Verification of yield QTL through realized molecular marker- assisted selection responses in a barley cross

Verification of putative quantitative trait loci (QTL) is an essential step towards implementing the use of marker-assisted selection (MAS) in cultivar improvement. In a previous study with 150 doubled haploid lines derived from the 6-row cross Steptoe/Morex (S/M), four regions (QTL1–4) of the barley genome were associated with differential genotypic expression for grain yield across environments. The objectives of this study were to verify the value of these four QTL for selection and to compare the efficiency of alternative MAS strategies using these QTL vs. conventional phenotypic selection for grain yield. A total of 92 DHLs derived from the S/M cross that were not used in the original mapping efforts were used for QTL verification. Confirmation of QTL effects was first accomplished by assessing yield differences between individuals carrying alternative alleles at each putative locus in three environments. QTL1 on chromosome 3 was confirmed as the most important and consistent locus to determine yield across sites, with the S allele being favorable. The M allele at QTL3 on chromosome 6 was beneficial for grain yield across sites, but to a lesser degree than QTL1. Magnitudes of allele effects at QTL2 (chromosome 2) and QTL4 (chromosome 7) were highly influenced by the environment where the genotypes were grown. Verification of QTL effects was best achieved by comparing realized selection response. Genotypic (MAS) and tandem genotypic and phenotypic selection were at least as good as phenotypic selection. Consistent selection responses were detected for QTL1 alone and together with QTL3. Genotypic selection for lines carrying the S allele at QTL1 resulted in the identification of high-yielding genotypes. Selection responses increased when the M allele at QTL3 was combined with the S allele at QTL1. Significant qualitative QTL × environment interactions for QTL2 and QTL4 were detected through differential realized selection responses at different sites. Without a thorough understanding of the physiological and agronomic particulars of any QTL and the target environment, MAS for QTL showing qualitative interactions should be minimized This revised version was published online in June 2006 with corrections to the Cover Date.     

Single large-scale marker-assisted selection (SLS-MAS)

This paper presents a new approach for plant improvement that interactively combines the use of DNA markers and conventional breeding. This approach involves selecting plants at early generation with a fixed, favorable genetic background at specific loci, conducting a single large-scale marker-assisted selection (SLS-MAS) while maintaining as much as possible the allelic segregation in the rest of the genome. First, the identification of elite lines presenting high allelic complementarity and being outstanding for traits of interest is required to capture favorable alleles from different parental lines. Second, after identification of the most favorable genomic regions for each selected parental line, those lines are intercrossed to develop segregating populations from which plants homozygous for favorable alleles at target loci are selected. One objective of the scheme is to conduct the marker-assisted selection only once, and it requires the selection of a minimum number of plants to maintain sufficient allelic variability at the unselected loci. Therefore, the selection pressure exerted on the segregating population is quite high and the screening of large populations is required to achieve the objectives of the scheme. No selection is applied outside the target genomic regions, to maintain as much as possible the Mendelian allelic segregation among the selected genotypes. After selection with DNA markers, the genetic diversity at un-selected loci may allow breeders to generate new varieties and hybrids through conventional breeding in response to various local needs. Although the single large-scale MAS scheme described here is oriented toward maize and large-scale breeding programs with substantial resources, the flexibility of this scheme would allow breeding programs to develop options compatible with local resources.     

