甘肃肉羊新品种选育群GDF9基因外显子2多态性及其与产羔性状关联分析

根据GenBank发布的绵羊GDF9基因外显子2的序列设计4对引物,采用PCR-SSCP技术分析GDF9基因外显子2在甘肃内羊新品种选育群羊中的单核苷酸多态性,并与产羔性状进行关联分析.结果表明,GDF9基因的扩增片段在所检测的新品种群羊中存在PCR-SSCP多态性,检测到3种基因型(AA、AB和BB),而在32只无角陶赛特母羊群中只检测到AA和AB基因型.测序结果显示,GDF9基因编码区第978位碱基发生A→G突变,但没有导致氨基酸的改变;第994位碱基发生G→A突变,导致Ⅴ变成Ⅰ(缬氨酸→异亮氨酸).新品种选育群羊产羔数的最小二乘均值关系为AB＞ AA＞ BB,统计分析结果初步表明3种基因型之间差异不显著(P＞0.05).故该区域可能不是影响新品种群羊繁殖力的功能结构区城.     

五指山猪GH、IGFBP3基因SNPs检测及与生长性状的关联分析

以五指山猪为试验材料,采用PCR-SSCP和测序相结合技术,对GH基因和IGFBP3基因进行多态性检测,并分析其与生长性状(体质量,体高,体长和胸围)的相关性。结果表明,GH基因第2外显子存在一处G→A转换,属沉默突变,该位点与生长性状关联分析显示,BB基因型的体重显著高于AA基因型和AB基因型;IGFBP3基因第2外显子存在一处G→C颠换,属沉默突变,该位点与生长性状关联分析显示,AA基因型的体高显著高于BB基因型;IGFBP3基因第4内含子存在一处碱基C缺失,该位点与生长性状关联分析显示,AA基因型体重显著高于AB基因型和BB基因型。研究将为五指山猪生长发育规律、系统选育及矮小机制等方面研究提供了遗传学依据。     

FSHβ 基因PCR-SSCP多态性及其与济宁青山羊高繁殖力关系的研究

采用PCR-SSCP技术检测促卵泡素b亚基(follicle-stimulating hormone β, FSHβ)基因5′调控区、外显子1和外显子2在高繁殖力山羊品种(济宁青山羊)和低繁殖力山羊品种(辽宁绒山羊、波尔山羊、安哥拉山羊)中的单核苷酸多态性, 同时研究该基因对济宁青山羊高繁殖力的影响。结果表明: 山羊与绵羊的FSHβ 基因该段核苷酸序列同源性为98%。9对引物中, 只有P9的扩增片段存在多态性。P9的扩增片段在济宁青山羊和辽宁绒山羊中检测到AA、AB和AC 3种基因型; 在波尔山羊中检测到AA、CC和AC 3种基因型; 在安哥拉山羊中检测到AA、BB、CC、AB、AC和BC共6种基因型。测序分析发现BB型与AA型相比在外显子2的第94 bp处有G→A突变, 并引起氨基酸改变(丙氨酸→苏氨酸); CC型与AA型相比在外显子2的第174 bp有一处C→T沉默突变。济宁青山羊AA、AB和AC基因型频率分别为0.686、0.137和0.177。AA基因型济宁青山羊产羔数最小二乘均值比AB基因型的多0.78只(P<0.05), 比AC基因型的多0.64只(P<0.05)。     

牛生长激素受体基因多态性与体尺、体重指标的相关性分析

利用PCR-SSCP技术对南阳牛、利木赞、盖洛威共100头牛的生长激素受体基因的外显子10(GHR10)部分序列进行单核苷酸多态性研究, 并分析了该基因的不同基因型与3个品种牛生产性状的关系。结果表明: 南阳牛、利木赞、盖洛威3个品种中共存在6种基因型(AA、BB、CC、AB、AC、BC), c2检验表明该实验群体在这一位点上处于Hardy-Weinberg平衡状态(P > 0.05), 利木赞和盖洛威的PIC表现为中度多态,而南阳牛表现高度多态。测序结果显示: 所扩增GHR10部分片段共有5处碱基突变, 分别是495 bp (A/T), 622 bp(C/T), 650 bp(A/C), 702 bp(T/C), 730 bp(A/G); 并导致3处氨基酸替代: 其中622 bp Pro/Ser(脯氨酸/丝氨酸), 650 bp Asn/Thr(天冬酰胺/苏氨酸), 730 bp Ser/Gly(丝氨酸/甘氨酸)。最小二乘分析表明: 基因型AB、BC所对应的12月龄体重最小二乘均值显著高于基因型CC所对应的最小二乘均值(P < 0.05, P < 0.01); 18月龄体重BC基因型显著高于CC基因型(P < 0.01), 基因型AB所对应的18月龄胸围最小二乘均值显著高于基因型CC所对应的最小二乘均值(P < 0.05)。     

