中外十个猪种H-FABP基因遗传变异的研究

利用PCR-RFLP技术（限制性内切酶采用HinfI,HaeⅢ和Msp I）检测了杜洛克、长白、大约克、南昌白、二花脸、梅山、玉山黑猪、乐平花猪、金华两头乌及上高两头乌10个猪种共计561头猪在心肌脂肪酸结合蛋白（H-FABP）基因位点的遗传变异。结果表明：（1）在HinfI-RFLP位点上,南昌白猪均为HH纯合子,而其他猪种在此位点上均存在变异,外来品种以纯合子HH居多。（2）在HaeⅢ-RFLP位点上,外来猪种杜、长、大及培育品种南昌白猪均在该位点表现为多态,但中国6个地方猪种在该位点均无变异;（3）在Msp I-RFLP位点上,仅有杜洛克在该位点表现为多态。而所有检测的地方猪种均表现为AADD-型。     

MyoG基因对金华猪繁殖性状的影响

用PCR-RFLP方法对金华猪I系（109头）、II系（15头）、III系（25头）的MyoG基因的第二内含子内（PCR1- MspⅠ-RFLP）和3'-端（PCR2- MspⅠ-RFLP）位点进行了多态性分析。结果表明：PCR1- MspⅠ-RFLP位点有3种基因型（AA、AB、BB）,其基因型频率分别为0.1544,0.3826,0.4631;PCR2- MspⅠ-RFLP位点只有2种基因型（MM、MN）,其基因型频率分别为0.9953和0.0047。采用SPSS程序分析MyoG基因对金华猪繁殖性状的影响。结果表明：MyoG基因对金华母猪初胎的总产仔数、二胎的产活仔数影响显著（P<0.05）,而对其他胎次组合的总产仔数、产活仔数以及初生窝重没有显著影响（P<0.05）。     

不同猪种E.coli F18受体基因的多态性

采用PCR—RFLP技术检测了大约克、长白、杜洛克、宁乡、沙子岭和大围子6个品种共867头猪的E.coli F18受体(ECF18R)基因座的遗传变异。结果表明：Hin6 Ⅰ-RFLP位点上,大约克、长白、杜洛克3个外来猪种均存在多态,且以敏感型(GG型和AG型)居多,平均占94％,3个外来猪种的G等位基因频率平均为0．76,AA抗性型个体占少数,平均为6％,猪群中M307处G→A的突变频率并不高。宁乡、沙子岭和大围子3个本地猪种的所有检测样品都表现为GG型,在该位点上均不存在G→A的突变。各猪种ECF18受体基因座的PCR—RFLP基因型分布X^2检验结果表明,每个外来猪种ECF18受体基因座的PCR—RFLP基因型分布与3个本地猪种的相比均差异显著或极显著,3个本地猪种间的ECF18受体基因座的PCR—RFLP基因型分布完全一致。外来猪种间只有长白与杜洛克各基因型的分布差异显著,其余均不显著。     

猪核糖体蛋白基因RPLP0的cDNA全长、染色体定位和多态性分析

RPLPO基因编码酸性核糖体磷蛋白大亚基P0,是核糖体60S亚基的组成成分之一。从本室构建的猪胚胎骨骼肌cDNA文库中分离得到猪RPLPO基因的全长cDNA,并提交GenBank数据库。比较猪RPLPO基因和人及小鼠同源基因的cDNA序列和蛋白质序列,结果表明该基因在3个物种中具有高的相似性。用PCR．RFLP方法在猪RPLPO基因cDNA545处检测到C—A的单碱基突变,为Csp6Ⅰ的酶切位点。统计分析结果表明3种基因型（AA,AC,CC）在外来品种杜洛克,大约克,长白和中国地方品种通城猪,小梅山,玉山猪中的分布各不相同。同时使用体细胞杂种板（SCHP）和辐射杂种板（IMpRH）对RPLPO基因进行染色体定位,该基因被定位于SSC14q22．q24并且和SW1321微卫星标志紧密连锁（25cR,LOD=14．54）。     

