PRNP gene variation in Pakistani cattle and buffaloes

Bovine spongiform encephalopathy (BSE) is a neurodegenerative prion protein misfolding disorder of cattle. BSE is of two types, classical BSE and atypical BSE which in turn is of two types, H-type BSE and L-type BSE. Both H-type BSE and L-type BSE are primarily sporadic prion disorders. However, one case of H-type BSE has recently been associated with E211K polymorphism in the prion protein gene (PRNP). Two polymorphisms in the bovine PRNP are also associated with susceptibility to classical BSE: a 23 bp insertion/deletion (indel) in the PRNP promoter region and a 12 bp indel in the first intron. No information regarding BSE susceptibility in Pakistani cattle is available. The present study aimed at achieving this information. A total of 236 cattle from 7 breeds and 281 buffaloes from 5 breeds were screened for E211K polymorphism and 23 bp and 12 bp indels employing triplex PCR. The E211K polymorphism was not detected in any of the animals studied. The 23 bp insertion allele was underrepresented in studied cattle breeds while the 12 bp insertion allele was overrepresented. Both 23 bp and 12 bp insertion alleles were overrepresented in studied buffalo breeds. Almost 90% of alleles were insertion alleles across all studied buffalo breeds. The average frequency of 23 bp and 12 bp insertion alleles across all studied cattle breeds was found to be 0.1822 and 0.9407, respectively. There were significant differences between Pakistani and worldwide cattle in terms of allele, genotype and haplotype frequencies of 23 bp and 12 bp indels. The higher observed frequency of 12 bp insertion allele suggests that Pakistani cattle are relatively more resistant to classical BSE than European cattle. However, the key risk factor for classical BSE is the dietary exposure of cattle to contaminated feedstuffs.     

Polymorphism of cattle prolactin gene: microsatellites, PSR-RFLP]

In the samples of Russian Ayrshire and Gorbatov Red cattle breeds, distribution of frequencies of prolactin (PRL) gene alleles generated due to the presence of polymorphic RsaI site in exon 3 were studied. In the breeds, the frequencies of the B allele of the PRL gene (with RsaI(+) site) detected by the PCR-RFLP method were 14.1 and 8.6%, respectively. In Black Pied, Ayrshire and Gorbatov Red cattle breeds, variation of the microsatellite dinucleotide repeat in the regulatory region of the gene PRL was also studied. Gorbatov Red breed was monomorphic at the microsatellite locus with the only allele 164 bp in length. Two alleles (164 bp and 162 bp) were detected in the other breeds studied. The frequencies of 164-bp allele of the microsatellite locus were 93.7 and 90.0% in Black Pied and Ayrshire breeds, respectively. In Gorbatov Red breed of dairy type with good beef qualities and low milk-fat yield, lower level of heterozygosity for PRL gene was demonstrated compared to Ayrshire and Black Pied breeds with high milk-fat yield. In three cattle breeds, higher mean estimate of polymorphism information content of PCR-RFLP in exon 3 (PIC = 0.21) was revealed compared with the same estimate (PIC = 0.09) for the microsatellite locus variability in the regulatory region of the PRL gene. Characteristics of allele B distribution of the PRL gene in the representatives of the Bovidae family are considered.     

Genetic polymorphism of the kappa-casein gene in Brazilian cattle

Frequencies of kappa-casein gene alleles were determined in 1316 animals from the Brazilian Bos indicus genetic groups (Sindhi cows, Gyr sires, Gyr cows, Guzerat sires, Guzerat cows, Nellore sires, and Gyr x Holstein crossbreds) by means of polymerase chain reaction-restriction fragment length polymorphism analysis using two independent restriction nucleases (Hinf I and HaeIII). The genotyping of kappa-casein alleles (A and B) is of practical importance, since the B allele is found to correlate with commercially valuable parameters of cheese yielding efficiency. The frequencies of the B allele of kappa-casein among breeds ranged from 0.01 to 0.30. The Sindhi breed had the highest frequency for the B allele (0.30), while the frequencies of this allele in other breeds ranged from 0.01 to 0.18. A wide variation in the B allele frequency among B. indicus breeds was found suggesting that molecular selection for animals carrying the B allele could impact breeding programs for dairy production.     

