Epistatic kinship a new measure of genetic diversity for short-term phylogenetic structures – theoretical investigations

The epistatic kinship describes the probability that chromosomal segments of length x in Morgan are identical by descent. It is an extension from the single locus consideration of the kinship coefficient to chromosomal segments. The parameter reflects the number of meioses separating individuals or populations. Hence it is suggested as a measure to quantify the genetic distance of subpopulations that have been separated only few generations ago. Algorithms for the epistatic kinship and the extension of the rules to set up the rectangular relationship matrix are presented. The properties of the epistatic kinship based on pedigree information were investigated theoretically. Pedigree data are often missing for small livestock populations. Therefore, an approach to estimate epistatic kinship based on molecular marker data are suggested. For the epistatic kinship based on marker information haplotypes are relevant. An easy and fast method that derives haplotypes and the respective frequencies without pedigree information was derived based on sampled full‐sib pairs. Different parameters of the sampling scheme were tested in a simulation study. The power of the method decreases with increasing segment length and with increasing number of segments genotyped. Further, it is shown that the efficiency of the approach is influenced by the number of animals genotyped and the polymorphism of the markers. It is discussed that the suggested method has a considerable potential to allow a phylogenetic differentiation between close populations, where small sample size can be balanced by the number, the length, and the degree of polymorphism of the chromosome segments considered.     

利用SNP标记进行北京地区中国荷斯坦牛亲子推断的研究

本研究旨在利用SNP标记对北京地区中国荷斯坦牛群进行亲子推断,并分析场、母牛出生年月、公牛家系对系谱错误率的影响,以期为指导奶牛育种和生产管理提供依据。共选取了255个最小等位基因频率大于0.45的高多态SNPs标记,利用似然法,采用Cervus3.0软件对北京地区84头荷斯坦公牛和1 927头母牛进行亲子推断研究。结果显示,试验群体平均系谱错误率为20.9%,不同的场、出生年份和月份的母牛的系谱错误率有显著差异(P<0.05),而各公牛家系间系谱错误率差异不显著(P>0.05)。结果说明,错误系谱的发生主要是由于牛场本身记录不完善造成的。在我国亟需建立利用遗传标记监测、校正系谱准确性的制度,采取措施提高系谱的准确性,加快我国荷斯坦牛遗传改良进程。     

Effective population size of an indigenous Swiss cattle breed estimated from linkage disequilibrium

Effective population size is an important parameter for the assessment of genetic diversity within a livestock population and its development over time. If pedigree information is not available, linkage disequilibrium (LD) analysis might offer an alternative perspective for the estimation of effective population size. In this study, 128 individuals of the Swiss Eringer breed were genotyped using the Illumina BovineSNP50 beadchip. We set bin size at 50 kb for LD analysis, assuming that LD for proximal single nucleotide polymorphism (SNP)‐pairs reflects distant breeding history while LD from distal SNP‐pairs would reflect near history. Recombination rates varied among different regions of the genome. The use of physical distances as an approximation of genetic distances (e.g. setting 1 Mb = 0.01 Morgan) led to an upward bias in LD‐based estimates of effective population size for generations beyond 50, while estimates for recent history were unaffected. Correction for restricted sample size did not substantially affect these results. LD‐based actual effective population size was estimated in the range of 87–149, whereas pedigree‐based effective population size resulted in 321 individuals. For conservation purposes, requiring knowledge of recent history (<50 generations), approximation assuming constant recombination rate seemed adequate.     