山西猪种及其杂种群体H-FABP基因的PCR-RFLP研究

利用PCR－RFLP技术对马身猪、山西白猪及其杂种群体共286头猪的心脏脂肪酸结合蛋白（H-FABP）基因5′－上游区（HinfⅠ-RFLP）和第二内含子内（HinfⅠ*-RFLP和Hae Ⅲ-RFLP）的遗传变异进行了研究。结果表明：（1）在Hae Ⅲ-RFLP位点上,马身猪均为DD纯合子,而其他猪群在此位点上均存在变异,马身猪的杂种群体在该位点上只有两种基因型（DD、Dd）;（2）在5′－上游区的HinfⅠ-RFLP位点上,杜洛克猪×山西白猪的杂种群体只有HH基因型,而其他群体都表现出多态性,马身猪等位基因h的频率为0.9727;（3）在第二内含子内的HinfⅠ*-RFLP位点上,马身猪表现出两种基因型（BB、Bb）,等位基因B的频率为0.9667;（4）在HinfⅠ*-RFLP和Hae Ⅲ-RFLP位点上,所有猪群均处于Hardy –Weinberg 平衡状态。     

西部地区主要猪种H-FABP基因多态性,IMF含量及不同基因型脂肪细胞脂滴量的关系

H-FABP是FABP家族成员之一,在长链脂肪酸的吸收和代谢平衡中发挥关键作用,但它对猪IMF含量的影响还知之甚少,对中国西部地区的猪种更是如此.文章利用PCR-RFLP（HinfⅠ、HaeⅢ和Msp Ⅰ 3种限制性内切酶）分子标记技术,检测了杜洛克猪、长白猪、大白猪、内江猪、荣昌猪、汉江黑猪、汉中白猪、八眉猪和野猪共计265头猪H-FABP基因5＇-上游区和第二内含子区的遗传变异,利用最小二乘模型分析了H-FABP基因对猪肌内脂肪含量的遗传效应,并运用猪脂肪细胞培养,油红O染色和TG测定等技术检测了H-FABP基因不同基因型脂肪细胞内脂滴的形态和沉积的量.结果表明：（1）在HinfⅠ-RFLP位点上,上述品种和野猪均存在多态性,其中大白猪、八眉猪、汉江黑猪、汉中白猪和野猪表现为低度多态,杜洛克、长白猪、内江猪和荣昌猪为中度多态;除汉江黑猪（P＜0.05）和野猪（P＜0.01）外,其他猪种基因频率和基因型频率都处于Hardy-Weinderg平衡状态（P＞0.05）;而在HaeⅢ-RFLP和Msp Ⅰ-RFLP位点上,仅内江猪、荣昌猪、汉江黑猪和八眉猪为单态;（2）9种基因型对肌内脂肪（IMF）含量的影响,HH＞Hh＞hh,DD＜Dd＜dd,AA＜Aa＜aa,遗传效应值分别为：3.89,3.42,3.17,2.27,2.49,2.91,2.28,2.70,2.95,H-FABP基因可显著地提高IMF含量（P＜0.05）;（3）aaddHH型的脂肪细胞脂滴的形态,密度和含量与其他基因型细胞差异显著（P＜0.05）.结果提示：可通过提高＂aaddHH＂基因型的频率来增加IMF含量,达到改善猪肉质的目的.     

西部地区主要猪种和野猪H-FABP基因分子标记

利用PCR-RFLP(Hinf I、Hae Ⅲ和Msp I 3种限制性内切酶)分子标记技术,检测了杜洛克猪、长白猪、大白猪、内江猪、荣昌猪、汉江黑猪、汉中白猪、八眉猪和野猪共计265头猪H-FABP基因5′-上游区和第二内含子区的遗传变异,并利用最小二乘模型分析了H-FABP基因对猪肌内脂肪含量的遗传效应。结果表明：（1）在Hinf I-RFLP位点上,上述品种和野猪均存在多态性,其中大白猪、八眉猪、汉江黑猪、汉中白猪和野猪表现为低度多态,杜洛克、长白猪、内江猪和荣昌猪为中度多态;除汉江黑猪（P<0.05）和野猪（P<0.01）外,其他猪种基因频率和基因型频率都处于Hardy-Weinderg平衡状态（P>0.05）;而在Hae Ⅲ-RFLP和Msp I-RFLP位点上,仅内江猪、荣昌猪、汉江黑猪和八眉猪为单态;（2）9种基因型对肌内脂肪（IMF）含量的影响,HH>Hh>hh,DD相似文献   

