From QTL to QTN identification in livestock--winning by points rather than knock-out: a review

Many quantitative trait loci (QTL) affecting economic traits in livestock have now been identified. However, the confidence interval (CI) of individual QTL as determined by linkage analysis often spans tens of map units, containing hundreds of genes. Linkage disequilibrium (LD) mapping can reduce the CI to individual map units, but this reduced interval will still contain tens of genes. Methods suitable for model animals to find and validate specific quantitative trait nucleotides (QTN) underlying the QTL cannot be easily applied to livestock species because of their long generation intervals, the cost of maintaining each animal and the difficulty of producing transgenics or 'knock-outs'. Considering these limitations, we review successful approaches for identifying QTN in livestock and outline a schematic strategy for QTN determination and verification. In addition to linkage and LD mapping, the methods include positional cloning, selection of candidate genes, DNA sequencing and statistical analyses. Concordance determination and functional assays are the critical tests for validation of a QTN; we provide a generalized formula for the probability of concordance by chance. Three genes that meet the burden of proof for QTN identification--DGAT1 in cattle, IGF2 in swine and GDF8 in sheep--are discussed in detail. The genetic and economic ramifications of identified QTN and the horizon for selection and introgression are also considered.     

Genetic mapping of quantitative trait loci for resistance to Haemonchus contortus in sheep

This paper presents results from a mapping experiment to detect quantitative trait loci (QTL) for resistance to Haemonchus contortus infestation in merino sheep. The primary trait analysed was faecal worm egg count in response to artificial challenge at 6 months of age. In the first stage of the experiment, whole genome linkage analysis was used for broad-scale mapping. The animal resource used was a designed flock comprising 571 individuals from four half-sib families. The average marker spacing was about 20 cM. For the primary trait, 11 QTL (as chromosomal/family combinations) were significant at the 5% chromosome-wide level, with allelic substitution effects of between 0.19 and 0.38 phenotypic standard deviation units. In general, these QTL did not have a significant effect on faecal worm egg count recorded at 13 months of age. In the second stage of the experiment, three promising regions (located on chromosomes 1, 3 and 4) were fine-mapped. This involved typing more closely spaced markers on individuals from the designed flock as well as an additional 495 individuals selected from a related population with a deeper pedigree. Analysis was performed using a linkage disequilibrium–linkage approach, under additive, dominant and multiple QTL models. Of these, the multiple QTL model resulted in the most refined QTL positions, with resolutions of <10 cM achieved for two regions. Because of the moderate size of effect of the QTL, and the apparent age and/or immune status specificity of the QTL, it is suggested that a panel of QTL will be required for significant genetic gains to be achieved within industry via marker-assisted selection.     

Additive and Over-dominant Effects Resulting from Epistatic Loci Are the Primary Genetic Basis of Heterosis in Rice

A set of 148 F9 recombinant inbred lines （RILs） was developed from the cross of an indica cultivar 93-11 and japonica cultivar DTI13, showing strong F1 heterosis. Subsequently, two backcross F1 （BCFI） populations were constructed by backcrossing these 148 RILs to two parents, 93-11 and DT713. These three related populations （281BCF1 lines, 148 RILs） were phenotyped for six yield-related traits in two locations. Significant inbreeding depression was detected in the population of RILS and a high level of heterosis was observed in the two BCF1 populations. A total of 42 main-effect quantitative trait loci （M-QTLs） and 109 epistatic effect QTL pairs （E-QTLs） were detected in the three related populations using the mixed model approach. By comparing the genetic effects of these QTLs detected in the RILs, BCF1 performance and mid-parental heterosis （HMp）, we found that, in both BCF1 populations, the QTLs detected could be classified into two predominant types： additive and over-dominant loci, which indicated that the additive and over-dominant effect were more important than complete or partially dominance for M-QTLs and E-QTLs. Further, we found that the E-QTLs detected collectively explained a larger portion of the total phenotypic variation than the M-QTLs in both RILs and BCF1 populations. All of these results suggest that additive and over-dominance resulting from epistatic loci might be the primary genetic basis of heterosis in rice.     