A novel method for rapid and reliable detection of complex vertebral malformation and bovine leukocyte adhesion deficiency in Holstein cattle

Background: Complex vertebral malformation (CVM) and bovine leukocyte adhesion deficiency (BLAD) are two autosomal recessive lethal genetic defects frequently occurring in Holstein cattle, identifiable by single nucleotide polymorphisms. The objective of this study is to develop a rapid and reliable genotyping assay to screen the active Holstein sires and determine the carrier frequency of CVM and BLAD in Chinese dairy cattle population. Results: We developed real-time PCR-based assays for discrimination of wild-type and defective alleles, so that carriers can be detected. Only one step was required after the DNA extraction from the sample and time consumption was about 2 hours. A total of 587 Chinese Holstein bulls were assayed, and fifty-six CVM-carriers and eight BLAD-carriers were identified, corresponding to heterozygote carrier frequencies of 9.54% and 1.36%, respectively. The pedigree analysis showed that most of the carriers could be traced back to the common ancestry, Osborndale Ivanhoe for BLAD and Pennstate Ivanhoe Star for CVM. Conclusions: These results demonstrate that real-time PCR is a simple, rapid and reliable assay for BLAD and CVM defective allele detection. The high frequency of the CVM allele suggests that implementing a routine testing system is necessary to gradually eradicate the deleterious gene from the Chinese Holstein population.     

Study on the Strategies of Genotype Imputation

Genomic data is more and more widely used in livestock breeding. Genotype imputation is an important tool to handle missing values in genotypic data, and the quality of imputation results directly affects the subsequent analysis. To obtain good imputation results, a comprehensive imputation strategy needs to be formulated. We studied on the effects of several factors on genotype imputation by simulation. The factors included reference population size, genetic relationship (distance) between the target population and the reference population, the number of target sites (proportion), the minimum allele frequency (MAF), and the imputation algorithm. The results showed that the number of target sites was the main factor affecting the genotype imputation, and it showed significantly positive correlation with the quality of imputation(P<0.05). The reference population size was the main factor affecting the imputation error rate in Beagle5.1. Correspondingly, the number of target sites was the main factor affecting the imputation error rate in Minimac4. Genetic distance between the target population and the reference population had a more significant effect on the imputation quality of Beagle5.1 than Minimac4. In general, the imputation error rate increased as the increases of MAF in a site. When the number of individuals in the reference population was small and the number of target sites was large, the speed of Minimac4 was superior to Beagle5.1, but there was a reverse trend as the reference population size increased. On the premise of ensuring the imputation quality, Beagle5.1 had relatively lower requirements for the above factors. In contrast, when the number of target sites was low and reference population size was large, the imputation effect of Beagle5.1 was better, while Minimac4 was more suitable for the imputation of a small reference population size and a higher number of target sites. In this study, different strategies were formulated for different imputation purposes, and the study results would provide a reference for genotype imputation.     

基因型填充策略研究

基因组数据在畜禽遗传育种中的应用越来越广泛，基因型填充作为基因组数据处理的重要工具，填充结果的好坏直接影响后续分析，为了得到好的填充结果，需要制定完善的填充策略。本研究通过模拟数据探讨参考群体大小、目标群体与参考群体间遗传关系（距离）远近、目标位点数目（比例）、最小等位基因频率以及填充算法等因素对基因型填充效果的影响。结果表明，目标位点数目与填充效果呈显著的正相关（P<0.05），是影响基因型填充准确性的主要因素；参考群体大小是影响Beagle5.1填充错误率的主要因素，目标位点数目是影响Minimac4填充错误率的主要因素；目标群体和参考群体的遗传距离对Beagle5.1填充效果的影响较Minimac4更为显著；一般情况下，最小等位基因频率越高的位点填充错误率越高；在参考群体个体数量少且目标位点数目多的情况下，Minimac4的填充速度优于Beagle5.1，但随参考群体个体数目增加有逆趋势。在保证填充质量的前提下，Beagle5.1对本研究中几种因素的标准要求相对较低。相对地，当目标群体位点数目较低，参考群体个体数目较多时，Beagle5.1的填充效果更好，而Minimac4更适合参考群体个体数目较少，目标群体位点数目较高的填充中。本研究针对不同的填充目的制定了不同策略，为基因型填充标准提供了参考。     

Pedigree analysis of factor XI deficiency in Japanese black cattle

Using a DNA-based diagnostic test for factor XI deficiency in Japanese black cattle, we surveyed 123 cattle (42 sires and 81 dams) in Gifu and Hyogo prefectures, and calculated gene frequencies. In sires, we drew up the pedigree network of the cattle with the factor XI deficiency. Results showed that the mutated allele of factor XI deficiency was retroactive in at least 6 or more generations of sires. Frequencies of the mutant gene were higher at 26.4% in total, and at 33.3% in sires. All 7 cattle with the homozygote of mutated allele were clinically normal, and showed no bleeding episodes. The mutated allele of factor XI deficiency might be widespread among Japanese black cattle.     

