Quantitative trait loci and candidate gene mapping of aluminum tolerance in diploid alfalfa

Aluminum (Al) toxicity in acid soils is a major limitation to the production of alfalfa (Medicago sativa subsp. sativa L.) in the USA. Developing Al-tolerant alfalfa cultivars is one approach to overcome this constraint. Accessions of wild diploid alfalfa (M. sativa subsp. coerulea) have been found to be a source of useful genes for Al tolerance. Previously, two genomic regions associated with Al tolerance were identified in this diploid species using restriction fragment length polymorphism (RFLP) markers and single marker analysis. This study was conducted to identify additional Al-tolerance quantitative trait loci (QTLs); to identify simple sequence repeat (SSR) markers that flank the previously identified QTLs; to map candidate genes associated with Al tolerance from other plant species; and to test for co-localization with mapped QTLs. A genetic linkage map was constructed using EST-SSR markers in a population of 130 BC₁F₁ plants derived from the cross between Al-sensitive and Al-tolerant genotypes. Three putative QTLs on linkage groups LG I, LG II and LG III, explaining 38, 16 and 27% of the phenotypic variation, respectively, were identified. Six candidate gene markers designed from Medicago truncatula ESTs that showed homology to known Al-tolerance genes identified in other plant species were placed on the QTL map. A marker designed from a candidate gene involved in malic acid release mapped near a marginally significant QTL (LOD 2.83) on LG I. The SSR markers flanking these QTLs will be useful for transferring them to cultivated alfalfa via marker-assisted selection and for pyramiding Al tolerance QTLs.     

Quantitative trait locus (QTL) analysis of wood quality traits in <Emphasis Type="Italic">Eucalyptus nitens</Emphasis>

To identify the chromosomal regions affecting wood quality traits, we conducted a genome-wide quantitative trait locus (QTL) analysis of wood quality traits in Eucalyptus nitens. This information is important to exploit the full potential of the impending Eucalyptus genome sequence. A three generational mapping population consisting of 296 progeny trees was used to identify QTL associated with several wood quality traits in E. nitens. Thirty-six QTL positions for cellulose content, pulp yield, lignin content, density, and microfibril angle (MFA) were identified across different linkage groups. On linkage groups (LG)2 and 8, cellulose QTL cluster with pulp yield and extractives QTL while on LG4 and 10 cellulose and pulp yield QTLs cluster together. Similarly, on LG4, 5, and 6 QTL for lignin traits were clustered together. At two positions, QTL for MFA, a physical trait related to wood stiffness, were clustered with QTL for lignin traits. Several cell wall candidate genes were co-located to QTL positions affecting different traits. Comparative QTL analysis with Eucalyptus globulus revealed two common QTL regions for cellulose and pulp yield. The QTL positions identified in this study provide a resource for identifying wood quality genes using the impending Eucalyptus genome sequence. Candidate genes identified in this study through co-location to QTL regions may be useful in association studies.     

Improving the quality of African robustas: QTLs for yield- and quality-related traits in Coffea canephora

Coffea canephora breeding requires combining sustainable productivity with improved technological and cup quality characteristics. Beverage quality is a complex and subjective trait, and breeding for this trait is time consuming and depends on knowledge of the genetics of its components. A highly variable C. canephora progeny resulting from an intraspecific cross was assessed for 63 traits over 5?years. To identify quantitative trait loci (QTLs) controlling agronomic, technological, and quality-related traits, a genetic map comprising 236 molecular markers was constructed, and composite interval mapping was performed. Beverage quality was evaluated in relation to biochemical and cup tasting traits. QTLs were identified for almost half of the traits evaluated, with effects ranging from 6% to 80% of phenotypic variation. Most of them present a consistent detection over years. The strongest QTLs explained a high percentage of the variation for yield in 2006 (34% to 57%), bean size (25% to 35%), content of chlorogenic acids (22% to 35%), sucrose and trigonelline content (29% to 81%), and acidity and bitterness of coffee beverages (30% to 55%). Regions of the C. canephora genome influencing beverage quality were identified. Five QTL zones were co-localized with candidate genes related to the biosynthesis of the analyzed traits: two genes coding for caffeine biosynthesis, one gene implicated in the biosynthesis of chlorogenic acids, and two genes implicated in sugar metabolism. This is one of the first studies on the identification of QTLs combining agronomic and quality traits in coffee. The high variability of quality traits within C. canephora and the presence of consistent QTLs offer breeders a promising tool to improve coffee cup quality.     

