Identification of novel QTL contributing resistance to aflatoxin accumulation in maize

The toxic metabolic product aflatoxin produced by the opportunistic fungus Aspergillus flavus (Link:Fr) in maize (Zea mays L.) can cause disease and economic harm when levels exceed very minute quantities. The selection of resistant germplasm has great potential to reduce the problem, but the highly quantitative nature of the trait makes this a difficult endeavor. The identification of aflatoxin accumulation resistance quantitative trait loci (QTL) from resistant donor lines and the discovery of linked markers could speed this task. To identify marker–trait associations for marker-assisted breeding, a genetic mapping population of F_2:3 families was developed from Mp715, a maize inbred line resistant to aflatoxin accumulation, and T173, a susceptible, southern-adapted maize inbred line. QTL, some with large phenotypic effects, were identified in multiple years on chromosomes 1, 3, 5, and 10, and smaller QTL identified in only 1 year were found on chromosomes 4 and 9. The phenotypic effect of each QTL ranged from 2.7 to 18.5%, and models created with multiple QTL could explain up to 45.7% of the phenotypic variation across years, indicating that the variation associated with the trait can be manipulated using molecular markers.     

Confirming quantitative trait loci for aflatoxin resistance from Mp313E in different genetic backgrounds

The fungus Aspergillus flavus (Link:Fr) causes ear rot of maize (Zea mays L.) and produces the toxic metabolic product aflatoxin. One particularly effective method of controlling the fungus is via host plant resistance, but while several resistant breeding lines have been identified, transferring the resistance genes from these lines into elite cultivars has been less effective than needed. A high number of genes involved with resistance, each with a small effect, and some only found under certain environmental conditions, has hampered resistance breeding. The identification of markers linked to genomic regions associated with resistance would aid in this effort. The goals of this study were to identify and characterize quantitative trait loci (QTL) conferring resistance to aflatoxin accumulation from resistant maize donor Mp313E in a background of the susceptible inbred line Va35; to compare them to the QTL identified from Mp313E in a background of B73; and to test the stability of the QTL identified in Mp313E × Va35 in multiple environments by remapping the phenotypic tails of the Mp313E × Va35 mapping population in new locations. Twenty different QTL were found in this study, 11 of which were also found in different environments using the phenotypic tail subset mapping population, and five of which were likely the same as those reported in the Mp313E × B73 mapping population. This indicates that many of the QTL are stable over the environments and genetic backgrounds tested, which will make them more valuable in breeding efforts.     

Identification of quantitative trait loci controlling resistance to maize chlorotic dwarf virus

Ineffective screening methods and low levels of disease resistance have hampered genetic analysis of maize (Zea mays L.) resistance to disease caused by maize chlorotic dwarf virus (MCDV). Progeny from a cross between the highly resistant maize inbred line Oh1VI and the susceptible inbred line Va35 were evaluated for MCDV symptoms after multiple virus inoculations, using the viral vector Graminella nigrifrons. Symptom severity scores from three rating dates were used to calculate area under the disease progress curve (AUDPC) scores for vein banding, leaf twist and tear, and whorl chlorosis. AUDPC scores for the F₂ population indicated that MCDV resistance was quantitatively inherited. Genotypic and phenotypic analyses of 314 F₂ individuals were compared using composite interval mapping (CIM) and analysis of variance. CIM identified two major quantitative trait loci (QTL) on chromosomes 3 and 10 and two minor QTL on chromosomes 4 and 6. Resistance was additive, with alleles from Oh1VI at the loci on chromosomes 3 and 10 contributing equally to resistance.     