Polymorphism analysis of prion protein gene in 11 Pakistani goat breeds

The association between caprine PrP gene polymorphisms and its susceptibility to scrapie has been investigated in current years. As the ORF of the PrP gene is extremely erratic in different breeds of goats, we studied the PrP gene polymorphisms in 80 goats which belong to 11 Pakistani indigenous goat breeds from all provinces of Pakistan. A total of 6 distinct polymorphic sites (one novel) with amino acid substitutions were identified in the PrP gene which includes 126 (A -> G), 304 (G -> T), 379 (A -> G), 414 (C -> T), 428 (A -> G) and 718 (C -> T). The locus c.428 was found highly polymorphic in all breeds as compare to other loci. On the basis of these PrP variants NJ phylogenetic tree was constructed through MEGA6.1 which showed that all goat breeds along with domestic sheep and Mauflon sheep appeared as in one clade and sharing its most recent common ancestors (MRCA) with deer species while Protein analysis has shown that these polymorphisms can lead to varied primary, secondary and tertiary structure of protein. Based on these polymorphic variants, genetic distance, multidimensional scaling plot and principal component analyses revealed the clear picture regarding greater number of substitutions in cattle PrP regions as compared to the small ruminant species. In particular these findings may pinpoint the fundamental control over the scrapie in Capra hircus on genetic basis.     

12个水牛群体催乳素基因第4外显子遗传特征分析

催乳素对水牛乳腺发育、泌乳和乳蛋白基因的表达有明显的调控作用。该研究采用直接测序并结合PCR-SSCP技术,分析沼泽型水牛和河流型水牛12个群体385个个体的催乳素基因(PRL)第4外显子(exon4)的遗传特征。结果表明:水牛PRLexon4由180个核苷酸组成,在不同物种中具有高度保守性。序列分析发现水牛中该外显子的第109位碱基处有C>T替换,为沉默突变,与泌乳性状之间无显著相关性。在9个沼泽型水牛群体中均检测到该突变位点,其中PBA基因在7个沼泽型水牛群体中为优势基因,PBB基因在德宏和富钟群体中为优势基因,沼泽型水牛群体中PBA基因频率在0.400～0.917之间,群体平均值为0.629±0.049,具有地域分布特征。在3个河流型水牛群体中,第109位核苷酸处均为C,未检测到多态。提示河流型水牛与沼泽型水牛已有较大的遗传分化。     

荷斯坦牛Nramp1基因遗传多态性及其与乳房炎相关性的研究

利用PCR-SSCP技术检测了344头中国荷斯坦牛Nramp1基因exon 11的基因多态性, 并分析了其不同基因型与乳房炎及产奶量性状的关系。结果表明: 实验群体发现3种基因型AA、AB、BB, 其中A等位基因为优势等位基因, 等位基因频率为0.767, 而B等位基因频率则为0.233。经χ2适合性检验, 群体处于Hardy-Weinberg平衡状态(P>0.05)。测序结果显示: 扩增片段分别在200 bp(C/G)和254 bp(T/G)存在碱基突变, 并导致了氨基酸改变, 分别为丙氨酸替换为脯氨酸(Ala356Pro)、亮氨基酸替换为蛋氨酸(Leu374Met)。通过构建最小二乘线性模型, 进行Nramp1基因多态性与产奶量、体细胞评分(SCS)的相关性分析表明, AA型个体的SCS最小二乘均值显著低于BB﹑AB型(P<0.05), 而AA型﹑AB个体的产奶量最小二乘均值显著高于BB型(P<0.01, P<0.05), AA基因型可作为乳房炎抗性的优良基因型。因此, 可将Nramp1作为奶牛乳房炎候选基因应用于分子标记辅助选择育种。     