Identification of quantitative trait loci for yield and yield components in an advanced backcross population derived from the<Emphasis Type="Italic"> Oryza sativa</Emphasis> variety IR64 and the wild relative<Emphasis Type="Italic"> O. rufipogon</Emphasis>

A BC₂F₂ population developed from an interspecific cross between Oryza sativa (cv IR64) and O. rufipogon (IRGC 105491) was used in an advanced backcross QTL analysis to identify and introduce agronomically useful genes from this wild relative into the cultivated gene pool. The objectives of this study were: (1) to identify putative yield and yield component QTLs that can be useful to improve the elite cultivar IR64; (2) to compare the QTLs within this study with previously reported QTLs in rice as the basis for identifying QTLs that are stable across different environments and genetic backgrounds; and (3) to compare the identified QTLs with previously reported QTLs from maize to examine the degree of QTL conservation across the grass family. Two hundred eighty-five families were evaluated in two field environments in Indonesia, with two replications each, for 12 agronomic traits. A total of 165 markers consisting of 131 SSRs and 34 RFLPs were used to construct the genetic linkage map. By employing interval mapping and composite interval mapping, 42 QTLs were identified. Despite its inferior performance, 33% of the QTL alleles originating from O. rufipogon had a beneficial effect for yield and yield components in the IR64 background. Twenty-two QTLs (53.4%) were located in similar regions as previously reported rice QTLs, suggesting the existence of stable QTLs across genetic backgrounds and environments. Twenty QTLs (47.6%) were exclusively detected in this study, uncovering potentially novel alleles from the wild, some of which might improve the performance of the tropical indica variety IR64. Additionally, several QTLs for plant height, grain weight, and flowering time detected in this study corresponded to homeologous regions in maize containing previously detected maize QTLs for these traits.     

Quantitative trait loci affecting amylose,amylopectin and starch content in maize recombinant inbred lines

The loci explaining the variability of quantitative traits related to starch content and composition (amylose, amylopectin and water soluble fraction) were searched for in maize kernels. Multifactorial genetic methods were used to detect and locate QTLs (quantitative trait loci) on a genetic map consisting mainly of RFLP markers for genes with known function. The genetic material was recombinant inbred lines originating from parents differing in starch structure (dent vs. flint). Kernels were harvested from field grown plants for two successive years and under two pollination systems. Main effect and epistasis QTLs were detected using two methods, composite interval mapping (MQTL) and ANOVA. Despite large year-to-year differences, physiologically meaningful co-locations were observed between trait QTLs. Moreover, the number of expressed sequences on our map allowed the search for co-locations between QTLs and genes involved in carbohydrate metabolism. The main co-location was between an amylose QTL and Shrunken 2 (SH2) locus, on chromosome 3 (SH2 encoding for the large subunit of ADPglucose pyrophosphorylase). The importance of this locus as a candidate gene for a starch QTL is in agreement with previous studies based either on QTL co-locations or on revertant analysis. Other co-locations were observed between amylose and amylopectin QTLs and the two loci of IVR1 invertase genes on chromosomes 2 and 10. Further comparison with previously detected QTLs for carbohydrate metabolism in maize leaves showed consistent co-location in map regions devoid of candidate genes, such as near chromosome 1S telomere. The possible contribution of regulatory genes in this region is discussed.     

