒pIRES2-acGFP-PID1与转染试剂共孵育后,对新西兰兔进行了睾丸打点注射试验。对繁殖的F1代个体进行了活体荧光检测、PCR和western blotting检测,以及抽样屠宰进行肌内脂肪含量等检测;将F1代阳性个体互交,繁殖了F2代兔,对其进行了阳性率检测以及肌内脂肪含量检测。结果外源PID1基因和荧光蛋白基因在后代中均成功表达,其中,F1代阳性率为35.88%,F2代阳性率为34.33%;转基因阳性兔与阴性和空白对照兔相比,PID1蛋白表达水平有所增加,肌内脂肪含量有显著提高(P0.05)。结论 PID1基因与肌内脂肪沉积密切相关,同时,进一步证明了睾丸注射法可以用于制备转基因动物,且外源基因可以稳定遗传。     

藏猪和大约克猪MYL1基因多态性及差异表达分析

为研究猪不同品种的MYL1基因的多态性及mRNA表达差异.本试验以藏猪和大约克猪作为实验动物,采用一代测序技术对藏猪(59头)和大约克猪(58头)MYL1基因起始密码子上游3 kb区域进行多态性检测;利用实时荧光定量PCR技术检测MYL1基因在背最长肌、背脂、肝脏组织差异表达情况.结果表明:藏猪和大约克猪在起始密码子上游3 kb区域发现T-159A突变位点并呈极显著差异(P＜0.01);MYL1基因在背最长肌表达量最高,其次是背脂,肝脏的最低,在背最长肌中藏猪MYL1基因表达量极显著高于大约克猪(P＜0.01),在背脂和肝脏组织中,藏猪MYL1基因表达量显著高于大约克猪(P＜0.05).推测T-159A位点可能是参与肌肉生长发育的重要调控位点.本研究为下一步开展MYL1基因对猪生长性能及其肉品质的影响提供参考依据.     

绵羊肌肉中FAS基因和HSL基因的发育性变化及其对肌内脂肪含量的影响

选取不同日龄的雄性哈萨克羊和新疆细毛羊共54只,屠宰后取背最长肌,用索氏抽提法检测肌内脂肪(intramuscular fat,IMF)含量,用荧光实时定量PCR法检测肌肉脂肪酸合成酶(fatty acid synthase,FAS)和激素敏感脂肪酶(hormone-sensitive lipase,HSL)基因表达的发育性变化,并分析基因表达对肌内脂肪沉积的影响。结果表明:1)随着日龄的增加,雄性哈萨克羊的IMF含量持续上升,各生长时期差异显著(P<0.05),而新疆细毛羊的IMF含量在各生长时期无显著差异(P>0.05)。雄性哈萨克羊的IMF含量30~90日龄期间极显著高于新疆细毛羊(P<0.01)。2)FAS基因mRNA水平在哈萨克羊肌肉中初生时最高(P<0.05),然后随日龄的增加呈下降趋势;在新疆细毛羊肌肉中,FAS mRNA水平表现出"下降-上升-下降-上升"的发育模式,其中60日龄显著高于90日龄(P<0.05),其余日龄之间差异不显著。HSL基因在2品种绵羊肌肉中的表达模式基本类似,在哈萨克羊肌肉中随年龄的增加而下降,初生时的水平显著高于60~90日龄(P<0.05);在新疆细毛羊中30日龄时达到最高(P<0.01),到60日龄时下降到最低(P<0.05),随后保持这种低表达水平。3)FAS和HSL基因mRNA的表达量均与哈萨克羊IMF含量呈负相关,相关系数分别为:r=-0.485(P=0.02),r=-0.423(P=0.05);在哈萨克羊中两基因表达量水平比值(FAS:HSL)与IMF呈极显著负相关r=-0.552(P=0.01)。在新疆细毛羊中两基因的表达水平及比值均与IMF无显著相关性(P>0.05)。     