猪肌细胞生成素基因对部分生长性状的影响

采用PCR-SSCP方法对长白猪、大白猪、杜洛克猪、山西黑猪和马身猪的肌细胞生成素（Myogenin,简称MyoG）基因进行单核苷酸多态性检测,并分析MyoG基因对猪的初生重、断奶重、6月龄重和背膘厚的影响。根据猪MyoG基因的DNA序列设计10对引物,发现In2-3引物扩增的片段有多态性。统计结果发现,3种基因型（AA、AB、BB）在各品种中的分布不一致。χ^2独立性检验表明,基因型频率在外来猪种（长白猪、大白猪、杜洛克猪）与地方猪种（山西黑猪、马身猪）间存在显著差异（P〈0．05）。固定效应模型分析结果表明,初生重和背膘厚基因型间差异显著（P〈0．05）,而断奶重和6月龄重基因型间差异不显著（P〉0．05）。最小二乘分析结果表明,BB基因型与其他两种基因型比较有较小的初生重,同AA和AB型比较差异极显著（P〈0．01）,3种基因型在初生重的大小排列顺序为AA〉AB〉BB;AA基因型与其他两种基因型比较有较小的背膘厚,同AB和BB型比较差异极显著（P〈0．01）,3种基因型在背膘厚的大小排列顺序为AA〈AB〈BB。因此,推测基因型对个体的初生重和胴体瘦肉率存在一定的影响,选择带有A等位基因的个体有望提高个体的初生重和胴体的瘦肉率。     

猪核糖体蛋白基因RPLP0的cDNA全长、染色体定位和多态性分析

RPLP0基因编码酸性核糖体磷蛋白大亚基P0, 是核糖体60S亚基的组成成分之一。从本室构建的猪胚胎骨骼肌cDNA文库中分离得到猪RPLP0基因的全长cDNA,并提交 GenBank 数据库。比较猪 RPLP0 基因和人及小鼠同源基因的cDNA序列和蛋白质序列,结果表明该基因在 3 个物种中具有高的相似性。用 PCR-RFLP 方法在猪 RPLP0基因cDNA 545处检测到 C→A 的单碱基突变,为 Csp6Ⅰ的酶切位点。统计分析结果表明 3 种基因型 (AA,AC,CC)在外来品种杜洛克,大约克, 长白和中国地方品种通城猪,小梅山,玉山猪中的分布各不相同。同时使用体细胞杂种板(SCHP)和辐射杂种板(IMpRH) 对 RPLP0 基因进行染色体定位,该基因被定位于 SSC 14q22-q24 并且和SW1321微卫星标志紧密连锁 (25cR, LOD = 14.54)。     

猪MyoG基因3''''端PCR-SSCP遗传多态性及其遗传效应

采用PCR—SSCP方法对长白猪、大白猪、杜洛克猪、山西黑猪和马身猪共636头猪的肌细胞生成素（Myogenin,简称MyoG）基因3’端的遗传多态性进行检测,分析MyoG基因对猪的初生体质量、断奶体质量、6月龄体质量和背膘厚的影响。根据已发表的猪MyoG基因3’端侧翼序列设计3对引物,发现F1／R1引物对扩增的片段有多态性。统计结果发现：长白、大白、杜洛克猪种B基因为优势基因,其基因频率分别为0．8807、0．7256和0．8581：山西黑猪种4基因为优势基因,其基因频率为0．9359;马身猪种只检测到4基因。χ^2独立性检验表明,基因型分布在外来猪种（长白猪、大白猪、杜洛克猪）与地方猪种（山西黑猪、马身猪）间存在极显著差异（P〈0．01）。固定效应模型分析结果表明,初生体质量基因型闻差异显著（P〈0．05）,而断奶体质量、6月龄体质量和背膘厚基因型闻差异不显著（P〉0．05）。最小二乘分析结果表明,BB基因型与其它2种基因型比较有较大的初生质量,同似和AB型比较差异极显著（P〈0．01）。因此,推测MyoG基因对个体的初生体质量存在一足的影响,选择带有B等位基因的个体有望提高个体的初生体质量。     