Genetic variation, population structure, and linkage disequilibrium in European elite germplasm of perennial ryegrass

Perennial ryegrass (Lolium perenne L.) is a highly valued temperate climate grass species grown as forage crop and for amenity uses. Due to its outbreeding nature and recent domestication, a high degree of genetic diversity is expected among cultivars. The aim of this study was to assess the extent of linkage disequilibrium (LD) within European elite germplasm and to evaluate the appropriate methodology for genetic association mapping in perennial ryegrass. A high level of genetic diversity was observed in a set of 380 perennial ryegrass elite genotypes when genotyped with 40 SSRs and 2 STS markers. A Bayesian structure analysis identified two subpopulations, which were confirmed by principal coordinate analysis (PCoA). One subpopulation consisted mainly of genotypes originating from the UK, while germplasm mostly from Continental Europe was grouped into the second subpopulation. LD (r²) decay was rapid and occurred within 0.4 cM across European varieties, when population structure was taken into consideration. However, an extended LD of up to 6.6 cM was detected within the variety Aberdart. High genetic diversity and rapid LD decay provide means for high resolution association mapping in elite materials of perennial ryegrass. However, different strategies need to be applied depending on the material used. Genome-wide association study (GWAS) with several hundred markers can be applied within synthetic varieties to identify large (up to 10 cM) genomic regions affecting trait variation. A combination of available and novel DNA markers is needed to achieve resolution required for GWAS in elite breeding materials. An even higher marker density of several million SNPs might be needed for GWAS in diverse ecotype collections, potentially resulting in quantitative trait polymorphism (QTP) identification.     

Genetic analysis of extended life span in Drosophila melanogaster II. Replication of the backcross test and molecular characterization of the N14 Locus

We are interested in localizing chromosomal regions that extend life span in Drosophila. Using stocks artificially selected for long life by Luckinbill and his colleagues, we have identified marker loci that are highly divergent in allelic frequencies between replicated long-lived lines and controls (Curtsinger et al., 1998). Several of the most divergent loci have been found to be associated with effects on life span in segregating backcross populations. Here we report an independent replication of the backcross test for the N14 marker locus, previously reported to extend male life spans by 12 days. The life span effect successfully replicates in males. N14 accounts for 30% of the total selection response in males. Life span extension occurs by a decrease in age-specific mortality rates at all ages, and is not attributable to modification of the slope of the age-specific mortality curve. The effect in females is small or nonexistent. Sequencing of the N14 locus shows that it is non-coding and not obviously regulatory, suggesting that the phenotypic effect arises from linkage disequilibrium with another locus or loci that directly affect life span. N14 DNA hybridizes to 63F/64A on the left arm of chromosome 3. The location is consistent with previous whole-chromosome substitution studies, and suggests new candidate genes for life span extension in Drosophila, including ras2. This revised version was published online in July 2006 with corrections to the Cover Date.     

Genetic mapping of monoterpene composition in an interspecific eucalypt hybrid

Genetic control of foliar oil composition was investigated amongst half-sib progeny of an interspecific eucalypt hybrid. The oil was found to be largely composed of the monoterpenes, limonene, α−pinene, γ−terpinene, 1,8 cineole and p-cymene. Due to difficulties in the interpretation of the compositional data based on raw proportions, further analysis was conducted using log-ratio variables. A high degree of intercorrelation amongst log-ratios was thought to be a consequence of commonality in the biosynthetic origins of the monoterpenes. Quantitative trait locus (QTL) analysis of log-ratio variables indicated that a significant (68–81%) proportion of the variation in four out of the ten possible log-ratios were controlled by a single genomic region of the maternal Eucalyptus grandis parent. The impact of this genomic region upon oil composition was thought to be a consequence of a gene, or genes, controlling the production of limonene, as limonene was the predominant oil constituent in many hybrid individuals and was common to all log-ratios associated with the identified genomic region. Received: 20 November 1998 / Accepted: 16 June 1999     

水稻对叶瘟和穗瘟部分抗性的遗传分析

在一个水稻籼籼交重组自交系群体中,选用由感病株系构成的2个亚群体和2个不同的稻瘟病菌小种,进行了水稻对叶瘟部分抗性的QTL定位,还选用由感病而且抽穗期相近的株系构成的亚群体和另一个病菌小种,进行了水稻对穗瘟部分抗性的QTL定位,将病叶面积百分比(DLA)、病斑大小(LS)和病斑数(LN)作为对叶瘟部分抗性的性状,将病斑长度(LL)和孢子量(CA)作为对穗瘟部分抗性的性状。所构建的图谱包含168个标记。应用QTLMapper 1．01b,共检测到11个表现主效应的QTL和28对双因子互作,有3个表现主效应的QTL参与对同一性状的互作。QTL的主效应对单一性状的贡献率为4．7％～38．8％,而上位性效应对单一性状的贡献率为16．0％～51．7％,QTL的主效应对大多数性状的贡献率小于互作效应,表明互作效应对于部分抗性的重要作用。对穗瘟部分抗性的两个性状LL和CA,所检测到QTL总效应的贡献率分别达到70．6％和82．6％,表明由排除了主效抗病基因的感病株系组成的亚群体适合于进行部分抗性QTL定位。     