牛RXRG基因遗传变异与双胎性状的关联分析

以视黄素X受体基因g(retinoid X receptor-gamma, RXRG)作为牛双胎性状的候选基因, 运用测序法寻找牛RXRG基因SNPs, 筛查到一个新的多态位点A1941G, 该位点位于3′UTR。运用PCR-RFLP法验证并分析该位点在鲁西牛双胎群体和单胎群体及中国西门塔尔牛、安格斯牛和西蒙杂交牛单胎群体间的多态性, 结果表明, 在鲁西牛双胎和单胎群体中分布A、B两个等位基因,处于中度多态。经χ2适合性检验, 鲁西双胎牛群体在该位点未达到Hardy-Weinberg平衡状态(P < 0.05)。将鲁西牛群体的A1941G位点的基因型效应与双胎性状进行关联分析, 卡方独立性检测结果显示, 基因型分布在鲁西单、双胎牛群体上差异达到极显著水平(P < 0.01)。     

Genetic aspects of twinning in cattle

Twinning in cattle ranges from about 1% for beef breeds to about 4% for dairy breeds. The incidence of double births may have both positive and negative effects, which mainly depends on the purpose for which the cattle are raised. Because of freemartinism, as well as management problems connected e.g. with a greater risk of dystocia and retained placenta, it is an undesirable trait in dairy herds. In beef cattle, however, twinning can considerably increase the efficiency of production. Low heritability, a long generation interval for progeny testing, sex-limited expression and an unfavourable correlation with milk yield make twinning difficult to control by selection. Hence, it is the type of trait for which the identification of the genetic marker - quantitative trait loci (QTL) linkage and the implementation of marker-assisted selection in breeding strategies are expected to be especially beneficial. Searching for QTL influencing the reproductive rate in cattle was performed mainly in the US Meat Animal Research Center twinning herd and in the commercial Norwegian cattle population. Among several genome regions that appear to control twinning and ovulation rates, the most interesting seem to be chromosomes 5, 7, 19 and 23.     

Genetic variation in the bovine myostatin gene in UK beef cattle: allele frequencies and haplotype analysis in the South Devon

Work on Belgian Blue cattle revealed that an 11 base pair (bp) deletion within the bovine myostatin gene (GDF8) is associated with the double-muscled phenotype seen in this breed. Investigations focusing on other European breeds known to show double-muscling identified several mutations within the coding region of the gene associated with the double-muscled phenotype in different breeds. The number of mutations found suggest that myostatin is highly variable within beef cattle. Variations that alter the structure of the gene product such that the protein is inactivated are associated with the most pronounced form of double-muscling as seen in the Belgian Blue. However, other mutations may have a less extreme affect on muscle development. While overt double-muscling gives rise to a high incidence of dystocia (calving difficulty), it is possible that some variants may give enhanced muscling, but with limited calving problems. We describe sequence analysis of the myostatin gene in ten beef breeds commonly used in the UK and show that the 11-bp deletion responsible for double-muscling in the Belgian Blue is also present in the South Devon cattle population. Allele frequencies and haplotypes in the South Devon and a polymerase chain reaction (PCR) based test for the deletion are described. PCR amplification across the deleted region provides a quick and effective test with clear identification of heterozygous individuals. We discuss our results with regard to the effect of genotype on phenotype and differences observed between the Belgian Blue and the South Devon.     

