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)。     

Single nucleotide polymorphisms of the prolactin receptor (PRLR) gene and its association with growth traits in chinese cattle

Genetic polymorphism of the prolactin receptor (PRLR) gene was detected by PCR-SSCP and DNA sequencing methods in 665 individuals from five Chinese cattle breeds. The results showed that at the P1 locus, three observed genotypes (AA, AB and BB), two linked SNPs (G1267A and T1268C), and one missense mutation (S18N) within a putative signal peptide were determined. The frequencies of haplotypes A and B in the five breeds were 0.596–0.802 and 0.198–0.404, respectively. Polymorphism of the PRLR gene was shown to be significantly associated with growth traits in the Nanyang breed. Individuals with genotype BB had greater hucklebone width, body weight and average daily gain than those with genotype AA at 6 months old (P < 0.01), as well as better body height, body length and heart girth when 6 months (P < 0.05). This study revealed for the first time that the PRLR gene is a promising candidate gene that affects growth traits in cattle.     

Novel SNPs in the bovine Transmembrane protein 18 gene, their linkage and their associations with growth traits in Nanyang cattle

Transmembrane protein 18 (TMEM18) gene is expressed in the central nervous system that has recently been linked to human obesity and body mass index (BMI) in genome wide association studies (GWAS). In this study, two novel single nucleotide polymorphisms (SNPs), g.3835 G > A (aa. Gly > Ser) and g.3865 A > G were detected in cattle TMEM18 gene by DNA sequencing. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and forced PCR-RFLP methods were used to genotype the two SNPs in 1218 Chinese native cattle individuals. The association between novel SNPs and growth traits (especially on body weight and body height) in Nanyang cattle showed that individuals with GG genotype was higher than those with AA genotype at g.3835G>A locus (P < 0.05) in Nanyang cattle. Significant association also revealed that individuals of AA genotype at g. 3865 A>G locus were higher in terms of body weight, average daily gain, chest circunmference, body length (P < 0.05), body height and hucklebone width (P < 0.01) than GG genotypes in Nanyang cattle. Moreover, the effect of XspI-MluI combinations with growth traits in Nanyang cattle revealed that AAGG type had higher body weight, body length, chest circumference and average daily gain (P < 0.05) than GGAG in Nanyang cattle aged at 6 months.     

<Emphasis Type="Italic">IGF-1</Emphasis> and <Emphasis Type="Italic">IGF-1R</Emphasis> gene polymorphisms in East Anatolian Red and South Anatolian Red cattle breeds

The aim of this study was to determine the allele and genotype frequencies of the Insulin like growth factor 1 gene (IGF-1) and Insulin like growth factor 1 receptor gene (IGF-1R) polymorphisms in East Anatolian (EAR) and South Anatolian Red cattle (SAR). Polymorphisms in both genes are claimed to affect economic parameters like body weight and subcutaneous back fat. For the study, blood samples were collected from 50 SAR and 50 EAR cattle. In both breeds, high frequencies of allele B of IGF-1, which was supposed to have positive effect on carcass traits and allele A of IGF-1R, related to milk traits, were observed. Therefore, no relationship of the polymorphisms studied with economic traits was observed, as both breeds have respectively low carcass and milk parameters. However, distribution of allele frequencies of IGF-1 and IGF-1R in SAR and EAR cattle was similar with Zebu cattle that support Bos indicus introgression to Anatolian breeds.     

郏县红牛ZAG基因多态性与生长性状的相关性

摘要: ZAG基因的功能主要是促进脂肪分解, 减少脂肪含量。文章利用PCR-SSCP和DNA测序技术研究了145头郏县红牛ZAG基因编码区4个位点(Z1、Z2、Z3、Z4)的多态性, 发现Z1、Z3、Z4 位点存在SSCP多态。对不同SSCP带型的对应片段进行了测序分析, 共发现6个新的SNP多态位点(C115T、A3257G、A4013G、T4027C、C4032T、T4120C)。Z3位点处于Hardy-Weinberg平衡状态, Z1、Z4 位点处于非平衡状态。不同基因型与生长发育性状的相关性分析显示, Z4位点上, AC基因型个体的体斜长、胸围、管围、体重指标显著(P<0.05)或极显著(P<0.01), 大于AA、AB基因型个体, 暗示该位点有可能作为郏县红牛生长性状标记辅助选择的标记之一。     

Novel SNPs of the mtDNA ND5 Gene and Their Associations with Several Growth Traits in the Nanyang Cattle Breed

Polymorphism of the mtDNA ND5 gene was detected by PCR-SSCP and DNA sequencing methods in 714 individuals from six Chinese cattle breeds. The results showed that at the P2 locus, two kinds of haplotypes, named A and B, with three SNPs (T12900C, A12923T, C12924T) were detected. The frequencies of haplotype A in six breeds were 0.041–0.167. Polymorphism of the ND5 gene was shown to be associated with growth in the Nanyang breed. Individuals with haplotype B had greater hucklebone width than those with haplotype A when 6 months old (P < 0.01), as well as better body height, body length, body weight, and average daily gain at 6 months (P < 0.05). Therefore, haplotype B of the ND5 gene was likely to have a positive effect on growth traits at 6 months in the Nanyang breed.     