Increased Vulnerability to 3-Nitropropionic Acid in an Animal Model of Huntington's Disease

Abstract: There is substantial evidence for both metabolic dysfunction and oxidative damage in Huntington's disease (HD). In the present study, we used in vivo microdialysis to measure the conversion of 4-hydroxybenzoic acid to 3,4-dihydroxybenzoic acid (3,4-DHBA) as a measure of hydroxyl radical production in a transgenic mouse model of HD, as well as in littermate controls. The conversion of 4-hydroxybenzoic acid to 3,4-DHBA was unchanged in the striatum of transgenic HD mice at baseline. Following administration of the mitochondrial toxin 3-nitropropionic acid (3-NP), there were significant increases in 3,4-DHBA generation in both control and transgenic HD mice, and the increases in the transgenic HD mice were significantly greater than those in controls. Furthermore, administration of 3-NP produced significantly larger striatal lesions in transgenic HD mice than in littermate controls. The present results show increased sensitivity to the mitochondrial toxin 3-NP in transgenic HD mice, which suggests metabolic dysfunction in this mouse model of HD.     

Toward positional cloning of Vgt1, a QTL controlling the transition from the vegetative to the reproductive phase in maize

Vgt1 (Vegetative to generative transition 1) is a quantitative trait locus (QTL) for flowering time in maize (Zea mays L.). Vgt1 was initially mapped in a ca. 5-cM interval on chromosome bin 8.05, using a set of near-isogenic lines (NILs) in the genetic background of the late dent line N28, with the earliness allele introgressed from the early variety Gaspé Flint. A new large mapping population was produced by crossing N28 and one early NIL with a ca. 6-cM long Gaspé Flint introgression at the Vgt1 region. Using PCR-based assays at markers flanking Vgt1, 69 segmental NILs homozygous for independent crossovers near the QTL were developed. When the NILs were tested in replicated field trials for days to pollen shed (DPS) and plant node number (ND), the QTL followed a Mendelian segregation. Using bulk segregant analysis and AFLP profiling, 17 AFLP markers linked to the QTL region were identified. Statistical analysis indicated a substantial coincidence of the effects of Vgt1 on both DPS and ND. Vgt1 was mapped at ca. 0.3 cM from an AFLP marker. As compared to DPS, the higher heritability of ND allowed for a more accurate assessment of the effects of Vgt1. The feasibility of the positional cloning of Vgt1 is discussed.     

Role of DAMPs and of Leukocytes Infiltration in Ischemic Stroke: Insights from Animal Models and Translation to the Human Disease

Stroke is a leading cause of death and disability worldwide. Several mechanisms are involved in the pathogenesis of ischemic stroke (IS). The contributory role of the inflammatory and immunity processes was demonstrated both in vitro and in animal models, and was confirmed in humans. IS evokes an immediate inflammatory response that involves complex cellular and molecular mechanisms. All components of the innate and adaptive immunity systems are involved in several steps of the ischemic cascade. In the early phase, inflammatory and immune mechanisms contribute to the brain tissue damage, whereas, in the late phase, they participate to the tissue repair processes. In particular, damage-associated molecular patterns (DAMPs) appear critical for the promotion of altered blood brain barrier permeability, leukocytes infiltration, tissue edema and brain injury. Conversely, the activation of regulatory T lymphocytes (Tregs) plays protective effects. The identification of specific cellular/molecular elements belonging to the inflammatory and immune responses, contributing to the brain ischemic injury and tissue remodeling, offers the advantage to design adequate therapeutic strategies. In this article, we will present an overview of the knowledge on inflammatory and immunity processes in IS, with a particular focus on the role of DAMPs and leukocytes infiltration. We will discuss evidence obtained in preclinical models of IS and in humans. The main molecular mechanisms useful for the development of novel therapeutic approaches will be highlighted. The translation of experimental findings to the human disease is still a difficult step to pursue. Further investigations are required to fill up the existing gaps.