Analysis of Two Chinese Yak(Bos grunniens) Population Using Bovine Microsatellite Primers

Two Chinese domestic yak populations representing the Plateau type and the Huanhu Alpine type were analysed with 12 bovine microsatellite primers. All primer pairs functioned in the yak genome and polymorphism was found at all loci. The allele size ranges and frequencies of the two yak populations were similar and there was considerable overlap with the allele size ranges observed in cattle. Data for European cattle breeds was obtained from the Cattle Diversity Database(CaDBase)to interpret the heterozygosity and genetic distance estimates in yak populations. Heterozygosity estimated for the two yak populations was comparable to that of European cattle while Nei's Genetic Distance DA between the two yak populations was less than distances between the most closely related German cattle breeds. Bovine microsatellite primers proved to be a valuable tool for characterization of yak populations.     

Technical note: prediction of breeding values using marker-derived relationship matrices

In livestock populations, estimation of breeding values for selection requires a matrix describing the additive relationship between individuals in the population. This matrix can be derived from pedigree information. In some livestock populations, pedigree information may be unavailable, incomplete, or in error. Here we use simulated data to demonstrate that marker-derived relationship matrices can be used to predict breeding values and estimate additive variance components, provided the markers are sufficiently dense. The approach is demonstrated for an Angus data set with 9,323 SNP markers genotyped.     

Marker-based estimates of between and within population kinships for the conservation of genetic diversity

In this article coefficients of kinship between and within populations are proposed as a tool to assess genetic diversity for conservation of genetic variation. However, pedigree-based kinships are often not available, especially between populations. A method of estimation of kinship from genetic marker data was applied to simulated data from random breeding populations in order to study the suitability of this method for livestock conservation plans. Average coefficients of kinship between populations can be estimated with low Mean Square Error of Prediction, although a bias will occur from alleles that are alike in state in the founder population. The bias is similar for all populations, so the ranking of populations will not be affected. Possible ways of diminishing this bias are discussed. The estimation of kinships between individuals is imprecise unless the number of marker loci is large (> 200). However, it allows distinction between highly related animals (full sibs, half sibs and equivalent relations) and animals that are not directly related if about 30–50 polymorphic marker genes are used. The marker-based estimates of kinship coefficients yielded higher correlations than genetic distance measures with pedigree-based kinships and thus to this measure of genetic diversity, although correlations were high overall. The relation between coefficients of kinship and genetic distances are discussed. Kinship-based diversity measures conserve the founder population allele frequencies, whereas genetic distances will conserve populations in which allele frequencies are the most different. Marker-based kinship estimates can be used for the selection of breeds and individuals as contributors to a genetic conservation programme.     

A novel method for rapid and reliable detection of complex vertebral malformation and bovine leukocyte adhesion deficiency in Holstein cattle

Background

Complex vertebral malformation (CVM) and bovine leukocyte adhesion deficiency (BLAD) are two autosomal recessive lethal genetic defects frequently occurring in Holstein cattle, identifiable by single nucleotide polymorphisms. The objective of this study is to develop a rapid and reliable genotyping assay to screen the active Holstein sires and determine the carrier frequency of CVM and BLAD in Chinese dairy cattle population.

Results

We developed real-time PCR-based assays for discrimination of wild-type and defective alleles, so that carriers can be detected. Only one step was required after the DNA extraction from the sample and time consumption was about 2 hours. A total of 587 Chinese Holstein bulls were assayed, and fifty-six CVM-carriers and eight BLAD-carriers were identified, corresponding to heterozygote carrier frequencies of 9.54% and 1.36%, respectively. The pedigree analysis showed that most of the carriers could be traced back to the common ancestry, Osborndale Ivanhoe for BLAD and Pennstate Ivanhoe Star for CVM.