Novel SNPs of the bovine <Emphasis Type="Italic">GAD1/gad67</Emphasis> gene and their association with growth traits in three native Chinese cattle breeds

In this study, polymorphism in the exon 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 and 11 of bovine NUCB2 gene was detected by PCR-SSCP and DNA sequencing methods in 686 individuals from three Chinese cattle breeds. Two haplotypes (M and N), three observed genotypes (MM, MN and NN) and two SNPs (NC_007313: g. 27451G>A, NC_007313: g. 27472T>C) were detected. The frequencies of haplotypes M and N in inland Chinese three breeds were 0.531–0.721 and 0.279–0.469 respectively. The studied showed that Nanyang, Jiaxian Red and Qinchuan cattle populations were in Hardy–Weinberg equilibrium at SNPs locus of NUCB2 gene (P > 0.05). Polymorphism of the NUCB2 gene was shown to be associated with growth traits in Qingchuan and Nanyang cattle breed. The linkage of two mutant sites in the bovine NUCB2 gene had significant effects on body length, body weight, heart girth, and average daily gain at 24 months (P < 0.05). Results of this study suggested that the NUCB2-gene-specific SNP may be a useful marker for growth traits in future marker-assisted selection programmes in inland Chinese cattle.     

Identification of two loci for resistance to clubroot (<Emphasis Type="Italic">Plasmodiophora brassicae</Emphasis> Woronin) in <Emphasis Type="Italic">Brassica rapa</Emphasis> L.

In an analysis of 114 F₂ individuals from a cross between clubroot-resistant and susceptible lines of Brassica rapa L., 'G004' and 'Hakusai Chukanbohon Nou 7' (A9709), respectively, we identified two loci, Crr1 and Crr2, for clubroot (caused by Plasmodiophora brassicae Woronin) resistance. Each locus segregated independently among the F₂ population, indicating that the loci reside on a different region of chromosomes or on different chromosomes. Genetic analysis showed that each locus had little effect on clubroot resistance by itself, indicating that these two loci are complementary for clubroot resistance. The resistance to clubroot was much stronger when both loci were homozygous for resistant alleles than when they were heterozygous. These results indicate that clubroot resistance in B. rapa is under oligogenic control and at least two loci are necessary for resistance.Communicated by H.C. Becker     

The novel 5bp deletion polymorphism in the promoter region of bovine <Emphasis Type="Italic">ACRP30</Emphasis> gene

ACRP30 gene was located nearby the QTL affecting the marbling, ribeye muscle area and fat thickness on the BAT1 in Angus. In this study, a 5bp deletion mutation within the bovine ACRP30 gene was firstly detected and confirmed in 991 cattle by PCR-SSCP, DNA sequencing and direct PCR amplification. The deletion mutation was appeared in Qinchuan, Nanyang, Jiaxian and Hasake, but was not found in Jinnan, Chinese Holsteins and Angus. The association of the deletion polymorphism with growth traits (including birth weight, body weight, average daily gain and body sizes in different growth periods (6/12/18/24 month-old)) was analyzed in 224 Nanyang cattle. No signification association of the deletion polymorphism with growth traits were observed (P > 0.05). The deletion was located in the promoter region and it resulted in a new putative CCAAT/enhancer binding protein-β response element (C/EBP-RE).     

The polymorphisms of bovine PCSK1 gene and their associations with growth traits

Proprotein convertase 1 (PCSK1) is an endopeptidase involved in proteolytic processing of peptide hormone precursors in granules of the regulated secretory pathway of endocrine cells and mutations in PCSK1 gene are thought to cause obesity. In the present study, based on PCR-SSCP and DNA sequencing methods, polymorphisms of the PCSK1 gene were detected in 858 individuals from five breeds (Nanyang cattle, Qinchuan cattle, Jiaxian cattle, Luxi cattle and Chinese Holstein). The results showed that only P₈ locus showed polymorphisms and 3 synonymous SNPs of PCSK1 gene were identified. Additionally, significant statistical difference was found in bovine birth weight and diplotype MM were 7.35% higher than diplotype XY.     