ADRP基因在宗地花猪及其杂交猪不同组织中的表达分析

为研究ADRP基因在宗地花猪及其二元杂交猪不同组织中的表达规律,探讨该基因与脂肪含量的关系,试验以宗地花猪及其二元杂交猪为材料,采用实时荧光定量PCR技术,检测ADRP基因在心脏、肝脏、脾脏、肺脏、肾脏、小肠、背最长肌及脂肪中的相对表达量。结果表明,ADRP基因在宗地花猪及其杂交猪各组织中均有表达,不同组织之间及不同猪种之间的表达存在差异,表达丰度关系宗地花猪的为小肠脂肪肺脏肝脏心脏背最长肌脾脏肾脏,宗江二元猪的为脂肪小肠脾脏肺脏肾脏肝脏背最长肌心脏;显著性检验结果显示,宗江二元猪肝脏、脾脏、肺、小肠、背最长肌中ADRP基因的相对表达量显著高于宗地花猪的(p0.01),心脏、肾脏和脂肪组织中的表达量显著低于宗地花猪的(p0.01)。结果提示,ADRP基因在宗地花猪及其二元杂交猪不同组织中的差异表达可能与脂肪沉积有关。     

莱芜猪和杜洛克猪肌肉肌球蛋白重链组成对肉质性状的影响

为探讨莱芜猪肌肉肌球蛋白重链(MyHC)慢速氧化型(Ⅰ)、快速氧化型(Ⅱa)、中间型(Ⅱx)、快速酵解型(Ⅱb)组成与肉质性状间的关系, 阐释莱芜猪肉质优良的机理. 本实验在评价肉质性状的基础上, 用荧光定量RT-PCR方法检测莱芜猪和杜洛克猪背最长肌和半膜肌MyHC 4种亚型mRNA的表达量, 并进行相关分析. 结果表明, 莱芜猪背最长肌和半膜肌MyHCⅡa, Ⅱx mRNA表达量显著高于杜洛克猪, MyHC Ⅱb mRNA表达量则显著低于杜洛克猪, 而MyHCⅠmRNA表达量无明显变化. MyHCⅠ, Ⅱa, Ⅱx mRNA表达量与肉色、pH、大理石纹、肌内脂肪含量正相关, 与嫩度剪切值和肌纤维直径负相关; MyHC Ⅱb mRNA的表达量与大理石纹和肌内脂肪含量显著负相关(P<0.05), 与嫩度剪切值极显著正相关(P<0.01), 与肌纤维直径正相关(P>0.05). 由此可见, MyHCⅠ, Ⅱa, Ⅱx, Ⅱb的组成影响肉质性状, 而且其mRNA的表达量与肉质性状, 尤其是与肌肉食用品质显著相关, 也就是说MyHC 4种亚型的mRNA表达量可准确、客观评价肉质.     

不同猪品种肌肉组织FoxO1与MyoD基因mRNA的表达及其相关性分析

分别取6月龄八眉猪、长白猪和长×八杂交猪背最长肌,提取总RNA,设计并合成猪FoxO1和MyoD 引物,以β肌动蛋白作为内参,优化反应条件和体系,利用RT-PCR方法检测八眉、长白和长×八杂交猪肌肉组织中FoxO1和MyoD基因mRNA 的表达.结果表明,在不同经济类型猪品种肌肉组织中,FoxO1和MyoD基因mRNA的表达丰度均存在显著差异,FoxO1在八眉猪肌肉组织中的表达普遍高于长白猪（P<0.01）,在杂交组合长×八肌肉组织中的表达也高于长白猪（P<0.01）;而MyoD恰好相反,即在长白猪肌肉组织中的表达普遍高于八眉猪（P<0.01）,在杂交组合长×八肌肉组织中的表达也高于八眉猪（P<0.01）．结果提示,FoxO1和MyoD基因mRNA的表达在肌肉发育中存在负相关（r = 0.728 , P < 0.05）,FoxO1在肌肉组织中的上调作用,可能是造成的MyoD基因mRNA表达降低的原因之一,进而负调控肌肉的发育和骨骼肌的量．     

Developmental changes and effect on intramuscular fat content of H-FABP and A-FABP mRNA expression in pigs