Molecular marker-assisted selection for enhanced yield in malting barley

Brewers are reluctant to change malting barley (Hordeum vulgare ssp. vulgare L.) cultivars due to concerns of altered flavor and brewing procedures. The U.S. Pacific Northwest is capable of producing high yielding, high quality malting barley but lacks adapted cultivars with desirable malting characteristics. Our goal was to develop high yielding near isogenic lines that maintain traditional malting quality characteristics by transferring quantitative trait loci (QTL) associated with yield, via molecular marker-assisted backcrossing, from the high yielding cv. Baronesse to the North American two-row malting barley industry standard cv. Harrington. For transfer, we targeted Baronesse chromosome 2HL and 3HL fragments presumed to contain QTL that affect yield. Analysis of genotype and yield data suggests that QTL reside at two regions, one on 2HL (ABG461C-MWG699) and one on 3HL (MWG571A-MWG961). Genotype and yield data indicate that additional Baronesse genome regions are probably involved, but need to be more precisely defined. Based on yield trials conducted over 22 environments and malting analyses from 6 environments, we selected one isogenic line (00-170) that has consistently produced yields equal to Baronesse while maintaining a Harrington-like malting quality profile. We conclude there is sufficient data to warrant experiments testing whether the 2HL and 3HL Baronesse QTL would be effective in increasing the yield of other low yielding barley cultivars.     

The conversion of RFLP markers to allele specific amplicons linked to QTLs governing malting quality in barley

Application of marker-assisted selection with RFLP based markers has been constrained by high cost and time requirements in situations involving a large number of plants. RFLP markers mapped on a Harrington/TR306 population have been identified elsewhere as linked to quantitative trait loci (QTL) governing malting quality. The probes ABG610, ABC622, as well as probes for the Nar1, Amy1 and Nar7 were sequenced and locus specific primers developed. These locus specific primers were applied to genomic DNA from both Harrington and TR306. Sequence analysis of the resultant monomorphic fragments revealed sequence divergence for the Xabg610, Xabc622, Amy1 and Nar1 loci, but not for the Nar7 locus. Application of a set of Hor2 primers to genomic DNA from the barley lines Harrington and TR306 led to the direct amplification of codominant alleles. Allele-specific primers were designed based on the sequence divergence identified among the Xabg610, Xabc622 and Nar1 alleles. Amplification conditions were optimized for each of these alleles such that only the favourable allele from Harrington was amplified. The usefulness of these primers for selecting Harrington alleles was demonstrated by their failure to amplify the corresponding alleles from the lines, Sterling, Stella and WM872. The Amy1 allele-specific amplicon was only capable of differentiating this locus between Harrington and TR306. The conversion of these markers into PCR amplifiable, allele-specific amplicons would greatly facilitate their application to barley breeding programs.     

Studies on the patterns of nitrogen uptake and translocation to the grain of winter barley intended for malting

In five experiments carried out between 1987 and 1989, crops of winter barley were grown under a range of agronomic treatments to study the way the treatments influenced the patterns of nitrogen uptake, dry matter production, translocation of assimilates and malting quality. Rate and timing of nitrogen fertiliser had large effects. Increased and late applications invariably increased nitrogen concentration in the grain by affecting the partitioning of nitrogen between the vegetative and reproductive parts of the plant. Foliar fungal diseases also affected nitrogen partitioning, apparently by interfering with the transport of stored nitrogen to the developing grain. Seasonal influences, especially rainfall during the grain filling period, were found to have profound effects on the pattern of change in ear and grain nitrogen concentration, and hence malting quality. The ability to predict final grain nitrogen concentration from measurements made earlier in the season is not therefore likely to be feasible.     

Accuracy of marker-assisted selection with auxiliary traits

Genetic information on molecular markers is increasingly being used in plant and animal improvement programmes particularly as indirect means to improve a metric trait by selection either on an individual basis or on the basis of an index incorporating such information. This paper examines the utility of an index of selection that not only combines phenotypic and molecular information on the trait under improvement but also combines similar information on one or more auxiliary traits. The accuracy of such a selection procedure has been theoretically studied for sufficiently large populations so that the effects of detected quantitative trait loci can be perfectly estimated. The theory is illustrated numerically by considering one auxiliary trait. It is shown that the use of an auxiliary trait improves the selection accuracy; and, hence, the relative efficiency of index selection compared to individual selection which is based on the same intensity of selection. This is particularly so for higher magnitudes of residual genetic correlation and environmental correlation having opposite signs, lower values of the proportion of genetic variation in the main trait associated with the markers, negligible proportion of genetic variation in the auxiliary trait associated with the markers, and lower values of the heritability of the main trait but higher values of the heritability of the auxiliary trait.     