Conclusions

These results demonstrate that real-time PCR is a simple, rapid and reliable assay for BLAD and CVM defective allele detection. The high frequency of the CVM allele suggests that implementing a routine testing system is necessary to gradually eradicate the deleterious gene from the Chinese Holstein population.     

Application of individual increase in inbreeding to estimate realized effective sizes from real pedigrees

The objective of this study was to test the performance of a recently proposed methodology for the estimation of realized effective size (N(e)) based on individual increase in inbreeding (DeltaF(i)) on several real pedigrees: (a) an experimental mice population; (b) a closed pedigree of fighting bulls; (c) the Spanish Purebred (SPB, Andalusian) horse pedigree; (d) the Carthusian strain of SPB pedigree; (e) the Spanish Arab horse pedigree; and (f) the Spanish Anglo-Arab horse pedigree. Several reference subpopulations were defined on the basis of generation length in order to consider only animals in the last generation, to assess the influence of the pedigree content on the estimates of N(e). The estimates of realized N(e) computed from DeltaF(i) (Ne) tended to be higher than those obtained from regression on equivalent generations. The new parameter Ne remained approximately stable when pedigree depth achieved about five equivalent generations. Estimates of take into account the genetic history of the populations, the size of their founder population, and the mating policy or bottlenecks caused by poor use of reproducing individuals. The usefulness of the realized N(e) computed from individual increase in inbreeding in real pedigrees is also discussed.     

Quantitative trait locus mapping based on selective DNA pooling

Concepts of a simple method to map quantitative trait loci (QTL) based on selective DNA pooling in half-sib family, backcross, and F₂ designs were developed. It is shown that the position of a QTL can be estimated from differences in allele frequencies for two flanking markers between individuals with high and low phenotypes and does not depend on the phenotypic means of the selected groups. An estimate of the QTL effect was obtained by relating group differences in phenotypic means to differences in QTL frequencies, which can be estimated from the QTL position and marker allele frequencies. Simulation of a half-sib family and a F₂ family of 2000 individuals showed that the method gives close to unbiased results when power is high. Biases increased when measurement errors on marker allele frequencies increased and when the effect of the QTL was small. Similarities of QTL mapping based on selective DNA pooling data and on individual genotyping data are discussed, as are opportunities to extend the selective DNA pooling method to the use of multiple markers and multiple half-sib family designs. This study shows that the use of selective DNA pooling can be extended from the detection of marker associations to the mapping of QTL. Selective DNA pooling can greatly reduce the number of genotypings required.     

Use of bovine single nucleotide polymorphism markers to verify sample tracking in beef processing

OBJECTIVE: To determine whether a selected set of 20 single nucleotide polymorphism (SNP) markers derived from beef cattle populations can be used to verify sample tracking in a commercial slaughter facility that processes primarily market (ie, culled) dairy cows. DESIGN: Prospective, blinded validation study. ANIMALS: 165 cows and 3 bulls from 18 states (82% Holstein, 8% other dairy breeds, and 10% beef breeds). PROCEDURE: Blood was collected by venipuncture from randomly chosen animals just prior to slaughter. The purported corresponding liver samples were collected during beef processing, and genotype profiles were obtained for each sample. RESULTS: On the basis of SNP allele frequencies in these cattle, the mean probability that 2 randomly selected individuals would possess identical genotypes at all 20 loci was 4.3 x 10(-8). Thus, the chance of a coincidental genotype match between 2 animals was 1 in 23 million. Genotype profiles confirmed appropriate matching for 152 of the 168 (90.5%) purported blood-liver sample pairs and revealed mismatching for 16 (9.5%) pairs. For the 16 mismatched sample pairs, 33% to 76% of the 20 SNP genotypes did not match (mean, 52%). Discordance that could be attributed to genotyping error was estimated to be < 1% on the basis of results for split samples. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that this selected set of 20 bovine SNP markers is sufficiently informative to verify accuracy of sample tracking in slaughter plants that process beef or dairy cattle. These or similar SNP markers may facilitate high-throughput, DNA-based, traceback programs designed to detect drug residues in tissues, control of animal diseases, and enhance food safety.     