Association of gene-linked SSR markers to seed glucosinolate content in oilseed rape (Brassica napus ssp. napus)

Breeding of oilseed rape (Brassica napus ssp. napus) has evoked a strong bottleneck selection towards double-low (00) seed quality with zero erucic acid and low seed glucosinolate content. The resulting reduction of genetic variability in elite 00-quality oilseed rape is particularly relevant with regard to the development of genetically diverse heterotic pools for hybrid breeding. In contrast, B. napus genotypes containing high levels of erucic acid and seed glucosinolates (++ quality) represent a comparatively genetically divergent source of germplasm. Seed glucosinolate content is a complex quantitative trait, however, meaning that the introgression of novel germplasm from this gene pool requires recurrent backcrossing to avoid linkage drag for high glucosinolate content. Molecular markers for key low-glucosinolate alleles could potentially improve the selection process. The aim of this study was to identify potentially gene-linked markers for important seed glucosinolate loci via structure-based allele-trait association studies in genetically diverse B. napus genotypes. The analyses included a set of new simple-sequence repeat (SSR) markers whose orthologs in Arabidopsis thaliana are physically closely linked to promising candidate genes for glucosinolate biosynthesis. We found evidence that four genes involved in the biosynthesis of indole, aliphatic and aromatic glucosinolates might be associated with known quantitative trait loci for total seed glucosinolate content in B. napus. Markers linked to homoeologous loci of these genes in the paleopolyploid B. napus genome were found to be associated with a significant effect on the seed glucosinolate content. This example shows the potential of Arabidopsis-Brassica comparative genome analysis for synteny-based identification of gene-linked SSR markers that can potentially be used in marker-assisted selection for an important trait in oilseed rape. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

AFLP mapping and detection of quantitative trait loci (QTLs) for economically important traits in Pinus sylvestris: a preliminary study

We have applied a two-way pseudo-testcross strategy in an analysis of Pinus sylvestris for genetic mapping and detection of quantitative trait loci (QTLs) associated with economically important traits targeted in the Swedish tree-breeding programme. Based on 94 full-sib progeny of a cross between two plus-trees from northern Sweden we generated two parental maps using AFLP markers. The female map was comprised of 94 markers assigned to 15 linkage groups giving a size of 796 cM. On the male map 155 markers were assigned to 15 linkage groups, giving a total size of 1335 cM. The recombination frequency was found to be sex-dependent, being 29.3% higher in male than in female gametes. On the female map, 12 QTLs were detected (but none for branch diameter or wood density). Three QTLs for tree height accounted for 25.8% of the total phenotypic variation of this trait. When the QTLs detected for all the traits were taken independently, the percentages of phenotypic variance ranged from 9.3% to 22.7%. The highest value was observed for frost hardiness, an important trait in northern Sweden for which a major gene seemed to be involved. A cluster of QTLs for tree height, trunk diameter and volume was located on one linkage group. On the male map, four QTLs for trunk diameter and volume were detected. Due to the reduced number of individuals under study, the results are preliminary and have to be validated on more trees.     

Identification of genetic loci affecting the establishment and development of Echinococcus multilocularis larvae in mice

Alveolar echinococcosis (AE) is a severe hepatic disorder caused by larval infection by the fox tapeworm Echinococcus multilocularis. The course of parasitic development and host reactions are known to vary significantly among host species, and even among different inbred strains of mice. As reported previously, after oral administration of parasite eggs, DBA/2 (D2) mice showed a higher rate of cyst establishment and more advanced protoscolex development in the liver than C57BL/6 (B6) mice. These findings strongly suggest that the outcome of AE is affected by host genetic factor(s). In the present study, the genetic basis of such strain-specific differences in susceptibility/resistance to AE in murine models was studied by whole-genome scanning for quantitative trait loci (QTLs) using a backcross of (B6 × D2)F₁ and D2 mice with varying susceptibility to E. multilocularis infection. For cyst establishment, genome linkage analysis identified one suggestive and one significant QTL on chromosomes (Chrs.) 9 and 6, respectively, whereas for protoscolex development, two suggestive and one highly significant QTLs were detected on Chrs. 6, 17 and 1, respectively. Our QTL analyses using murine AE models revealed that multiple genetic factors regulated host susceptibility/resistance to E. multilocularis infection. Moreover, our findings show that establishment of the parasite cysts in the liver is affected by QTLs that are distinct from those associated with the subsequent protoscolex development of the parasite, indicating that different host factors are involved in the host–parasite interplay at each developmental stage of the larval parasite. Further identification of responsible genes located on the identified QTLs could lead to the development of effective disease prevention and control strategies, including an intensive screening and clinical follow-up of genetically high-risk groups for AE infection.     