Simultaneous marker-assisted selection for multiple traits in autogamous crops

A method is presented for the selection of parents with the aim of obtaining improved genotypes in the progeny of a cross. The procedure is designed to select in several unrelated traits simultaneously and is based on the selection of molecular markers that are linked to QTLs. The method was compared with conventional phenotypic selection in simulation experiments for a number of genetic structures underlying the traits and several types of parental populations. Although the method in general provides good results, some of the underlying assumptions may be violated quite easily, thereby reducing the applicability of the procedure in practice. Received: 10 September 1999 / Accepted: 24 August 2000     

Optimal selection on two quantitative trait loci with linkage

A mathematical approach to optimize selection on multiple quantitative trait loci (QTL) and an estimate of residual polygenic effects was applied to selection on two linked or unlinked additive QTL. Strategies to maximize total or cumulative discounted response over ten generations were compared to standard QTL selection on the sum of breeding values for the QTL and an estimated breeding value for polygenes, and to phenotypic selection. Optimal selection resulted in greater response to selection than standard QTL or phenotypic selection. Tight linkage between the QTL (recombination rate 0.05) resulted in a slightly lower response for standard QTL and phenotypic selection but in a greater response for optimal selection. Optimal selection capitalized on linkage by emphasizing selection on favorable haplotypes. When the objective was to maximize total response after ten generations and QTL were unlinked, optimal selection increased QTL frequencies to fixation in a near linear manner. When starting frequencies were equal for the two QTL, equal emphasis was given to each QTL, regardless of the difference in effects of the QTL and regardless of the linkage, but the emphasis given to each of the two QTL was not additive. These results demonstrate the ability of optimal selection to capitalize on information on the complex genetic basis of quantitative traits that is forthcoming.     

QTL analysis of malting quality in barley based on the doubled-haploid progeny of two elite North American varieties representing different germplasm groups

Characterization of the determinants of economically important phenotypes showing complex inheritance should lead to the more effective use of genetic resources. This study was conducted to determine the number, genome location and effects of QTLs determining malting quality in the two North American barley quality standards. Using a doubled-haploid population of 140 lines from the cross of Harrington×Morex, malting quality phenotype data sets from eight environments, and a 107-marker linkage map, QTL analyses were performed using simple interval mapping and simplified composite interval mapping procedures. Seventeen QTLs were associated with seven grain and malting quality traits (percentage of plump kernels, test weight, grain protein percentage, soluble/total protein ratio, α-amylase activity, diastatic power and malt-extract percentage). QTLs for multiple traits were coincident. The loci controlling inflorescence type [vrs1 on chromosome 2(2H) and int-c on chromosome 4(4H)] were coincident with QTLs affecting all traits except malt-extract percentage. The largest effect QTLs, for the percentage of plump kernels, test weight protein percentage, S/T ratio and diastatic power, were coincident with the vrs1 locus. QTL analyses were conducted separately for each sub-population (six-rowed and two-rowed). Eleven new QTLs were detected in the subpopulations. There were significant interactions between the vrs1 and int-c loci for grain-protein percentage and S/T protein ratio. Results suggest that this mating of two different germplasm groups caused a disruption of the balance of traits. Information on the number, position and effects of QTLs determining components of malting quality may be useful for maintaining specific allele configurations that determine target quality profiles. Received: 28 May 1999 / Accepted: 9 November 1999     

A cautiously optimistic vision for marker-assisted breeding

Even though marker-assisted selection now plays a prominent role in the field of plant breeding, examples of successful, practical outcomes are rare. It is clear that DNA markers hold great promise, but realizing that promise remains elusive. Despite innovations like better marker systems and improved genetic mapping strategies, most marker associations are not sufficiently robust for successful marker-assisted selection. In large part this is due to inadequate experimental design. Molecular breeders must reassess their research programs so that DNA marker work leads to useful selection tools and valuable germplasm. As molecular breeders adopt more rigorous experimental guidelines and ambitious goals, they also need to integrate the growing body of knowledge from genomics and bioinformatics.     