Isolation and characterization of polymorphic microsatellite markers for the endangered tree <Emphasis Type="Italic">Handeliodendron bodinieri</Emphasis> (Sapindaceae)

Nine polymorphic microsatellite loci were developed and characterized for Handeliodendron bodinieri, an endangered tree endemic to karst forest in southwest China. Each locus displayed between 4 and 11 alleles. The observed heterozygosity ranged from 0.345 to 0.931 and expected heterozygosity varied from 0.414 to 0.824. The combined probabilities of paternity exclusion over all loci were 0.931 for P _r(Ex₁) and 0.993 for P _r(Ex₂), respectively. These loci represent a powerful tool in investigating gene flow and population structure in natural populations of H. bodinieri.     

奶牛TLR4基因的多态性与乳房炎的相关性研究

TLR4通过识别病原体而激活免疫细胞,在先天免疫和适应性免疫防御中起着重要作用。以中国荷斯坦奶牛、三河牛和中国西门塔尔牛为研究对象,以TLR4为乳房炎抗性的候选基因,分别扩增316bp和382bp2个片段,分别采用SSCP和RFLP-AluⅠ方法来检测TLR4基因的多态性。结合测序发现:在intron1的第4,525bp处的A→G的突变,和exon3的第1,397bp处的T→C突变,使得产生多态。2个位点的A、B等位基因在3个群体中都有分布,且处于中度多态。χ2适合性检验表明,3个群体在这2个位点的突变达到Hardy-Weinberg平衡状态(P>0.05)。运用SAS8.0软件采用最小二乘法拟合线性模型,将基因座不同基因型与奶牛乳房炎进行了关联分析,结果表明:T4CRBR1的AA基因型为乳房炎抗性基因型(P<0.05),A等位基因为乳房炎抗性的有利基因,T4CRBR2的各基因型个体间的体细胞评分差异不显著(P>0.05)。     

Association between IGF-IR, m-calpain and UCP-3 gene polymorphisms and growth traits in Nanyang cattle

The objective of this study was to estimate the allele and genotype frequencies of the IGF-IR/TaqI, m-calpain/HhaI, and UCP-3/BglI polymorphisms and to determine associations between these polymorphisms and growth traits in Chinese indigenous cattle breeds. Genotyping was performed on 321 animals including 135 Nanyang, 80 Qinchuan, and 106 Jiaxian cattle. No significant differences in growth traits were observed between the genotypes of IGF-IR/TaqI polymorphism in Nanyang cattle. The m-calpain/HhaI, and UCP-3/BglI polymorphisms were associated with body weight, withers height, and body length of 6 months (P < 0.05 or P < 0.01), body length of 18 months (P < 0.05), and body length and heart girth of 24 months (P < 0.01) in Nanyang cattle. In addition, Nanyang cattle aged 12 months with AB genotype of m-calpain/HhaI polymorphism had higher body length and heart girth than those with BB genotype (P < 0.01). The withers height were greater (P < 0.01) in Nanyang cattle aged 12 months with genotype AB of UCP-3/BglI polymorphism than those with genotype BB. The Nanyang cattle aged 24 months with genotype AA had higher withers height than those with genotype BB (P < 0.05). In conclusion, the m-calpain/HhaI and UCP-3/BglI polymorphisms may be used as DNA markers for selection in the breeding process of Nanyang cattle.     

Haplotype distribution in the GLI3 gene and their associations with growth traits in cattle

The glioma-associated oncogene family zinc finger 3 gene (GLI3) mediates in all vertebrates hedgehog (Hh) signaling that plays an essential role in the induction and patterning of numerous cell types during invertebrate and vertebrate development. In this study, a total of 6 single nucleotide polymorphisms (SNPs: 1–6) were identified by polymerase chain reaction–single stranded conformational polymorphism (PCR–SSCP) and DNA pool sequencing, including all 13 exons and 12 exon–intron boundaries within the bovine GLI3 gene. 16 haplotypes and 13 combined genotypes were revealed and the linkage disequilibrium was assessed in 708 individuals representing three main cattle breeds from China. The statistical analyses indicated that the SNP2, 3 and 4 are associated with the body weight at birth and 6 months in Nanyang cattle population (P < 0.05). No significant association was detected between 11 combined genotypes and body weight at five different ages. Our results provide evidence that polymorphisms in the GLI3 gene are associated with growth traits, and may be used for marker-assisted selection in beef cattle breeding program.     