三个绵羊群体MC4R基因多态性及生物信息学分析

旨在探讨绵羊黑素皮质素受体-4(melanocortin-4 receptor,MC4R)的分子机理,采用PCR-SSCP方法对3个绵羊群体(甘肃肉用绵羊新品种群、小尾寒羊和湖羊)的MC4R基因外显子进行多态性检测和生物信息学分析。结果表明,3个绵羊群体均存在3种基因型AA型、AB型和BB型,优势基因型为BB,其中优势等位基因为B;测序结果表明,野生型BB型和突变型AB型相比,AB型个体在该基因编码区第511位点发生G→A突变,第495位发生C→T突变;AA型个体在该基因编码区第511位点发生G→A突变,出现AA的纯合,第495位发生C→T突变,出现CC纯合;3个绵羊群体中小尾寒羊的多态信息含量属于中度多态(0.25PIC0.50),甘肃肉用绵羊新品种群羊和湖羊属于低度多态(PIC0.25);χ2适合性检验表明除湖羊之外,其余2个绵羊品种均处于Hardy-Weinberg平衡状态。生物信息学分析发现MC4R氨基酸序列有明显的疏水性区域,有7个跨膜螺旋区及信号肽,其编码蛋白主要的二级结构元件是α螺旋和无规则卷曲;同源性比对发现绵羊MC4R基因与山羊、牛、野猪、人类及大猩猩的相似度分别为97%、94%、81%、83%及83%,说明MC4R是一个非常保守的蛋白,在绵羊的生长发育中起着重要作用。     

甘肃河西地区西门塔尔杂交类群NGB基因第3外显子的遗传变异研究

旨在对甘肃河西的临泽、甘州、武威、金昌、高台5个地区283头西门塔尔杂交类群NGB基因第3外显子的遗传多态性及变异特征进行系统分析,采用PCR-SSCP方法检测了283头西门塔尔杂交类群NGB基因第3外显子和部分内含子的多态性,且对群体内各等位基因进行了测序。结果显示,5个地区西门塔尔杂交类群共检测出5个等位基因(A、B、C、D、E),表现为5种基因型(AA、AB、AC、AD、AE)。其中甘州、武威、金昌西门塔尔杂交类群NGB基因均只检测到AA、AB 2种基因型,高台西门塔尔杂交类群检测到AA、AE 2种基因型,临泽西门塔尔杂交类群检测到AA、AB、AC、AD 4种基因型。A等位基因和AA基因型的频率在5个群体中最高,为优势基因和优势基因型。对不同SSCP带型的对应片段进行测序分析,共发现6个核苷酸突变位点(75 bp C→T,78 bp C→G,128 bp G→A,214 bp G→A,232 bp C→T,233 bp G→A),其中第75 bp和第78 bp处的突变位点位于内含子区域,其余4处突变位点均位于外显子区域。第214 bp处的核苷酸突变导致甘氨酸(Gly)突变为丝氨酸(Ser),第232 bp处核苷酸突变导致精氨酸(Arg)突变为色氨酸(Trp),第233 bp处核苷酸突变导致精氨酸(Arg)突变为谷氨酰胺(Gln),经χ2检验结果显示,5个地区的西门塔尔杂交类群在此3个突变位点上都处于Hardy-Weinberg平衡状态(P0.05)。群体遗传学分析结果表明,临泽、甘州、武威、金昌、高台西门塔尔杂交类群的多态信息含量(PIC)分别为0.0582、0.0196、0.0196、0.0161、0.0159,均属于低度多态(PIC0.25)。     