Mapping Quantitative Trait Loci Affecting Variation in Drosophila Triacylglycerol Storage

Objective: Recent genetic studies indicate that Drosophila melanogaster could be a powerful model to identify genes involved in mammalian adipocyte differentiation and fat storage. The objective of our study was to identify quantitative trait loci (QTLs) that contribute to variation in triacylglycerol (TAG) storage in two D. melanogaster laboratory strains. Research Methods and Procedures: We used two genetic mapping procedures to identify loci with main and epistatic effects on TAG storage. First, using 68 recombinant inbred lines derived from the unrelated Oregon R and Russian 2b strains, we mapped the location of QTLs affecting TAG storage using both composite interval mapping and Bayesian epistatic methods. Second, we used the quantitative deficiency mapping procedure to identify candidate genes affecting this trait within one of the QTLs identified on the second chromosome. For both mapping experiments, flies were cultured in standard conditions. TAG content of 4‐ to 5‐day‐old flies, adjusted for live body mass and total proteins, was used as the phenotypic measure. Results: Multiple QTLs associated with variation in TAG storage were identified by the genome‐wide recombination mapping method, and some of them were sex‐specific. The QTLs had main effects, but a male‐specific epistatic interaction between two QTLs was also found. Finally, two closely linked QTLs were detected by deficiency mapping at 57E1‐57E3 and 57E4‐57F1 on chromosome 2, the first of which causes female‐specific variation in TAG between the Oregon R and 2b strains. Discussion: Our results suggest that variation in TAG storage in D. melanogaster is controlled by different genetic mechanisms and different sets of QTLs in male and female flies.     

Salt tolerance in Lycopersicon species. III. Detection of quantitative trait loci by means of molecular markers

A segregating population derived from a cross between L. esculentum cv Madrigal and a line of L. pimpinellifolium was used to identify genetic markers linked to QTLs involved in salinity tolerance in terms of yield, under a conductivity of 15 dS/m (171.1 mM NaCl). Six markers resulted, associated with QTLs affecting average fruit weight, fruit number and total weight under salinity. One of them, Aco-1, behaves reversely to the expectation from parental means; this and other features make it a promising target to obtain salt-tolerant tomatoes. Epistatic interactions were also found, thus affecting the criteria for marker-assisted selection. Although only 41% of the loci assayed were polymorphic, a high efficiency in identifying QTLs was achieved, since 43% of the marker loci are linked to QTLs for the trait under study.     

Nucleotide variability and gene expression reveal new putative genes related to seed shattering in weedy rice

Seed shattering is one of the main traits related with the domestication of cultivated rice and with the invasiveness and persistence of weedy rice. Two independent studies in 2006 have indicated that qSH1 in Japonica and Sh4 in Indica rice are major genes governing this trait. However, a wide variation of seed shattering occurs in weedy rice ecotypes from the same geographic region and even within the same ecotype. The aim of this study was to evaluate the nucleotide variability of known and putative genes related to seed shattering in cultivated rice and to identify and validate new genes related to this trait in weedy rice. The qSH1 gene was not associated with seed shattering in the evaluated genotypes. The nucleotide variability of the genes Os01g0849100 and Os08g0512400, previously identified based on a genome‐wide resequencing study, was related to seed shattering in rice. The nucleotide variability of three single nucleotide polymorphisms (SNPs) of the OsXTH8 gene, which is related to cell wall biosynthesis, was not associated with seed shattering. However, the high expression of this gene was related to the occurrence of this trait. This study evaluated jointly a series of genes involved in rice seed shattering and indicated that the genes OsXTH8, Os08g0512400 and Os01g0849100 are important for the regulation of this trait in weedy rice in addition to previously described genes. Seed shattering in weedy rice has a more complex regulation than in cultivated rice where few major genes were identified.     