Identification of quantitative trait loci controlling resistance to gray leaf spot disease in maize

Breeding maize for gray leaf spot (GLS) resistance has been hindered by the quantitative nature of the inheritance of GLS resistance and by the limitations of selection under less than optimumal disease pressure. In order to identify the quantitative trait loci (QTLs) controlling GLS resistance, a cross was made between B73 (susceptible) and Va14 (resistant) to generate a large F₂ population. Six GLS disease assessments were made throughout the disease season for over 1000 F₂ plants in 1989, and for 600 F₂-derived F₃ lines replicated in two blocks in 1990. RFLP analysis for78 marker loci representing all ten maize chromosomes was conducted in 239 F₂ individuals including those with the extreme GLS disease phenotypes. The GLS disease scores of the three field evaluations, each averaged over six ratings, were separately used for the interval mapping in order to determine the consistency of the QTL effects. The heavy GLS disease pressure, meticulous disease ratings, and large population size of this study afforded us the sensitivity for detecting QTL effects. QTLs located on three chromosomes (1, 4, and 8) had large effects on GLS resistance, each explaining 35.0–56.0%, 8.8–14.3%, and 7.7–11.0% of the variance, respectively. These three QTL effects were remarkably consistent across three disease evaluations over 2 years and two generations. Smaller QTL effects were also found on chromosomes 2 and 5, but the chromosome-5 effect might be a false positive because it was not repeatable even in the same location. The chromosome-1 QTLs had the largest effect or highest R² reported for any quantitative trait to-date. Except for the chromosome-4 gene, which was from the susceptible parent B73, the resistance alleles at all QTL were derived from Va14. The resistance QTLs on chromosomes 1 and 2 appear to have additive effects, but those on chromosomes 4 and 8 are dominant and recessive, respectively. Significant interaction between the QTLs on chromosomes 1 and 4 was detected in all three evaluations. Cumulatively, the four QTLs identified in this study explained 44, 60, and 68% of the variance in F₂, and in F₃ replications 1 and 2, respectively.     

Identification of quantitative trait loci that affect endoreduplication in maize endosperm

Endoreduplication in maize endosperm precedes the onset of starch and storage protein synthesis, and it is generally thought to influence grain filling. We created four backcross populations by reciprocally crossing the F₁ progeny of a cross between Sg18 and Mo17 to the parental inbreds, which differ in endoreduplication by two parameters—mean ploidy and percentage of endoreduplicated nuclei. This four-backcross design allowed us to estimate and test the additive and dominant genetic effects of quantitative trait loci (QTLs) affecting endoreduplication. An analysis of endosperm from the four backcross populations at 16 days after pollination using a modified triploid mapping approach identified three endosperm QTLs influencing mean ploidy and two endosperm QTLs affecting the percentage of endoreduplicated nuclei. Some of these QTLs may manifest their effects on endoreduplication via expression in the embryo. The QTLs detected display strong dominance or over-dominance and interacted epistatically with an embryo-expressed QTL. This helps to explain the genetic basis for transgressive segregation in the backcross progeny. Although the favorable alleles that increase mean ploidy and percentage of endoreduplicated nuclei can be contributed by both parents, the Mo17-derived alleles for endoreduplication were often dominant or over-dominant to the Sg18-derived allele. One QTL on chromosome 7 that may be expressed in both the embryo and endosperm exerted a pleiotropic effect on two different parameters of endoreduplication. The results from this study shed light on the regulation of endoreduplication in maize endosperm and provide a marker-assisted selection strategy for potentially improving grain yield. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. C. M. Coelho and S. Wu contributed equally to this work and should be considered as first authors.     

Identification of quantitative trait loci for nitrogen use efficiency in maize

Intensively managed crop systems are normally dependent on nitrogen input to maximize yield potential. Improvements in nitrogen- use efficiency (NUE) in crop plants may support the development of cropping systems that are more economically efficient and environment friendly. The objective of this study was to map and characterize quantitative trait loci (QTL) for NUE in a maize population. In preliminary experiments, inbred lines contrasting for NUE were identified and were used to generate populations of F2:3 families for genetic study. A total of 214 F2:3 families were evaluated in replicated trials under high nitrogen (280 kg/ha) and low nitrogen (30 kg/ha) conditions in 1996 and 1997. Analysis of ear-leaf area, plant height, grain yield, ears per plant, kernels number per ear, and kernel weight indicated significant genetic variation among F2:3 families. The heritability of these traits was found to be high (h2=0.57–0.81). The mapping population were genotyped using a set of 99 restriction fragment length polymorphism (RFLP) markers. A linkage map of these markers was developed and used to identify QTL. Between two and six loci were found to be associated with each trait. The correspondence of several genomic regions with traits measured under nitrogen limited conditions suggests the presence of QTL associated with NUE. QTLs will help breeders to improve their maize ideotype of a low-nitrogen efficiency by identifying those constitutive and adaptive traits involved in the expression of traits significantly correlated with yield, such as ear leaf area and number of ears per plant. This revised version was published online in June 2006 with corrections to the Cover Date.     