猪Mx1基因第14外显子多态性分析及新突变位点的 发现

采用PCR-RFLP方法对国内外7个猪种Mx1基因第14外显子的多态性进行分析, 共检测到3个等位基因, 6种基因型。其中杜洛克中仅存在AA基因型, 苏太猪中存在全部基因型, 只有在梅山猪和具有梅山猪血统的苏太猪中出现基因型BB。所有猪种中, 只有在地方猪种和培育猪种中出现等位基因B, 所有猪种除松辽黑猪外均以A为优势等位基因。卡方检验结果表明, 不同猪种间基因型分布差异较大, 梅山猪和松辽黑猪与其他所有猪种的基因型频率差异极显著(P＜0.01) , 苏太猪与除皮特兰猪外的所有猪种的基因型频率差异也极显著(P＜0.01) , 淮猪与杜洛克和约克夏这两个国外猪种基因型频率差异不显著(P＞0.05), 而与皮特兰和其他地方猪种的基因型频率均存在极显著差异(P＜0.01) 。通过测序在扩增片段中新发现了3种类型的碱基突变, 前2个分别导致了Thr和Glu向Ala和Arg的替换, 最后一个突变不引起氨基酸的变化, 且后两个突变位点为BB基因型所特有。     

猪ATF4基因多态性与生产性状的关联及基因表达分析

为进一步了解和认识ATF4基因的功能,揭示ATF4对猪脂肪代谢的影响,寻找与肉质性状相关联的分子标记,文章采用PCR方法扩增了ATF4基因部分序列,通过序列比对发现在翻译起始密码子ATG下游159 bp处存在A159G转换,通过PCR-AluⅠ-RFLP对大白猪、长白猪、梅山猪和通城猪进行酶切分型,发现在大白猪和长白猪中均为AA基因型,在梅山猪和通城猪中均为GG基因型。进一步对大白猪×梅山F2群体资源家系进行了酶切分型,并分析该位点的多态性与生产性状的关系。结果表明,ATF4的多态性与臀部平均膘厚存在极显著相关(P<0.01),与胸腰椎间膘厚、平均膘厚、眼肌高、眼肌面积存在显著相关(P<0.05)。采用Real-time PCR分析了ATF4基因在大白猪与梅山猪背最长肌不同发育阶段的表达模式。结果表明,ATF4基因在大白猪和梅山猪胚胎期65 d和出生后3 d中的表达水平相对都比较低,且在两品种间无明显差异;而在出生后60 d和120 d,ATF4基因在大白猪中与梅山猪均出现了上调表达,并且在梅山猪中的相对表达水平要显著高于大白猪。研究结果为进一步深入研究猪ATF4基因在脂肪代谢中的分子机理奠定了基础。     

Bi-PASA genotyping of a new polymorphism in the APOB gene shows no evidence for an association with fatness in pigs

Sequence analysis of PCR products of a 343-bp fragment from exon 29 of the porcine APOB gene of four Erhualian and four Landrace pigs revealed a missense G/C substitution at position of 6117 in this gene. Two allele-specific primers were designed to genotype this polymorphism using the Bi-PASA technique. Genotyping of 146 animals from Erhualian, Hampshire, Large White, Landrace and Duroc breeds revealed large breed differences in allele frequency. No association with fatness was observed within each of the four European breeds, where animals had been selected to be those with the highest and those with the lowest backfat depth at 100 kg liveweight from a large carcass dissection project.     

SIM1基因第8外显子的SNP对猪背膘厚的影响

采用PCR-SSCP方法检测了约克夏、杜洛克、皮特兰、长白猪、嘉兴黑猪和金华猪6个品种共169头猪的SIM1基因外显子8的SNP及其基因型频率。结果共发现CC、CT、TT 3种基因型, 其基因型频率在国内外猪品种之间具有较大差异。其中, 国内猪种嘉兴黑猪和金华猪只存在TT基因型, 而国外猪种约克夏、杜洛克、皮特兰、长白猪则都存在3种基因型。用最小二乘法分析SNP对长白猪、约克夏猪和杜洛克猪的背膘厚的效应的结果表明, 纯合基因型个体的背膘厚大于杂合基因型个体。SIM1基因型对国外猪种背膘厚有显著效应(P< 0.05), 并且不同部位效应不同。     