西农萨能奶山羊CSN1S2基因多态与产奶量、体尺指标的相关分析

利用PCR2RFLP技术对69只西农萨能奶山羊的CSN1S2基因进行多态性分析。扩增产物为310bp的CSN1S2F基因片段,被Alw26I限制性内切酶消化后表现多态性。F等位基因为310bp,N等位基因为179和131bp,相应地,FF基因型为310bp,NF基因型为310,179和131bp,NN基因型为179和131bp。在该群体中,F等位基因频率为010870,N等位基因频率为019130,处于Hardy2Weinberg平衡状态。将西农萨能奶山羊CSN1S2F基因座不同基因型与平均产奶量进行相关分析,结果表明:CSN1S2的不同基因型对平均产奶量有显著影响,NN基因型个体产奶量最高,FF基因型个体产奶量最低,且FF基因型个体平均产奶量显著低于NN基因型个体(P<0105)。将西农萨能奶山羊CSN1S2F基因座不同基因型与体尺指标进行相关分析,结果表明:在初生重和体高指标上,NF基因型个体显著优于NN基因型个体(P<0105);在体斜长和管围指标上,NF基因型个体略优于NN基因型个体,但差异不显著(P>0105);在成年体重和胸围指标上,不同的基因型个体间无差异。     

Cosegregation of a factor VIII microsatellite marker with mild hemophilia A in Golden Retriever dogs

Mild hemophilia A (factor VIII deficiency) was diagnosed in Golden Retrievers and pedigree studies were undertaken to test the cosegregation of an intragenic factor VIII marker with the disease phenotype. The study population consisted of 30 client-owned dogs (22 males and 8 females). Hemophilic males (n = 12) typically demonstrated prolonged bleeding after trauma or surgery rather than spontaneous hemorrhagic events. The affected males had a proportionate reduction in factor VIII coagulant activity (mean FVIII:C = 4%) and factor VIII protein concentration (mean FVIII:Ag = 3%). Twenty-five dogs (10 affected males, 8 clear males, 2 obligate carrier dams, and 5 suspect carrier daughters) were genotyped for a factor VIII microsatellite marker, with allele size assigned by an automated capillary electrophoresis system. Five distinct marker alleles were present in the study pedigree and a 300-base pair allele was found to segregate with the hemophilia A phenotype. The inheritance of the hemophilia-associated allele defined carrier status for 5 suspect daughters of obligate carrier dams. The limitations inherent to linkage analyses (i.e., lack of access to key family members and homozygosity at the marker locus) did not preclude carrier detection in this pedigree. We conclude that genotype analysis for the intragenic factor VIII marker can aid in control of canine hemophilia A through enhanced carrier detection.     

四川常羽乌骨鸡群体遗传多样性分析

利用选自家禽基因组的10个微卫星标记,对四川常羽乌骨鸡5个群体(四川山地乌骨鸡白羽系、黑羽系;黄忠山地乌骨鸡;黄忠山地乌骨鸡绿壳蛋系;草科乌骨鸡)的遗传多样性进行了检测,计算了各群体的群体杂合度、群体间遗传距离,并根据遗传距离进行了聚类分析。结果表明所选择的微卫星标记在各群体中表现出较高的多态性;四川5个乌骨鸡群体遗传多样性比较丰富,群体平均杂合度为0.5383到0.6659;各群体间的遗传距离有一定的差异;聚类分析将5个群体聚为三类。     

Different methods of selecting animals for genotyping to maximize the amount of genetic information known in the population