Genetic analysis by chromosome-mediated gene transfer

Summary A general method is presented for stable transfer of genetic information to eukaryotic cells, utilizing metaphase chromosomes as the vehicle. Recent progress, current problems and large areas of uncertainty in this field are reviewed; particular consideration is given to frequency of transfer, size of the transgenome, evidence for cotransfer of linked genes and serial chromosome transfer. A reasonable model for chromosome transfer is considered with respect to the available information, and various discrepancies are noted. The utility of this method for fine structural mapping, cloning small regions of the eukaryotic genome and other potential applications are discussed. Presented in the formal symposium on Somatic Cell Genetics at the 27th Annual Meeting of the Tissue Culture Association, Philadelphia, Pennsylvania, June 7–10, 1976.     

Genetic diversity analysis of BMY cattle based on microsatellite DNA markers

BMY cattle (1/2 Brahman, 1/4 Murray Grey and 1/4 Yunnan Yellow cattle) has been inter se breeding since 1980s. Genetic diversity of BMY cattle was extensively investigated using 16 microsatellite markers. A total of 130 microsatellite alleles and high allele size variance were detected. All loci displayed high genetic diversity with overall mean of N(a) = 8.13, PIC = 0.7224 and H(e) = 0.7666, which were higher than those of many other beef breeds. The allele-sharing neighbour-joining tree clearly displayed the new genotypic combinations and the minglement from both BMY cattle and Brahman. The results provided the genetic information to match the standards of new beef breed in South China.     

Genetic diversity analysis of six Spanish native cattle breeds using microsatellites

Six native Spanish cattle breeds have been characterized by using 30 microsatellite markers. The studied populations can be divided into three groups: Brown orthoid (Asturian Mountain, Asturian Lowland and the Nord-west Brown Group), Red convex (Pyrenean and Menorquina) and the Iberian bovine (Fighting bull). Allele frequencies were calculated and used for the characterization of the breeds and the study of their genetic relationships. Different genetic distance measures were calculated and used for dendogram construction. The closest populations were those representing Asturian breeds, the most divergent being Menorquina and Fighting Bull. The latter also showed the lowest diversity values (mean number of alleles per locus and heterozygosity). Genetic distances obtained between the other populations under analysis were similar to those reported for different European cattle breeds. This work analyzes the recent origin of these populations and contributes to the knowledge and genetic characterization of European native breeds.     

Genetic diversity analysis of BMY cattle based on microsatellite DNA markers

cattle (1/2 Brahman, 1/4 Murray Grey and 1/4 Yunnan Yellow cattle) has been inter se breeding since 1980s. Genetic diversity of BMY cattle was extensively investigated using 16 microsatellite markers. A total of 130 microsatellite alleles and high allele size variance were detected. All loci displayed high genetic diversity with overall mean of N _a = 8.13, PIC = 0.7224 and H _e = 0.7666, which were higher than those of many other beef breeds. The allele-sharing neighbour-joining tree clearly displayed the new genotypic combinations and the minglement from both BMY cattle and Brahman. The results provided the genetic information to match the standards of new beef breed in South China.     

Polymorphism analysis of the prion gene in BSE-affected and unaffected cattle

Polymerase chain reaction (PCR) primers designed to amplify the octapeptide repeat region of the bovine prion gene were used to test the association of genotypes with bovine spongiform encephalitis (BSE) in 56 BSE-affected and 177 unaffected animals. Three alleles (A, B, C) were detected as single-strand conformation polymorphisms (SSCPs) and two alleles (1,2 representing six or five copies of the octapeptide repeat respectively) were detected as amplified double-strand fragment length polymorphisms (AMFLPs). Observed genotypes of SSCPs and AMFLPs were analysed by x-square. The SSCP genotypes of nuclear family members of animals with BSE and BSE-affected animals were different (P < 0.001, P < 0.01) from unrelated animals of the same breed without BSE. No genotypic differences were found between the BSE-affected animals and their relatives (P > 0.469). No AMFLP genotypic differences were detected between BSE-affected animals, their relatives, unrelated animals of the same breed or animals of different breeds (P > 0.05). These data suggest that BSE-affected animals and their relatives are more likely to have the AA SSCP genotype than unrelated animals of the same breed or animals of different breeds.