GHR基因第10外显子与塞北兔生长发育性能相关性研究

本研究利用单链构象多态性(SSCP)和测序技术,对塞北兔群体GHR基因第10外显子进行多态性研究,并与生长发育性能进行关联性分析。结果发现:有两处点突变,214处为同义突变,均编码苯丙氨酸;AB基因型在255处出现A位点的缺失,引起移码突变;塞北兔整个群体呈现哈代-温伯格平衡状态,但母兔群体不平衡;AA基因型比AB基因型对塞北兔活体重,胴体重,前腿重均有提高,差异显著(p0.05)。表明:母兔群体在育种过程中受到人工选择而引起遗传漂变,AA基因型为有利基因型,因此可作为影响塞北兔生长性状的主效候选基因,进行早期辅助选择。     

水貂GH基因SNP_S与皮张长度的相关性研究

以水貂生长激素(GH)基因作为控制水貂皮张长度性状主基因的候选基因,以大兴安岭水貂养殖基地养殖的水貂群为试验材料,通过PCR-SSCP方法对GH基因进行多态性检测。在该基因内含子1中发现1处碱基突变:C→A,并检测到3种基因型(AA、AB、BB),BB基因型个体与AA基因型个体皮张长度有一定的差异(P0.05)。在外显子2中发现2处碱基突变:T→A、C→G,并由此检测到了3种基因型,分别命名CC、CD、DD,但3种基因型对水貂皮长的影响没有显著的差异(P0.05)。统计各基因型之间的组合给水貂皮长带来的影响时,发现多数组合基因型对所检测的水貂皮长有显著影响(P0.05)。     

奶牛β-Lg基因第1外显子PCR-SSCP多态性与泌乳性能的相关性

采用PCR-SSCP技术并结合测序对233头奶牛β乳球蛋白(β-Lg)基因5’端部分序列和外显子1全部序列进行了多态性研究,分析了该基因与奶牛泌乳性状的相关性。结果表明:β-Lg基因5’端和外显子1共存在2个等位基因3种基因型,BB型为优势基因型,B为优势等位基因。该群体在这一位点上偏离Hardy-Weinberg平衡状态,多态信息含量(PIC)为0.3548。测序结果显示,与普通牛该基因序列(X14710)相比,B等位基因在2073 bp、2202 bp和2206 bp处发生了G→C、C→T和A→G的碱基突变,其中2202 bp处的C→T突变导致第11位氨基酸由苏氨酸变为异亮氨酸,而A等位基因在3个位点上与X14710相同。最小二乘法分析表明,BB型305 d乳蛋白量显著高于AA型和AB型(P<0.05);AB型305 d乳脂量显著高于AA型(P<0.05),BB型与AB型之间差异不显著(P>0.05);等位基因B为高乳蛋白量和乳脂量的优势基因,可作为奶牛选育的分子遗传标记。     

中国美利奴羊H-FABP基因多态性及其与部分肉质性状相关性的研究

利用聚合酶链式反应-单链构象多态（PCR-SSCP）技术检测中国美利奴羊(Chinese Merino)心脏型脂肪酸结合蛋白基因（H-FABP）外显子2的单核苷酸多态性（SNPs）和遗传多态性,分析其与肌内脂肪（IMF）含量、肌纤维直径和肌纤维密度的相互关系,为该品种绵羊的分子标记辅助选择提供理论依据。结果显示,H-FABP基因外显子2有AA、AB和BB 3种基因型,AA型和BB型在778位均发生了C缺失,939位均发生了A→G转换,BB型还在789位发生了T→C转换,该突变导致所编码氨基酸发生了缬氨酸→丙氨酸的替换;BB型为IMF的优势基因型,与AB型相比差异显著(P<0.05),与AA型相比差异极显著（P<0.01);BB型对肌纤维直径存在负相关。结果提示,中国美利奴羊H-FABP基因外显子2具有多态性,该基因可能是中国美利奴羊肉质性状的主效基因,或者与控制这些性状的主效基因相连锁。     