Both stable and unstable QTLs for resistance to powdery mildew are detected in apple after four years of field assessments

Powdery mildew, caused by the ascomycete fungus Podosphaera leucotricha, is one of the most damaging diseases of apple worldwide. Polygenically determined resistance might contribute to a significant increase of resistance to this disease in new cultivars. A quantitative trait locus (QTL) analysis was performed in an F₁ progeny derived from a cross between the apple cultivar Discovery and the apple hybrid TN10-8. Powdery mildew incidence was assessed during four years (five seasons) in spring and/or autumn in a French local orchard. Seven additive and/or dominant QTLs were detected over the five seasons, with effects (R ²) ranging from 7.5% to 27.4% of the progeny phenotypic variation. Two QTLs, on linkage groups (LGs) 2 and 13, were consistently identified and accounted together from 29% to 37% of the phenotypic variation according to the year of assessment. The other QTLs were identified during one (LGs 1, 14), two (LG10), or three (LGs 8, 17) seasons. Their instability indicated a changing genetic determinism according to the year of assessment, for which several hypotheses may be put forward. The QTLs on LGs 2 and 8 mapped close to clusters of resistance gene analogs (RGAs) and major genes for resistance to mildew or apple scab previously identified. The stable QTLs identified on LGs 2 and 13, together with the strong effect QTL located on LG 8, are of special interest for breeding purposes, especially if combined with other major resistance genes.     

水稻蒸煮品质相关QTL定位及候选基因分析

挖掘与稻米蒸煮品质相关的数量性状基因座(quantitative trait locus, QTL)，分析候选基因，并通过遗传育种手段改良稻米蒸煮品质相关性状，可有效提升稻米的口感。以籼稻华占(Huazhan, HZ)、粳稻热研2号(Nekken2)及由其构建的120个重组自交系(recombinant inbred lines, RILs)群体为实验材料，测定成熟期稻米的糊化温度(gelatinization temperature, GT)、胶稠度(gel consistency, GC)和直链淀粉含量(amylose content, AC)。结合高密度分子遗传图谱进行QTL定位，共检测到26个与稻米蒸煮品质相关的QTLs (糊化温度相关位点1个、胶稠度相关位点13个、直链淀粉含量相关位点12个)，其中最高奇数的可能性(likelihood of odd, LOD)值达30.24。通过实时荧光定量PCR (quantitative real-time polymerase chain reaction, qRT-PCR)分析定位区间内候选基因的表达量，发现6个基因在双亲间的表达量差异显著，推测LOC_Os04g20270和LOC_Os11g40100的高表达可能会极大地提高稻米的胶稠度，而LOC_Os01g04920和LOC_Os02g17500的高表达以及LOC_Os03g02650和LOC_Os05g25840的低表达有助于降低直链淀粉含量。这些结果为培育优质水稻新品种奠定了分子基础，并为揭示稻米蒸煮品质的分子调控机制提供了重要的遗传资源。     

A high-density consensus map of barley to compare the distribution of QTLs for partial resistance to Puccinia hordei and of defence gene homologues

A consensus map of barley was constructed based on three reference doubled haploid (DH) populations and three recombinant inbred line (RIL) populations. Several sets of microsatellites were used as bridge markers in the integration of those populations previously genotyped with RFLP or with AFLP markers. Another set of 61 genic microsatellites was mapped for the first time using a newly developed fluorescent labelling strategy, referred to as A/T labelling. The final map contains 3,258 markers spanning 1,081 centiMorgans (cM) with an average distance between two adjacent loci of 0.33 cM. This is the highest density of markers reported for a barley genetic map to date. The consensus map was divided into 210 BINs of about 5 cM each in which were placed 19 quantitative trait loci (QTL) contributing to the partial resistance to barley leaf rust (Puccinia hordei Otth) in five of the integrated populations. Each parental barley combination segregated for different sets of QTLs, with only few QTLs shared by any pair of cultivars. Defence gene homologues (DGH) were identified by tBlastx homology to known genes involved in the defence of plants against microbial pathogens. Sixty-three DGHs were located into the 210 BINs in order to identify candidate genes responsible for the QTL effects. Eight BINs were co-occupied by a QTL and DGH(s). The positional candidates identified are receptor-like kinase, WIR1 homologues and several defence response genes like peroxidases, superoxide dismutase and thaumatin. Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