Ultra-simple DNA extraction method for marker-assisted selection using microsatellite markers in rice

We prevent an ultra-simple DNA extraction method for microsatellite analysis of rice. Each extraction requires only one microtube, one disposable pipette tip, TE buffer and few pieces (about 5 mm) of rice leaf tissue. This is sufficient for 200 PCR reactions. The extract can be kept in the freezer for long-term storage. Also, DNA can be extracted from 200–300 individuals in a few hours. These features enabled us to perform rapid largescale seedling genotyping required for marker-assisted selection. We have also examined the applicability of this method for other PCR-based markers: RAPDs, nuclear STS, chloroplast STS and chloroplast microsatellites.     

Validation of growth-related quantitative trait loci markers in different Exopalaemon carinicauda families for marker-assisted selection

We detected growth-related QTL and associated markers from the backcross population of Exopalaemon carinicauda in the previous study. Based on our previous study, the 47 SNP markers associated with candidate growth trait QTL were selected to analyze the association between these markers and body weight (BW), body length and abdominal segment length traits in four different populations including wild population, a full-sib family, a half-sib family and a backcross population for evaluating their potential application of marker-assisted selection in E. carinicauda. The general linear model (GLM) and mixed linear model were applied and the associations between SNP loci and three growth-related traits verified. The results showed that the Marker79268 and Marker100644 were significantly associated with the BW trait in more than three populations by the GLM method. The Marker100644 was significantly associated with BW in the full-sib family, half-sib family and backcross populations by the GLM and mixed linear model methods. Our findings will provide useful SNP markers to go forward to improve growth performance through more refined marker-assisted selection in E. carinicauda.     

Within-family marker-assisted selection for aquaculture species

A within-family marker-assisted selection scheme was designed for typical aquaculture breeding schemes, where most traits are recorded on sibs of the candidates. Here, sibs of candidates were tested for the trait and genotyped to establish genetic marker effects on the trait. BLUP breeding values were calculated, including information of the markers (MAS) or not (NONMAS). These breeding values were identical for all family members in the NONMAS schemes, but differed between family members in the MAS schemes, making within-family selection possible. MAS had up to twice the total genetic gain of the corresponding NONMAS scheme. MAS was somewhat less effective when heritability increased from 0.06 to 0.12 or when the frequency of the positive allele was < 0.5. The relative efficiency of MAS was higher for schemes with more candidates, because of larger fullsib family sizes. MAS was also more efficient when male:female mating ratio changed from 1:1 to 1:5 or when the QTL explained more of the total genetic variation. Four instead of two markers linked to the QTL increased genetic gain somewhat. There was no significant difference in polygenic genetic gain between MAS and NONMAS for most schemes. The rates of inbreeding were lower for MAS than NON-MAS schemes, because fewer full-sibs were selected by MAS.     

Detection of quantitative trait loci for meat quality traits in cattle

A whole-genome scan was carried out to detect quantitative trait loci (QTL) affecting sensory, organoleptic, physical and chemical properties of meat. The study used phenotypic data from 235 second-generation cross-bred bull calves of a Charolais × Holstein experimental population. Loin muscle samples were evaluated for yield force, intramuscular fat and nitrogen contents, myofibrillar fragmentation index, haem pigment concentration, moisture content and pH at 24 h postmortem. A sensory assessment was performed on grilled loin and roasted silverside joints by trained panellists. A linear regression analysis based on 165 markers revealed 35 QTL at the 5% chromosome-wide significance level (20 for sensory traits and 15 for physical and chemical traits), five of which were highly significant ( F -value: ≥9). The most significant QTL was located on chromosome 6 (with the best likely position at 39 cM) and affected haem pigment concentration. The Holstein allele for this QTL was associated with an increase of 0.53 SD in the haem scores. A QTL for pH_24h was identified on chromosome 14 (at 40 cM) and a QTL for moisture content was identified on chromosome 22 (at 21 cM). Two highly significant QTL were identified for sensory panel-assessed traits: beef odour intensity (grilled sample) on chromosome 10 (at 119 cM), and juiciness (roast sample) on chromosome 16 (at 70 cM). The proportion of phenotypic variance explained by the significant QTL ranged from 3.6% (for nitrogen content on chromosome 10) to 9.5% (for juiciness, roast sample on chromosome 16).     