SNP variation in ADRB3 gene reflects the breed difference of sheep populations

The β3-adrenergic receptor (ADRB3), a G-protein coupled receptor, plays a major role in energy metabolism and regulation of lipolysis and homeostasis. We detect the single nucleotide polymorphism (SNP) variation in full-length sequence of ovine ADRB3 gene in 12 domestic sheep populations within four types by polymerase chain reaction-single strand conformation polymorphism and sequencing to reveal the breed difference. Twenty-two SNPs, 12 of which in the exon 1 and ten in the intron, were detected, and 12 new exonic and four new intronic SNPs were found. Most SNPs presented in Shanxi Dam Line and least ones in Dorset. The average SNP number in both meat and dual purpose for meat and wool breeds was significantly higher than general and dual purpose breeds for wool and meat. Frequency of each SNP in studied breeds or types was different. The 18C Del and 1617T Ins majorly existed in dual purpose breeds for wool and meat. The 25A Del, 119C>G and 130C>T were mostly found in the meat and dual purpose for meat and wool breeds. The 1764C>A more frequently presented in meat than in other types. The majority of variations came from within the populations as suggested by analysis of molecular variance. Close relationship presented among the Chinese and western breeds, respectively. In conclusion, SNPs of ovine ADRB3 gene can reflect the breed difference and within- and between-population variations, and to a great extent, the breed relationship.     

BMP-15基因外显子Ⅰ在三种绵羊中的多态性及与繁殖力相关性分析

为研究绵羊繁殖力与绵羊BMP-15 (bone morphogenetic protein 15)基因的多态位点关系,并寻找调控绵羊繁殖力的分子标记,以甘肃高山细毛羊、蒙古羊、小尾寒羊三种绵羊为研究对象,采用PCR-SSCP技术与DNA碱基测序相结合的方法,检测该基因第一外显子67、92两个位点在上述三个不同品种绵羊中是否存在FecX^H (Q23→Ter)和FecX^I (V31→D)突变,同时根据检测结果与其繁殖力做相关性分析。结果表明,在三种绵羊中既未检测出与Inverdale绵羊相同的FecX^I突变,也未检测出与Romney绵羊相同的FecX^H突变,因此推测BMP-15基因中影响Romney与Inverdale绵羊高繁殖力的突变位点对以上三种绵羊均无显著影响。     

云南乌骨绵羊乌质性状与TYR基因多态性的相关分析

乌骨乌肉是乌骨绵羊的主要乌质性状。本文测定了乌骨绵羊、兰坪本地羊和罗姆尼羊血液TYR活性并分析了乌骨绵羊与乌骨鸡组织器官黑色素结构,结果表明：乌骨绵羊TYR活性显著高于兰坪本地绵羊和罗姆尼羊（P<0.05）;乌骨绵羊黑色素与乌骨鸡黑色素的IR光谱基本一致,黑色物质主要是真黑色素。首次克隆了绵羊TYR基因第一外显子长667 bp序列,并检测了乌骨绵羊和非乌骨绵羊TYR基因多态性。结果发现,检测的乌骨绵羊TYR基因第一外显子有两个变异位点,分别位于第64和154号编码氨基酸上,但都属于同义突变。通过对64号位置设计酶切位点检测TYR基因多态性,结果发现,该突变与乌质性状有关基因紧密连锁,TYR基因上可能存在乌质性状相关功能突变位点。另外,TYR基因多态性与毛色的表型相关极显著（P<0.01）,该基因可能也是毛色功能基因。     

In a shake of a lamb's tail: using genomics to unravel a cause of chondrodysplasia in Texel sheep

Chondrodysplasia in Texel sheep is a recessively inherited disorder characterized by dwarfism and angular deformities of the forelimbs. A genome-wide association study using the Illumina OvineSNP50 BeadChip on 15 sheep diagnosed as affected and eight carriers descended from three affected rams was conducted to uncover the genetic cause. A homozygous region of 25 consecutive single nucleotide polymorphism (SNP) loci was identified in all affected sheep, covering a region of 1 Mbp on ovine chromosome 4. Seven positional candidate genes - including the solute carrier family 13 (sodium/sulphate symporters), member 1 (SLC13A1) - were identified and used to search for new SNPs for fine mapping of the causal locus. The SLC13A1 gene, encoding a sodium/sulphate transporter, was the primary candidate gene attributable to similar phenotypes observed in the Slc13a1 knockout mouse model. We discovered a 1-bp deletion of T (g.25513delT) at the 107 bp position of exon 3 in the SLC13A1 gene. Genotyping by direct sequencing and restriction fragment length polymorphism analysis for this mutation showed that all 15 affected sheep were g.25513delT/g.25513delT; the eight carriers were g.25513delT/T and 54 normal controls were T/T. The mutation g.25513delT shifts the open reading frame of SLC13A1 to introduce a stop codon and truncate C-terminal amino acids. It was concluded that the g.25513delT mutation in the SLC13A1 gene was responsible for the chondrodysplasia seen in these Texel sheep. This knowledge can be used to identify carriers with the defective g.[25513delT] allele to avoid at-risk matings to improve animal welfare and decrease economic losses.     