Genetic Variation and Association of Insulin-Like Growth Factor Binding Protein-3 with Performance in Swine

Genetic variation of insulin-like growth factor binding protein-3 was analyzed in 17 pig breeds (14 native Chinese and 3 European). Using PCR-single strand conformation polymorphism, we found a polymorphism in intron 2, and this SNP was the combined mutation of G897T-G903A-C911T. The Chinese breeds carried a higher TAT/TAT genotype frequency (over 50%), except for Bamei (22%), Yujiang Black (0.0%), and Erhualian (10.0%); the European breeds had a higher GGC/GGC genotype frequency (Large White 1.67%, Landrace 13.89%, Duroc 0.0%). The allelic frequency of TAT in Chinese breeds was over 50%, except for Yujiang Black (12.5%); the allelic frequency of GGC was over 50% in all European breeds. The effect of genotype on 43 performance traits was investigated in one population (Lantang × Landrace). Pigs with the TAT/TAT genotype had higher B-point and C-point back-fat thickness than pigs with the GGC/GGC genotype. The TAT/TAT pigs also scored higher in meat color than the GGC/GGC pigs. These results implied that IGFBP-3 may affect meat quality and carcass traits.     

Male-specific DNA sequences in pigs

One hundred primers (Operon kits OPAA, OPAO, OPAV, OPC, and OPE series) were used for random amplified polymorphic DNA (RAPD) fingerprinting to determine male-specific fragments. Seventy-four percent of the primers yielded Yorkshire polymorphic fragments. One of these primers, OPAV-18, produced a novel 1098-bp DNA fragment found only in tested males. This male-specific fragment was isolated and constructed into plasmids for nucleotide sequencing. Two primers (5'-TTGCTCACGG TAGATAACAA GAGAG-3' and 5'-TTGCTCACGG ACCAGGTAGG GAATG-3') were designed according to the cloned male-specific sequence to amplify the male-specific band using polymerase chain reaction (PCR) for pig sexing. Sex-specific bands in the PCR gel products were represented in males but none were found in females when Yorkshire, Duroc, and Landrace genomic DNA samples were amplified with these two primers by PCR. The PCR products in the gel were transferred to nylon membranes and hybridized with a 32P-dCTP labeled probe of the cloned male-specific DNA fragment. There was a clear hybridization signal in samples from all of the male pigs, but not from those of female pigs. Male and female genomic DNA samples from these pigs were spotted onto nylon membranes and hybridized with the male-specific probe. The probe hybridized strongly to males only. A high degree of sequence homology was found among the novel male-specific DNA sequences in Yorkshire, Duroc and Landrace. The sex of these three breeds of pigs could be easily and effectively determined using these two primers.     

The effect of a c.-8G>T polymorphism on the expression of cytochrome b5A and boar taint in pigs

The level of cytochrome b5A ( CYB5A ) in pig testis is correlated with boar taint from androstenone and an AF016388:c.-8G>T polymorphism in CYB5A has been linked with low androstenone levels in the fat of pigs. In this study, we developed a polymerase chain reaction-based assay to genotype 1242 boars from eight lines for the c.-8G>T SNP. The c.-8T allele was found in all eight lines at a frequency ranging from 1.8% to 20.3% with an overall frequency of 8.6%. Significant deviations from Hardy–Weinberg equilibrium were found in the Hampshire, Landrace and Yorkshire breeds. The homozygous mutant c.-8TT occurred infrequently and was not found in some lines, but was consistently associated with low androstenone levels in fat. Both CYB5A mRNA and CYB5A protein levels were decreased in the c.-8TT genotype in a subset of Yorkshire boars, suggesting that low levels of CYB5A protein in the c.-8TT mutant were not due to inefficient translation of CYB5A mRNA. There were significant but modest marker effects on fat androstenone levels in Landrace, Yorkshire and a Large White/Duroc cross and fat skatole in Duroc and Sire Line breeds. There was no effect of CYB5A genotype on bulbourethral gland length, suggesting that this SNP will not affect reproductive traits. We conclude that the c.-8G>T SNP in the CYB5A gene has a significant but modest effect on boar taint in male pigs and could be useful in some breeds as part of a panel of SNP markers in a marker-assisted selection programme to produce low boar taint pigs.