Identification of quantitative trait loci linked to Ceratocystis wilt resistance in cacao

Ceratocystis wilt (CW) in cacao (Theobroma cacao L.), caused by Ceratocystis cacaofunesta, is a drastic disease that results in plant death. The pathogen was recently identified in the major cacao-producing region of Brazil?CBahia. The identification of genetic markers tightly linked to disease resistance loci is a valuable tool for the development of resistant cultivars using marker-assisted selection (MAS). Branches of 143 six-year-old individuals of an F2 Sca 6?×?ICS 1 population were wounded by making a 3-mm deep cut with a sterile scalpel, and inoculated with a 20-??l drop of a spore suspension of 3?×?10⁴?CFU/ml. The inoculation method used allowed the population to be quantitatively phenotyped. The length of the xylem discoloration followed a continuous distribution. These results imply that the resistance was quantitatively inherited. Quantitative trait loci (QTL) analysis revealed two genomic regions (in linkage groups 3 and 9) associated with CW resistance. The QTL explained individually from 6.9 to 8.6?% of the phenotypic variation. The QTL identified are crucial for identifying genes for resistance and can be applied in the genetic breeding of cacao using MAS.     

Breeding for resistance to aflatoxin accumulation in maize

Contamination of maize,Zea mays, grain with aflatoxin, a naturally occurring toxin produced byAspergillus flavus, frequently reduces the value and marketability of maize produced in the southern USA. Drought, high temperatures, and insect damage are often associated with high levels of maize aflatoxin contamination. Growing resistant maize hybrids is generally considered the most feasible method of reducing or eliminatingA. flavus infection and subsequent accumulation of aflatoxin. Developing appropriate screening techniques and identifying maize germplasm with resistance to aflatoxin contamination provides the foundation for a breeding program. Only a few sources of aflatoxin resistance have been identified. Four germplasm lines (Mp313E, Mp420, Mp715, and Mp717) have been developed and released by USDA-ARS at Mississippi State University. NC 388, developed at North Carolina State University, is reported as another putative source of aflatoxin resistance. Conventional phenotypic selection was used to successfully combine resistance to aflatoxin contamination from two of these lines, Mp313E and Mp715, with desirable agronomic qualities from Va35. The identification of quantitative trait loci (QTL) associated with resistance to aflatoxin contamination will also permit the use of marker assisted selection in transferring resistance into elite germplasm lines. Development of parental inbreds that combine aflatoxin resistance with superior agronomic quality is an essential component of a hybrid maize breeding program designed to reduce or eliminate aflatoxin contamination.     

Identification of quantitative trait loci for cadmium tolerance and accumulation in wheat

Quantitative trait loci (QTL) for Cadmium (Cd) tolerance and accumulation in wheat (Triticum aestivum L.) were identified, using 103 recombinant inbred lines (RILs) derived from a cross of Ch × Sh at germination and seedling stages. The traits of germination, growth and physiology were measured. Cd tolerance indexes (TI) were calculated for plants under Cd stress relative to control conditions. Cd concentrations in both root and shoot were determined and the amount of Cd accumulation and translocation calculated. The phenotypic variation of the above traits showed a continuous distribution pattern among the RILs. Twenty-six QTLs were detected, (16 of which were designated for the traits under the control and Cd stress, 8 for Cd tolerance and 2 for root Cd accumulation). These 26 QTLs individually could explain 7.97–60.16% of the phenotypic variation. Fourteen QTLs were positive (with the additive effects coming from Ch) while the remaining 12 QTLs were negative (with the additive effects contributed by Sh). No QTL were detected in the same region on the chromosomes of wheat. The results indicated that genetic mechanisms controlling the traits of Cd tolerance were independent from each other. Therefore, in this study, the properties of Cd tolerance and accumulation showed to be independent traits in wheat.     