Mapping QTL controlling yield and yield components in a spring barley (Hordeum vulgare L.) cross using marker regression

An RFLP map constructed from 99 doubled haploid lines of a cross between two spring barley varieties (Blenheim × Kym) was used to localize quantitative trait loci (QTL) controlling grain yield and yield components by marker regression and single-marker analysis. Trials were conducted over three years. Genotype-by-year interaction was detected for plant grain weight and ear grain weight so they were analysed separately for each year. None was detected for thousand-grain weight and ear grain number so data were pooled over years. A total of eleven QTL were detected for plant grain weight over two years and fourteen for ear grain weight over three years. Seven QTL were detected for plot yield. The locus with the largest effect was on chromosome 2(2H)L and accounted for 19% of the variation in the progeny. Eight QTL were detected for thousand-grain weight and five for ear grain number. Many of the QTL detected were in comparable positions in each year. Yield and yield components were only partly correlated. Comparisons based on common RFLP markers showed that some QTL were found in positions similar to those identified in other studies. For a number of QTL the identification of linked markers provided suitable opportunities for marker-assisted selection and improvement of barley and reference markers with which to analyse the homoeologous chromosome regions of wheat and other cereals.     

Recent insights into barley and Rhynchosporium commune interactions

Rhynchosporium commune is the causal pathogen of scald in barley (Hordeum vulgare), a foliar disease that can reduce yield by up to 40% in susceptible cultivars. R. commune is found worldwide in all temperate growing regions and is regarded as one of the most economically important barley pathogens. It is a polycyclic pathogen with the ability to rapidly evolve new virulent strains in response to resistance genes deployed in commercial cultivars. Hence, introgression and pyramiding of different loci for resistance (qualitative or quantitative) through marker-assisted selection is an effective way to improve scald resistance in barley. This review summarizes all 148 resistance quantitative trait loci reported at the date of submission of this review and projects them onto the barley physical map, where it is clear many loci co-locate on chromosomes 3H and 7H. We have summarized the major named resistance loci and reiterated the renaming of Rrs15 (CI8288) to Rrs17. This review provides a comprehensive resource for future discovery and breeding efforts of qualitative and quantitative scald resistance loci.     

Improvement of malting quality of barley by complementing the malt enzyme spectrum

The processing quality of cereals can be modified by altering the structural grain constituents or the enzyme activities that mobilize storage reserves of the seeds. In order to complement the malt enzyme spectrum, a gene encoding for a thermotolerant fungal endo-(1,4)--glucanase was introduced into two barley cultivars, Kymppi and Golden Promise. The gene was expressed in the seeds during germination, thus providing a thermotolerant enzyme that is active under mashing conditions. The amount of thermotolerant -glucanase produced by the seeds (ca. 0.025% soluble seed protein) has been shown to be sufficient to reduce wort viscosity by decreasing the soluble -glucan content. For the safe commercial cultivation of transgenic plants risk assessment of their cultivation is needed. In our study experimental estimates of the transgene flow from transgenic barley by pollen dispersal were produced. Field trials were conducted during the summers of 1996 and 1997. A transgenic barley line homozygous for the gene encoding for neomycin phosphotransferase was used as a source of pollen and male-sterile barley lines as recipients. In order to be able to transform the cross-fertilization frequencies to corresponding values of normal male-fertile barley, plots of normal barley were also included in the experimental plan. On the basis of our study, cross-fertilization in male-sterile recipient barley is possible with very low frequency up to 50 meters from the donor area. However, the frequency dramatically decreases with distance and due to self-pollination the possibility of cross-fertilization remains very low in normal cultivated barley.     