GDF9 as a candidate gene for prolificacy of Small Tail Han sheep

Growth differentiation factor 9 (GDF9) which controls the fecundity of Belclare, Cambridge, Santa Ines, Moghani, Ghezel and Thoka ewes was studied as a candidate gene for the prolificacy of Small Tail Han sheep. According to the sequence of ovine GDF9 gene, six pairs of primers were designed to detect single nucleotide polymorphisms of two exons of GDF9 gene in both high fecundity breed (Small Tail Han sheep) and low fecundity breed (Dorset sheep) by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP). Only the products amplified by primers 2-1 and 2-2 displayed polymorphisms. For primer 2-1, three genotypes (AA, AB and BB) were detected in both sheep breeds. Sequencing revealed one silent mutation (G477A) in exon 2 of GDF9 gene in the BB genotype in comparison with the AA, which was known as G3 mutation of GDF9 gene in Belclare and Cambridge ewes. The relationship of least squares means for litter size was AA?>?AB?>?BB in Small Tail Han sheep (P?>?0.05). For primer 2-2, two genotypes (CC and CD) were detected in both sheep breeds. Sequencing revealed one novel single nucleotide mutation (G729T) in exon 2 of GDF9 gene in the CD genotype in comparison with the CC, which resulted in an amino acid change (Gln243His). The ewes with mutation heterozygous genotype CD had 0.77 (P?相似文献   

Identification of Single Nucleotide Polymorphisms in the Nicastrese Goat and Sardinia Sheep Mannose-Binding Lectin

This study was undertaken to detect polymorphisms in the goat and sheep mannose-binding lectin encoding gene (MBL2) and to explore allelic variability of this gene in these two species. The analysis and comparison of the sequences obtained from sheep showed 13 polymorphic sites, six in the promoter and seven in exon 1, four of which were of the missense type. In the goats, 12 polymorphic sites were detected, five intronic, five in the promoter, and one exonic. The exon site was responsible for an amino acid change. Mutations detected at the MBL2 locus in the sheep are of particular interest, being potentially responsible for the alterations of gene expression. A population survey involved 102 ewes of the Sardinian breed and 218 goats of the Nicastrese breed, all reared in southern Italy.     

Molecular evolution of coding and non-coding sequences of the growth hormone receptor (GHR) gene in the family Bovidae

The GHR gene exon 1A and exon 4 with fragments of its flanking introns were sequenced in twelve Bovidae species and the obtained sequences were aligned and analysed by the ClustalW method. In coding exon 4 only three interspecies differences were found, one of which had an effect on the amino-acid sequence--leucine 152 proline. The average mutation frequency in non-coding exon 1A was 10.5 per 100 bp, and was 4.6-fold higher than that in coding exon 4 (2.3 per 100 bp). The mutation frequency in intron sequences was similar to that in non-coding exon 1A (8.9 vs 10.5/100 bp). For non-coding exon 1A, the mutation levels were lower within than between the subfamilies Bovinae and Caprinae. Exon 4 was 100% identical within the genera Ovis, Capra, Bison, and Bos and 97.7% identical for Ovis moschatus, Ammotragus lervia and Bovinae species. The identity level of non-coding exon 1A of the GHR gene was 93.8% between species belonging to Bovinae and Caprinae. The average mutation rate was 0.2222/100 bp/MY and 0.0513/100 bp/MY for the Bovidae GHR gene exons 1A and 4, respectively. Thus, the GHR gene is well conserved in the Bovidae family. Also, in this study some novel intraspecies polymorphisms were found for cattle and sheep.