H-FABP and A-FABP genes are considered as candidates for intramuscular fat (IMF) accretion. The aim of the present study was to assess the expression of H-FABP and A-FABP genes in m. longissimus dorsi (LD) and liver tissues of Laiwu and Lulai Black pig populations of different body weight (BW). Eighty-four barrows at different BW (30, 40, 50, 60, 70, 80, 90 and 100 kg, n?=?6 per group) of Laiwu Black pig (no 100 kg group) and Lulai Black pig (no 30 kg group) were used to study the development changes of A-FABP and H-FABP mRNA expression and their relationships to IMF content. The results showed that, in both breeds, the IMF content increased continuously with growing (P?<?0.05). The expression of H-FABP and A-FABP genes also increased with growing in LD tissue (P?<?0.05), and reached a peak at 50 and 70 kg BW in Laiwu and Lulai Black pig, respectively. However, this regularity was not observed in liver tissue in both breeds. A positive correlation was just found between the A-FABP mRNA expression level in LD tissue and IMF content and BW in both breeds (P?<?0.05). In conclusion, the A-FABP gene is strongly related to the development and function of IMF accretion in pigs.     

Effect of immunological stress to neuroendocrine and gene expression in different swine breeds

Immunological stress is the status of animal in active immune when they are challenged by bacterial, virus and endocrine. It is associated with immunological, neurological, and endocrinological response. An immunological stress model was established in this study using Chinese indigenous breed (Laiwu), crossbred (Lulai), and exotic breed (Yorkshire), to explore the capacity of immunological stress resistance among different breeds. The study was also to reveal the effect of chromium yeast to immunological stress. 48 post-weaning piglets were taken from three breeds, 16 piglets of each breed from Laiwu, Lulai and Yorkshire. The experiment was designed as 2 × 2 factors, immunological stress (Saline, LPS) and Chromium (with Cr, without Cr). There were four treatments: control, LPS, Cr, and Cr+LPS. Blood parameters related to immunological stress, such as IL-1β, TNF-α, GH, and cortisol, were examined after blood sample were taken at 0, 2, 5, and 7 h of post-injection. The results showed that IL-1β, TNF-α, and cortisol increased in group of LPS treatment while GH declined at 2 h of post-injection in comparison to the control (p < 0.01). However, IL-1β, TNF-α, and cortisol in group of Cr+LPS were lower than that in group of LPS while GH were higher (p < 0.05). Total RNA was extractedfrom blood lymphocytes separation samples at 2 h of post-injection. Q-PCR was applied to determine the gene expression of IL-1β, IL-6 and TNF-α. The results showed that LPS injection increased the gene expression of IL-1β, IL-6 and TNF-α. Among three breeds, the expression of IL-1β, IL-6 and TNF-α in Yorkshire were significantly higher than in Laiwu and Lulai (p < 0.05), but there was no difference between Laiwu and Lulai. Among four treatments, the expression of three genes in group of LPS was the highest, compared to the group of Cr+LPS (p < 0.05) and control (p < 0.01). This study concluded that Laiwu had stronger capacity of immunological stress resistance and next was Lulai among three breeds. Chromium yeast helped piglets relieve immunological stress.     

Single nucleotide polymorphism scanning and expression of the FRZB gene in pig populations

Secreted frizzled-related protein 3 (sFRP3), encoded by the gene FRZB, is a member of the sFRP family with important roles in inhibition of the Wnt signalling pathway through competitive binding of the Wnt receptor. Here, we investigated pig FRZB as a candidate gene for growth traits and identified three polymorphic sites, an insertion (A-532B) and two SNPs (G636A and C650T) in its 5′-UTR. The genotype distributions of G636A and C650T were significantly different among mini-type indigenous (Diannan Small-ear and Tibetan), normal indigenous (Laiwu and Huai), and introduced (Large Yorkshire and Landrace) breeds. In semi-quantitative PCR expression analysis, expression of FRZB mRNA was abundant in tissues of hypophysis, longissimus dorsi muscle, and adipose tissues, and low in the heart, hypothalamus, and brain. Quantitative determination of mRNA level and protein expression analysis were corresponding. The results demonstrated that FRZB gene expression in longissimus dorsi muscle and liver tissue was significantly higher in Diannan Small-ear and Tibetan pigs than in the Large Yorkshire breed (P < 0.05); however, in back fat tissue, the expression was significantly higher in Diannan Small-ear pig than in Tibetan or Large Yorkshire breeds (P < 0.05). Given the known growth and fat characteristics of the breeds, these results indicate that FRZB expression has a negative association with muscle growth and a positive association with fat deposition. In conclusion, FRZB may be a major candidate gene for growth traits in pigs.     