     

The genetic basis of quantitative variation in susceptibility of Arabidopsis thaliana to Pseudomonas syringae (Pst DC3000): evidence for a new genetic factor of large effect

* Pathogens represent an important threat to plant communities and agriculture, and can shape many aspects of plant evolution. Natural variation in plant disease susceptibility is typically quantitative, yet studies on the molecular basis of disease resistance have focused mainly on qualitative variation. * Here we investigated the genetic architecture of quantitative susceptibility to the bacterium Pseudomonas syringae by performing a quantitative trait locus (QTL) analysis on the F2 progeny of two natural accessions of Arabidopsis thaliana under two nutrient treatments. * We found that a single QTL explains most of the variation (77%) in susceptibility between accessions Columbia (Col-0) and San Feliu-2 (Sf-2), and its effect is independent of nutrients. The Sf-2 allele at this QTL is dominant and can reduce the bacterial population size by 31-fold, much like a classical resistance (R) gene. However, minor QTLs, whose effects are altered by nutrient treatment, were also detected. * Surprisingly, we found that none of the QTLs for susceptibility had any effect on fruit production, suggesting that the use of resistance genes for crop improvement and evolutionary analysis of plant-pathogen interactions requires caution.     

Developmental analysis of teosinte glume architecture1: A key locus in the evolution of maize (Poaceae)

A key event in the evolution of maize from teosinte was a reduction in the cupulate fruitcase and softening of the glumes, which increased the accessibility of kernels for harvest. The teosinte glume architecture1 (tga1) locus largely controls this difference between maize and teosinte, and thus may have played a pivotal role in maize evolution. The teosinte allele (tga1+teosinte) lengthens inflorescence internodes, shortens rachillae, and makes glumes longer, thicker, and harder. Developmental characterization of morphometric traits reveals that differences among genotypes are apparent early in female inflorescence development. Increased hardening in glumes homozygous for tga1+teosinte is correlated with a thicker abaxial mesoderm of lignified cells. Silica deposition in the abaxial epidermal cells of the glumes is also affected. In the maize background, glumes homozygous for tga1+teosinte deposit silica in both the short and long cells of the glume epidermis, whereas glumes homozygous for the maize allele (Tga1+Maize) concentrate silica only in the short cells. Silica deposition also appears to be affected by genetic background. The effects of tga1 appear largely to explain the differences in glume induration between maize and teosinte. The diverse pleiotropic effects of tga1 suggest that it is regulatory in nature.     

From Genes to Ecosystems: The Genetic Basis of Condensed Tannins and Their Role in Nutrient Regulation in a Populus Model System

Research that connects ecosystem processes to genetic mechanisms has recently gained significant ground, yet actual studies that span the levels of organization from genes to ecosystems are extraordinarily rare. Utilizing foundation species from the genus Populus, in which the role of condensed tannins (CT) has been investigated aboveground, belowground, and in adjacent streams, we examine the diverse mechanisms for the expression of CT and the ecological consequences of CT for forests and streams. The wealth of data from this genus highlights the importance of form and function of CT in large-scale and long-term ecosystem processes and demonstrates the following four patterns: (1) plant-specific concentration of CT varies as much as fourfold among species and individual genotypes; (2) large within-plant variation in CT occurs due to ontogenetic stages (that is, juvenile and mature), tissue types (that is, leaves versus twigs) and phenotypic plasticity in response to the environment; (3) CT have little consistent effect on plant–herbivore interactions, excepting organisms utilizing woody tissues (that is, fungal endophytes and beaver), however; (4) CT in plants consistently slow rates of leaf litter decomposition (aquatic and terrestrial), alter the composition of heterotrophic soil communities (and some aquatic communities) and reduce nutrient availability in terrestrial ecosystems. Taken together, these data suggest that CT may play an underappreciated adaptive role in regulating nutrient dynamics in ecosystems. These results also demonstrate that a holistic perspective from genes-to-ecosystems is a powerful approach for elucidating complex ecological interactions and their evolutionary implications. All authors made significant contributions of data, research or writing to the study described in this review.     

Genetic variation and biochemical properties of esterase-18 (ES-18) in the laboratory rat (Rattus norvegicus): A new locus of esterase cluster 2 in linkage group V

A new liver-specific rat carboxylesterase isozyme (EC 3.1.1.1) designated esterase-18 (ES-18) is described. Genetic variation of ES-18 was examined in 93 inbred strains and substrains and a structural locusEs-18 was suggested, coding for either the presence (Es-18 ^a) or the absence (Es-18 ^b) of the isozyme. Linkage studies involving two backcross series revealed thatEs-18 resides in cluster 2 of LGV. No recombination betweenEs-18 and other cluster 2 loci was found in 19 lines of two RI strain sets or in the backcross series.R. K. was supported by the Sonderforschungsbereich 146 (Versuchstierforschung). O.D. was supported by the Deutsche Forschungsgemeinschaft (De 315/2). This is communication No. 65 of a research program devoted to the cellular distribution, regulation, and genetics of nonspecific esterases.     