Identification of quantitative trait loci for race-nonspecific resistance to tan spot in wheat

Tan spot, caused by Pyrenophora tritici-repentis (Ptr), is an economically important foliar disease in the major wheat growing areas throughout the world. Multiple races of the pathogen have been characterized based on their ability to cause necrosis and/or chlorosis on differential wheat lines. In this research, we evaluated a population of recombinant inbred lines derived from a cross between the common wheat varieties Grandin and BR34 for reaction to tan spot caused by Ptr races 1–3 and 5. Composite interval mapping revealed QTLs on the short arm of chromosome 1B and the long arm of chromosome 3B that were significantly associated with resistance to all four races. The effects of the two QTLs varied for the different races. The 1B QTL explained from 13% to 29% of the phenotypic variation, whereas the 3B QTL explained from 13% to 41% of the variation. Additional minor QTLs were detected but not associated with resistance to all races. The host-selective toxin Ptr ToxA, which is produced by races 1 and 2, was not a significant factor in the development of disease in this population. The race-nonspecific resistance derived from BR34 may take precedence over the gene-for-gene interaction known to be associated with the wheat–Ptr system.     

Detection and verification of quantitative trait loci for resistance to Fusarium ear rot in maize

Fusarium ear rot caused by Fusarium verticillioides is a prevalent disease in maize which can severely reduce grain yields and quality. Identification of stable quantitative trait loci (QTL) for resistance to Fusarium ear rot is a basic prerequisite for understanding the genetic mechanism of resistance and for the use of marker-assisted selection. In this study, two hundred and ten F _2:3 families were developed from a cross between resistant inbred line BT-1 and susceptible inbred line Xi502, and were genotyped with 178 simple sequence repeat markers. The resistance of each line was evaluated in two environments by artificial inoculation using the nail-punch method. The resistance QTL were detected using the composite interval mapping method. Three QTL were detected on chromosomes 4, 5 and 10. Of them, the QTL on chromosome 4 (bin 4.05/06) had the largest resistance to Fusarium ear rot, and could explain 17.95?% of the phenotypic variation. For further verification of the QTL effect, we developed near-isogenic lines (NILs) carrying the QTL region on chromosome 4 using parental line Xi502 as the recurrent parent. In the NIL background, this QTL can increase the resistance by 33.7?C35.2?% if the resistance region is homozygous, and by 17.8?C26.5?% if the resistance region contains the heterozygous allele. The stable and significant resistance effect of the QTL on chromosome 4 lays the foundation for further marker-assisted selection and map-based cloning in maize.     

Mapping quantitative trait loci (QTLs) for resistance to Gibberella zeae infection in maize

The basic prerequisite for an efficient breeding program to improve levels of resistance to pathogens in plants is the identification of genes controlling the resistance character. If the response to pathogens is under the control of a multilocus system, the utilization of molecular markers becomes essential. Stalk and ear rot caused by Gibberella zeae is a widespread disease of corn: resistance to G. zeae is quantitatively inherited. Our experimental approach to understanding the genetic basis of resistance to Gibberella is to estimate the genetic linkage between available molecular markers and the character, measured as the amount of diseased tissue 40 days after inoculation of a suspension of Fusarium graminearum, the conidial form of G. zeae, into the first stalk internode. Sensitive and resistant parental inbreds were crossed to obtain F₁ and F₂ populations: the analysis of the segregation of 95 RFLP (restriction fragment length polymorphism) clones and 10 RAPD (random amplified polymorphic DNA) markers was performed on a population of 150 F₂ individuals. Analysis of resistance was performed on the F₃ families obtained by selfing the F₂ plants. Quantitative trait loci (QTL) detection was based either on analysis of regression coefficients between family mean value and allele values in the F₂ population, or by means of interval mapping, using MAPMAKER-QTL. A linkage map of maize was obtained, in which four to five genomic regions are shown to carry factors involved in the resistance to G. zeae.     