Genetic association of <Emphasis Type="Italic">Phosphodiesterase 1B</Emphasis> (PDE1B) with carcass traits in Korean cattle

Quantitative trait loci for fat deposition and carcass traits have been identified in the vicinity of the gene encoding phosphodiesterase 1B (PDE1B) on bovine chromosome 5. Therefore, the PDE1B gene can be considered as a positional and functional candidate gene for carcass traits in beef cattle. This study aimed to identify single nucleotide polymorphisms (SNPs) in the PDE1B gene and to evaluate their associations with carcass traits in Korean cattle. Eight SNPs, g.440T>G, g.17122A>G, g.17507A>C, g.17575A>G, g.17607T>C, g.17609C>A, g.17692C>T, and g.17707C>G, were identified in the region ranged from exon 1 to intron 6. Five of them were used for association analysis because of their availability of restriction fragment length polymorphisms. As a result, g.17122A>G in intron 3 was significantly associated with backfat thickness (BFT), and g.17507A>C in exon 5 was associated with longissimus dorsi muscle area (LMA, P < 0.05). Animals with the AG genotype of g.17122 had thicker BFT than those with the AA genotype. Animals with the AA or AC genotype of g.17507A>C had larger LMA than those with the CC genotype. We suggested the PDE1B gene as a candidate gene for carcass traits of beef cattle. Fine mapping would be required for application to marker-assisted selection.     

Polymorphisms of <Emphasis Type="Italic">STAT5A</Emphasis> gene and their association with milk production traits in Holstein cows

The STAT5A gene was studied as a candidate gene for five milk production traits (milk yield at 305 days, protein percentage, fat percentage, lactose percentage and dry matter percentage) in Holstein cows. According to the sequence of bovine STAT5A gene, two pairs of primers (P1 and P2) were designed to detect polymorphisms of STAT5A gene in 401 Holstein cows by PCR-RFLP and PCR-SSCP. The results showed that the products amplified by primers P1 and P2 displayed polymorphisms. For P1, three genotypes (AA, AG, and GG) were detected, and the frequency of AA/AG/GG was 0.252/0.486/0.262, respectively. Sequence analysis revealed a single nucleotide substitution A–G at 14217 bp (GenBank NC_007317) of bovine STAT5A gene while compared GG genotype with AA genotype. The differences of the least squares means for the four milk production traits (milk yield at 305 days, fat percentage, lactose percentage and dry matter percentage) between AA, AG and GG were not significant (P > 0.05). Least squares mean of protein percentage for AG or GG was significantly higher than that for AA (P < 0.05); the difference of the least squares mean for protein percentage was not significant between AG and GG (P > 0.05). For P2, three genotypes (CC, CT, and TT) were detected in Holstein cows, and the frequency of CC/CT/TT was 0.751/0.234/0.015, respectively. Sequencing revealed an insertion CCT at 17266 (NC_007317) of bovine STAT5A gene while compared CC genotype with TT genotype. The differences of the least squares means for the three milk production traits (protein percentage, lactose percentage and dry matter percentage) between CC, CT and TT were not significant (P > 0.05). Least squares mean of milk yield at 305 days for TT or CT was significantly higher than that for CC (P < 0.05); the difference of the least squares mean for milk yield at 305 days was not significant between TT and CT (P > 0.05). Least squares mean of fat percentage for CC or CT was significantly higher than that for TT (P < 0.05); the difference of the least squares mean for fat percentage was not significant between CC and CT (P > 0.05). The results preliminarily indicated that allele G of A14217G polymorphic site of STAT5A gene is a potential DNA marker for improving protein percentage in dairy cattle, 17266indelCCT polymorphic site of STAT5A gene is a potential DNA marker for improving milk yield at 305 days and fat percentage in dairy cattle.     

Effect of Genetic Variations of the POU1F1 Gene on Growth Traits of Nanyang Cattle

PCR-RFLP was applied to analyze the effect of the genetic variations of the POU1F1 gene on growth traits of 100 Nanyang cattle. The results showed that the 451 bp PCR product digested with Hinf I demonstrated polymorphism in the population, which was at Hardy-Weinberg equilibrium. Moreover, the frequencies of alleles A/B in the Nanyang population were 0.465/0.535. The association of the variations of the POU1F1 gene with the growth traits in the population was analyzed. The following parameters were greater in individuals with a genotype BB than in those with an genotype AB: birth weight, average weight increase before ablactation, body height at 12 months, body weight, body length, and chest girth at 6 months and 12 months (P<0.05). The body weight at 12 months was higher in the BB individuals than in the AA individuals (P <0.05). The body weight and body sizes also showed a trend of allele B> allele A in the other age groups. Therefore, the genotype BB maybe a dominant genotype and the allele B may be a dominant allele. These results imply that the allele B of the POU1F1 gene is likely to positively affect the growth traits.