Verification of barley seed dormancy loci via linked molecular markers

Seed dormancy is a relatively complex trait in barley (Hordeum vulgare L.). Several dormancy loci were identified previously by quantitative trait locus analysis. Three reciprocal crosses were made in the present study between parents carrying specific dormancy alleles via linked molecular markers to verify individual dormancy locus effects and potential expression. Analyses of F₂ progenies revealed that the dormancy allele at the locus flanked by the markers Ale and ABC302 on the long arm of chromosome 7 had a major effect on dormancy, and was at least partly epistatic to the dormancy locus in the ABC309-MWG851 interval near the telomere of the long arm of chromosome 7. In the absence of the dormancy allele in the Ale-ABC302 interval, the allele in the ABC309-MWG851 interval exerted moderate to large effects on dormancy. Cytoplasmic effects on dormancy were also observed. The germination percentages of progeny with relatively high levels of dormancy were more variable than those of non-dormant or less-dormant progeny, apparently due to environmental effects. Removal of the dormancy allele in the Ale-ABC302 interval, or introducing the dormancy allele in the ABC309-MWG851 interval, should suffice for adjusting dormancy levels in breeding programs to suit various production situations and end uses. The verification of dormancy loci via linked molecular markers allows manipulation of these loci in applied breeding programs.     

Advances to improve the eating and cooking qualities of rice by marker-assisted breeding

The eating and cooking qualities of rice are heavily emphasized in breeding programs because they determine market values and they are the appealing attributes sought by consumers. Conventional breeding has developed traditional varieties with improved eating and cooking qualities. Recently, intensive genetic studies have pinpointed the genes that control eating and cooking quality traits. Advances in genetic studies have developed molecular techniques, thereby allowing marker-assisted breeding (MAB) for improved eating and cooking qualities in rice. MAB has gained the attention of rice breeders for the advantages it can offer that conventional breeding cannot. There have been successful cases of using MAB to improve the eating and cooking qualities in rice over the years. Nevertheless, MAB should be applied cautiously given the intensive effort needed for genotyping. Perspectives from conventional breeding to marker-assisted breeding will be discussed in this review for the advancement of the eating and cooking qualities of fragrance, amylose content (AC), gel consistency (GC) and gelatinization temperature (GT) in rice. These four parameters are associated with eating and cooking qualities in rice. The genetic basis of these four parameters is also included in this review. MAB is another approach to rice variety improvement and development in addition to being an alternative to genetic engineering. The MAB approach shortens the varietal development time, and is therefore able to deliver improved rice varieties to farmers within a shorter period of time.     

Application of nontargeted metabolite profiling to discover novel markers of quality traits in an advanced population of malting barley

The process of breeding superior varieties for the agricultural industry is lengthy and expensive. Plant metabolites may act as markers of quality traits, potentially expediting the appraisal of experimental lines during breeding. Here, we evaluated the utility of metabolites as markers by assessing metabolic variation influenced by genetic and environmental factors in an advanced breeding setting and in relation to the phenotypic distribution of 20 quality traits. Nontargeted liquid chromatography–mass spectrometry metabolite profiling was performed on barley (Hordeum vulgare L.) grain and malt from 72 advanced malting barley lines grown at two distinct but climatically similar locations, with 2‐row and 6‐row barley as the main genetic factors. 27 420 molecular features were detected, and the metabolite and quality trait profiles were similarly influenced by genotype and environment; however, malt was more influenced by genotype compared with barley. An O2PLS model characterized molecular features and quality traits that covaried, and 1319 features associated with at least one of 20 quality traits. An indiscriminant MS/MS acquisition and novel data analysis method facilitated the identification of metabolites. The analysis described 216 primary and secondary metabolites that correlated with multiple quality traits and included amines, amino acids, alkaloids, polyphenolics and lipids. The mechanisms governing quality trait–metabolite associations were interpreted based on colocalization to genetic markers and their gene annotations. The results of this study support the hypothesis that metabolism and quality traits are co‐influenced by relatively narrow genetic and environmental factors and illustrate the utility of grain metabolites as functional markers of quality traits.