Identification of quantitative trait loci contributing to Fusarium wilt resistance on an AFLP linkage map of flax (Linum usitatissimum)

 An AFLP genetic linkage map of flax (Linum usitatissimum) was used to identify two quantitative trait loci (QTLs) on independent linkage groups with a major effect on resistance to Fusarium wilt, a serious disease caused by the soil pathogen Fusarium oxysporum (lini). The linkage map was constructed using a mapping population from doubled-haploid (DH) lines. The DH lines were derived from the haploid component of F₂ haploid-diploid twin seed originating from a cross between a polyembryonic, low-linolenic-acid genotype (CRZY8/RA91) and the Australian cultivar ‘Glenelg’. The AFLP technique was employed to generate 213 marker loci covering approximately 1400 cM of the flax genome (n=15) with an average spacing of 10 cM and comprising 18 linkage groups. Sixty AFLP markers (28%) deviated significantly (P<0.05) from the expected segregation ratio. The map incorporated RFLP markers tightly linked to flax rust (Melamspora lini) resistance genes and markers detected by disease resistance gene-like sequences. The study illustrates the potential of the AFLP technique as a robust and rapid method to generate moderately saturated linkage maps, thereby allowing the molecular analysis of traits, such as resistance to Fusarium wilt, that show oligogenic patterns of inheritance. Received: 8 December 1997 / Accepted: 7 April 1998     

In silico mapping of quantitative trait loci in maize

Quantitative trait loci (QTL) are most often detected through designed mapping experiments. An alternative approach is in silico mapping, whereby genes are detected using existing phenotypic and genomic databases. We explored the usefulness of in silico mapping via a mixed-model approach in maize (Zea mays L.). Specifically, our objective was to determine if the procedure gave results that were repeatable across populations. Multilocation data were obtained from the 1995–2002 hybrid testing program of Limagrain Genetics in Europe. Nine heterotic patterns comprised 22,774 single crosses. These single crosses were made from 1,266 inbreds that had data for 96 simple sequence repeat (SSR) markers. By a mixed-model approach, we estimated the general combining ability effects associated with marker alleles in each heterotic pattern. The numbers of marker loci with significant effects—37 for plant height, 24 for smut [Ustilago maydis (DC.) Cda.] resistance, and 44 for grain moisture—were consistent with previous results from designed mapping experiments. Each trait had many loci with small effects and few loci with large effects. For smut resistance, a marker in bin 8.05 on chromosome 8 had a significant effect in seven (out of a maximum of 18) instances. For this major QTL, the maximum effect of an allele substitution ranged from 5.4% to 41.9%, with an average of 22.0%. We conclude that in silico mapping via a mixed-model approach can detect associations that are repeatable across different populations. We speculate that in silico mapping will be more useful for gene discovery than for selection in plant breeding programs.     

Genetic analysis of two new quantitative trait loci for ear weight in maize inbred line Huangzao4

Ear weight is one of the most important agronomic traits considered necessary in maize (Zea mays L.) breeding projects. To determine its genetic basis, a population consisting of 239 recombinant inbred lines, derived from the cross Mo17 x Huangzao4, was used to detect quantitative trait loci (QTLs) for ear weight under two nitrogen regimes. Under a high nitrogen fertilization regime, one QTL was identified in chromosome bin 2.08-2.09, which explained 7.46% of phenotypic variance and an increase in ear weight of about 5.79 g, owing to an additive effect. Under a low nitrogen regime, another QTL was identified in chromosome bin 1.10-1.11; it accounted for 7.11% of phenotypic variance and a decrease of 5.24 g in ear weight, due to an additive effect. Based on comparisons with previous studies, these two QTLs are new loci associated with ear weight in maize. These findings contribute to our knowledge about the genetic basis of ear weight in maize.     