含不同比例莱芜猪血缘杂交猪胴体品质及 肉质特性的研究

摘 要： 以莱芜猪、3/4莱芜猪、1/2莱芜猪、1/4莱芜猪和大约克夏猪（共60头）为研究对象,在同样条件下饲养至90kg屠宰,研究比较不同比例莱芜猪血缘对肥育猪胴体品质和肉质特性的影响。结果表明：不同血缘结构的试验猪间在胴体重、胴体长、后腿比例和背膘厚方面存在显著的差异（P<0.05）;眼肌面积和瘦肉率方面则存在极显著的差异（P<0.01）,并且是随莱芜猪血缘含量的减少,其胴体长、后腿比例、眼肌面积和瘦肉率逐渐增高,而其背膘厚逐渐降低。在肉质特性方面,不同血缘结构的试验猪间肌肉的大理石纹、肉色、干物质和粗蛋白含量存在显著的差异（P<0.05）;而肌肉失水率、滴水损失及肌内脂肪含量存在极显著的差异（P<0.01）。在同样体重下,莱芜猪及其杂交猪与大约克夏猪相比,其肌肉具有鲜红的肉色、良好的持水性能和较丰富的肌内脂肪。研究结果提示：为兼顾产肉性能与肉质特性而进行的优质肉猪生产,莱芜猪有其独特的利用价值,其适宜的血缘比例应控制在1/4左右。Abstract: Sixty pigs, including pure Laiwu Black (LL), pure Large Yorkshire (YY), 1/2 Laiwu (Y♂×L♀), 3/4 Laiwu (L♂×YL♀) and 1/4 Laiwu (Y♂×YL♀), were housed in groups with the same diet until 90kg for slaughter. The objectives were to investigate the effects of graded proportions of Laiwu Black genes on carcass and meat quality performance. Results indicated that different consanguinity had significant effect on carcass weight, carcass length (CL), ham percentage (HP) and backfat thickness (BT) (P<0.05), and had highly significant effect on eye muscle area (EMA) and lean percentage (LP) (P<0.01). Furthermore, CL, HP, EMA and LP tended to increased gradually, but BT tended to decreased gradually as Laiwu Black genes decreased. For meat quality properties, different consanguinity had significant effect on meat color, marbling score, dry matter and crude protein content of muscles (P<0.05), and had highly significant effect on water loss, drip loss and intramuscular fat content of muscles (P<0.01). Compared with the muscle of Large Yorkshire, that of Laiwu Black and its crossbred pigs were bright red in meat color, high in water holding capacity and abundant in intramuscular fat. Results implied that to give attention to both quantity and quality, Laiwu Black surely has usable precious value in superior pig production. Commercial crossbred pigs should have about 1/4 of Laiwu Black genes.     

Molecular characterization and expression analysis of caveolin-1 in pig tissues

Members of the caveolin family played important roles during fundamental cellular processes,such as regulation of cell morphology,migration,and gene expression in muscle cells.In this study,caveolin-1 (Cav-1),one of the caveolins,was identified from longissimus dorsi muscle of Large Yorkshire pig and Chinese indigenous Lantang pig based on the results of mRNA differential display analysis.The deduced amino acids sequence of the porcine Cav-1 contained a caveolin domain,and was very conservative among different species.The Cav-1 mRNA was widely expressed in the eight tissues in this study,including heart,liver,kidney,encephalon,spleen,lung,longissimus dorsi muscle,and back fat, and the highest expression quantity was found in back fat of the two pig breeds.The expression quantity of porcine Cav-1 in back fat and longissimus dorsi muscle of Lantang pig was significantly higher than that of Large Yorkshire(P<0.01,and P<0.05,respectively).These results suggested that the Cav-1 might be a candidate gene for carcass traits,and might provide valuable information for understanding the mechanism of caveolae signaling in fat deposition by using the animal model of pig.