Approaches to interval mapping of QTL in a multigeneration pedigree: the example of porcine chromosome 4

Quantitative trait loci (QTLs) have been mapped in many studies of F2 populations derived from crosses between diverse lines. One approach to confirming these effects and improving the mapping resolution is genetic chromosome dissection through a backcrossing programme. Analysis by interval mapping of the data generated is likely to provide additional power and resolution compared with treating data marker by marker. However, interval mapping approaches for such a programme are not well developed, especially where the founder lines were outbred. We explore alternative approaches to analysis using, as an example, data from chromosome 4 in an intercross between wild boar and Large White pigs where QTLs have been previously identified. A least squares interval mapping procedure was used to study growth rate and carcass traits in a subsequent second backcross generation (BC2). This procedure requires the probability of inheriting a wild boar allele for each BC2 animal for locations throughout the chromosome. Two methods for obtaining these probabilities were compared: stochastic or deterministic. The two methods gave similar probabilities for inheriting wild boar alleles and, hence, gave very similar results from the QTL analysis. The deterministic approach has the advantage of being much faster to run but requires specialized software. A QTL for fatness and for growth were confirmed and, in addition, a QTL for piglet growth from weaning at 5 weeks up to 7 weeks of age and another for carcass length were detected.     

A novel form of resistance in rice to the angiosperm parasite Striga hermonthica

The root hemiparasitic weed Striga hermonthica is a serious constraint to grain production of economically important cereals in sub-Saharan Africa. Breeding for parasite resistance in cereals is widely recognized as the most sustainable form of long-term control; however, advances have been limited owing to a lack of cereal germplasm demonstrating postattachment resistance to Striga. Here, we identify a cultivar of rice (Nipponbare) that exhibits strong postattachment resistance to S. hermonthica; the parasite penetrates the host root cortex but does not form parasite-host xylem-xylem connections. In order to identify the genomic regions contributing to this resistance, a mapping population of backcross inbred lines between the resistant (Nipponbare) and susceptible (Kasalath) parents were evaluated for resistance to S. hermonthica. Composite interval mapping located seven putative quantitative trait loci (QTL) explaining 31% of the overall phenotypic variance; a second, independent, screen confirmed four of these QTL. Relative to the parental lines, allelic substitutions at these QTL altered the phenotype by at least 0.5 of a phenotypic standard deviation. Thus, they should be regarded as major genes and are likely to be useful in breeding programmes to enhance host resistance.     

Ribosomal DNAs: an exception to the conservation of gene order in rice genomes

rDNA (18S-5.8S-25S rDNA) and 5S rDNA loci were visualized on the chromosomes of six species of the genus Oryza by fluorescence in situ hybridization (FISH) and the labeled rice chromosomes were identified based on their condensation patterns. As a result, the chromosomes harboring rDNA and/or 5S rDNA loci were determined in the complement for all the known rice genomes. Variation in the location of the rDNA loci indicated the transpositional nature of the rDNAs in the genus Oryza, as also suggested in Triticeae and Allium. Comparative analysis of the locations of rDNA loci among rice, maize and wheat revealed that variability in the physical location of the rDNA loci was characteristic of the genus Oryza and also of the genera of Gramineae. This variability in the location of the rDNA loci between evolutionarily related species is in sharp contrast to the conservation of the general order of genes in their genomes.     

Pathway-Related Factors: The Potential for Human Data to Improve the Scientific Basis of Risk Assessment

The default uncertainty factors used for risk assessment are applied either to allow for different aspects of extrapolation of the dose-response curve or to allow for database deficiencies. Replacement of toxicokinetic or toxicodynamics defaults by chemical-specific data allows the calculation of a chemical-specific “data-derived factor”, which is the product of chemical-specific values and default uncertainty factors. Such chemical-specific composite values will improve the scientific basis of the risk assessment of that chemical, but the necessary chemical-specific data are rarely available. Categorical defaults related to pathways of elimination and mechanisms of toxicity could be used when the overall fate or mechanism is known, but there are no chemical-specific data sufficient to allow replacement of the default, and the development of an overall data-derived factor. The development of pathway-related categorical defaults is being undertaken using data on selected probe substrates for which adequate data are available. The concept and difficulties of this approach are illustrated using data for CYP1A2.