Identification of quantitative trait loci associated with resistance to cucumber mosaic virus in Capsicum annuum

QTL analysis for resistance to cucumber mosaic virus (CMV) was performed in an intraspecific Capsicum annuum population. A total of 180 F3 families were derived from a cross between the susceptible bell-type cultivar Maor and the resistant small-fruited Indian line Perennial and inoculated with CMV in three experiments carried out in the USA and Israel using two virus isolates. Mostly RFLP and AFLP markers were used to construct the genetic map, and interval analysis was used for QTL detection. Four QTL were significantly associated with resistance to CMV. Two digenic interactions involving markers with and without an individual effect on CMV resistance were also detected. The QTL controlling the largest percentage (16–33%) of the observed phenotypic variation (cmv11.1) was detected in all three experiments and was also involved in one of the digenic interactions. This QTL is linked to the L locus that confers resistance to tobacco mosaic virus (TMV), confirming earlier anecdotal observations of an association between resistance to CMV and susceptibility to TMV in Perennial. An advanced backcross breeding line from an unrelated population, 3990, selected for resistance to CMV was analyzed for markers covering the genome, allowing the identification of genomic regions introgressed from Perennial. Four of these introgressions included regions associated with QTL for CMV resistance. Markers in two genomic regions that were identified as linked to QTL for CMV resistance were also linked to QTL for fruit weight, confirming additional breeding observations of an association between resistance to CMV originating from Perennial and small fruit weight. Received: 17 July 2000 / Accepted: 16 October 2000     

Identification of molecular markers linked to quantitative trait loci for soybean resistance to corn earworm

 One hundred and thirty nine restriction fragment length polymorphisms (RFLPs) were used to construct a soybean (Glycine max L. Merr.) genetic linkage map and to identify quantitative trait loci (QTLs) associated with resistance to corn earworm (Helicoverpa zea Boddie) in a population of 103 F₂-derived lines from a cross of ‘Cobb’ (susceptible) and PI229358 (resistant). The genetic linkage map consisted of 128 markers which converged onto 30 linkage groups covering approximately 1325 cM. There were 11 unlinked markers. The F₂-derived lines and the two parents were grown in the field under a plastic mesh cage near Athens, Ga., in 1995. The plants were artificially infested with corn earworm and evaluated for the amount of defoliation. Using interval-mapping analysis for linked markers and single-factor analysis of variance (ANOVA), markers were tested for an association with resistance. One major and two minor QTLs for resistance were identified in this population. The PI229358 allele contributed insect resistance at all three QTLs. The major QTL is linked to the RFLP marker A584 on linkage group (LG) ‘M’ of the USDA/Iowa State University public soybean genetic map. It accounts for 37% of the total variation for resistance in this cross. The minor QTLs are linked to the RFLP markers R249 (LG ‘H’) and Bng047 (LG ‘D1’). These markers explain 16% and 10% of variation, respectively. The heritability (h²) for resistance was estimated as 64% in this population. Received: 15 October 1997 / Accepted: 4 November 1997     

Identification of quantitative trait loci for resistance to powdery mildew in a Spanish barley landrace

The Spanish landrace-derived inbred line SBCC97, together with other lines from the Spanish Barley Core Collection, displays high resistance to powdery mildew, caused by the fungus Blumeria graminis f. sp. hordei. The objective of this study was to map quantitative trait loci (QTLs) for resistance to powdery mildew in a recombinant inbred line population derived from a cross between SBCC97 and the susceptible cultivar ‘Plaisant’. Phenotypic analysis was performed using four B. graminis isolates, and genetic maps were constructed with mainly simple sequence repeat (SSR) markers, following a sequential genotyping strategy. Two major QTLs with large effects were identified on chromosome 7H, and they accounted for up to 45% of the total phenotypic variance. The alleles for resistance at each QTL were contributed by the Spanish parent SBCC97. One locus was mapped to the short arm of chromosome 7HS, and was flanked by the resistance gene analogue (RGA) marker S9202 and the SSR GBM1060. This corresponded to the same chromosomal region in which a major race-specific resistance gene from Hordeum vulgare ssp. spontaneum, designated as mlt, had been identified previously. The second QTL was linked tightly to marker EBmac0755, and it shared its chromosomal location with the qualitative resistance gene Mlf, which has only been described previously in the wild ancestor H. spontaneum. This is the first report of these two QTLs occurring together in cultivated barley, and it paves the way for their use in barley breeding programs that are designed to transfer resistance alleles into elite cultivars.