Molecular-marker-facilitated investigation of host-plant response to Exserohilum turcicum in maize (Zea mays L.): components of resistance

RFLPs were used to investigate components of host-plant response to Exserohilum turcicum in 150 unselected F₂₃ lines of a B52/Mo17 maize population. Following inoculation with spore suspensions of the pathogen (race 0), components of disease development were measured and then quantitative trait mapping was performed to identify the location and effects of quantitative trait loci (QTLs) determining host-plant response. Components of interest were the average number of lesions per leaf, the average percent leaf tissue diseased (severity) and the average size of lesions (cm²). Based on a LOD threshold of 2.31 (P<0.05), the number of lesions appears to be associated with QTLs on chromosomes 1S, 3L, 5S. Severity was associated with analogous regions and, in addition, QTLs on chromosomes 7L and 8L. Most QTLs, for either of these two components, involve additive gene action and partial dominance or overdominance. In contrast, lesion size was associated with QTLs on chromosomes 7L and 5L; recessive gene action may be involved at 7L.Journal Paper No. J-15178 of the Iowa Agriculture and Home Economic Experiment Station, Ames, Iowa. Project No. 3134     

Detection of grain protein content QTLs across environments in tetraploid wheats

Grain protein content (GPC) is an important quality factor in both durum and bread wheats. GPC is considered to be a polygenic trait influenced by environmental factors and management practice. The objectives of this study were both to compare the quantitative trait loci (QTL) for GPC in a population of 65 recombinant inbred lines of tetraploid wheats evaluated in three locations for several years (eight data sets), and to investigate the genetic relationship among GPC and grain yield. QTLs were determined based on the Messapia × dicoccoides linkage map which covers 217 linked loci on the 14 chromosomes with 42 additional loci as yet unassigned to linkage groups. The map extends to 1352 cM; the average distance between adjacent markers was 6.3 cM. Seven QTLs for GPC, located on the chromosome arms 4BS, 5AL, 6AS (two loci), 6BS, 7AS and 7BS, were detected that were significant in at least one environment at P<0.001 or in at least two environments at P<0.01. One QTL was significant in all but one environment, two were significant in four or five environments, and four were significant in two out of eight environments. Six out of seven protein content QTLs had pleiotropic effects or were associated to QTLs for grain yield and explained the negative correlation among GPC and yield components. The present results support the concept that studies conducted in a single environment are likely to underestimate the number of QTLs that can influence a trait and that the phenotypic data for a quantitative trait should be collected over a range of locations to identify putative QTLs and determine their phenotypic effects.     

Combined genetic and physiological analysis of a locus contributing to quantitative variation

The natural variation of many traits is controlled by multiple genes, individually referred to as quantitative trait loci (QTL), that interact with the environment to determine the ultimate phenotype of any individual. A QTL has yet to be described molecularly, in part because strategies to systematically identify them are underdeveloped and because the subtle nature of QTLs prevents the application of standard methods of gene identification. Therefore, it will be necessary to develop a systematic approach(es) for the identification of QTLs based upon the numerous positional data now being accumulated through molecular marker analyses. We have characterized a QTL by the following three-step approach: (1) identification of a QTL in complex populations, (2) isolation and genetic mapping of this QTL in near-isogenic lines, and (3) identification of a candidate gene using map position and physiological criteria. Using this approach we have characterized a plant height QTL in maize that maps to chromosome 9 near the centromere. Both map position and physiological criteria suggest the gibberillin biosynthesis gene dwarf3 as a candidate gene for this QTL.