It is possible to predict genotypes of some individuals based on genotypes of relatives. Different methods of sampling individuals to be genotyped from populations were evaluated using simulation. Simulated pedigrees included 5,000 animals and were assigned genotypes based on assumed allelic frequencies for a SNP (favorable/unfavorable) of 0.3/0.7, 0.5/0.5, and 0.8/0.2. A field data pedigree (29,101 animals) and a research pedigree (8,688 animals) were used to test selected methods using simulated genotypes with allelic frequencies of 0.3/0.7 and 0.5/0.5. For the simulated pedigrees, known and unknown allelic frequencies were assumed. The methods used included random sampling, selection of males, and selection of both sexes based on the diagonal element of the inverse of the relationship matrix (A(-1)) and absorption of either the A or A(-1) matrix. For random sampling, scenarios included selection of 5 and 15% of the animals, and all other methods presented concentrated on the selection of 5% of the animals for genotyping. The methods were evaluated based on the percentage of alleles correctly assigned after peeling (AK(P)), the probability of assigning true alleles (AK(G)), and the average probability of correctly assigning the true genotype. As expected, random sampling was the least desirable method. The most desirable method in the simulated pedigrees was selecting both males and females based on their diagonal element of A(-1). Increases in AK(P) and AK(G) ranged from 26.58 to 29.11% and 2.76 to 6.08%, respectively, when males and females (equal to 5% of all animals) were selected based on their diagonal element of A(-1) compared with selecting 15% of the animals at random. In the case of a real beef cattle pedigree, selection of males only or males and females yielded similar results and both selection methods were superior to random selection.     

Mutations in bone morphogenetic protein 15 and growth differentiation factor 9 genes are associated with increased litter size in fat-tailed sheep breeds

The objective of this study was to investigate association between GDF9 and BMP15 gene polymorphism and litter size in fat-tailed sheep, a total of 97 mature ewes from four breeds (Afshari=19; Baluchi=18; Makui=30 and Mehraban=30) were genotyped for the BMP15 HinfI and GDF9 HhaI polymorphisms by PCR-RFLP technique. The highest and lowest mutant allele frequencies were found in Makui (0.27) and Afshari (0.10) sheep for the BMP15 gene and in Afshari (0.24) and Mehraban (0.18) sheep for the GDF9 gene, respectively. Litter size was significantly influenced by genotype of the ewe for two genes (P < 0.01). Heterozygous genotypes for both loci showed higher litter size than homozygous genotypes (P < 0.01). None of the individuals carried homozygous genotype for both of the GDF9 and BMP15 variants in these breeds. The individuals carrying the mutant allele for one of the investigated candidate gene still showed fertile phenotype. Thus, existence of homozygosity at one of the BMP15 and GDF9 variant is not probably able to block normal hormonal pathway of reproduction in fat-tailed sheep.     

Marker‐assisted selection reduces expected inbreeding but can result in large effects of hitchhiking

We used computer simulations to investigate to what extent true inbreeding, i.e. identity‐by‐descent, is affected by the use of marker‐assisted selection (MAS) relative to traditional best linear unbiased predictions (BLUP) selection. The effect was studied by varying the heritability (h² = 0.04 vs. 0.25), the marker distance (MAS vs. selection on the gene, GAS), the favourable QTL allele effect (α = 0.118 vs. 0.236) and the initial frequency of the favourable QTL allele (p = 0.01 vs. 0.1) in a population resembling the breeding nucleus of a dairy cattle population. The simulated genome consisted of two chromosomes of 100 cM each in addition to a polygenic component. On chromosome 1, a biallelic QTL as well as 4 markers were simulated in linkage disequilibrium. Chromosome 2 was selectively neutral. The results showed that, while reducing pedigree estimated inbreeding, MAS and GAS did not always reduce true inbreeding at the QTL relative to BLUP. MAS and GAS differs from BLUP by increasing the weight on Mendelian sampling terms and thereby lowering inbreeding, while increasing the fixation rate of the favourable QTL allele and thereby increasing inbreeding. The total outcome in terms of inbreeding at the QTL depends on the balance between these two effects. In addition, as a result of hitchhiking, MAS results in extra inbreeding in the region surrounding QTL, which could affect the overall genomic inbreeding.