B73 Allele Research Articles

A QTL on Maize Chromosome 5 is Associated with Root-Knot Nematode Resistance.

This study provides the first report of a quantitative trait locus (QTL) in maize (Zea mays) for resistance to the southern root-knot nematode (SRKN) (Meloidogyne incognita). The SRKN can feed on the roots of maize in the U.S. Southern Coastal Plain region and can cause yield losses of 30% or greater in heavily infested fields. Increases in SRKN density in the soil may reduce the yield for subsequently planted susceptible crops. The use of maize hybrids with resistance to SRKN could prevent an increase in SRKN density, yet no genetic regions have been identified that confer host resistance. In this study, a B73 (susceptible) x Ky21 (resistant) S5 recombinant inbred line (RIL) population was phenotyped for total number of eggs (TE) and root weight. This population has been previously genotyped using single nucleotide polymorphisms (SNPs). By utilizing the SNP data with the phenotype data, a single QTL was identified on chromosome 5 that explained 15% of the phenotypic variation (PV) for the number of eggs and 11% of the PV for the number of eggs per g of root (EGR). Plants that were homozygous for the Ky21 allele for the most associated marker PZA03172.3 had fewer eggs and fewer EGR than the plants that were homozygous or heterozygous for the B73 allele. Thus, the first QTL for SRKN resistance in maize has been identified and could be incorporated into maize hybrids.

The northern corn leaf blight resistance gene Ht1 encodes an nucleotide-binding, leucine-rich repeat immune receptor.

Northern corn leaf blight, caused by the fungal pathogen Exserohilum turcicum, is a major disease of maize. The first major locus conferring resistance to E.turcicum race 0, Ht1, was identified over 50 years ago, but the underlying gene has remained unknown. We employed a map-based cloning strategy to identify the Ht1 causal gene, which was found to be a coiled-coil nucleotide-binding, leucine-rich repeat (NLR) gene, which we named PH4GP-Ht1. Transgenic testing confirmed that introducing the native PH4GP-Ht1 sequence to a susceptible maize variety resulted in resistance to E.turcicum race 0. A survey of the maize nested association mapping genomes revealed that susceptible Ht1 alleles had very low to no expression of the gene. Overexpression of the susceptible B73 allele, however, did not result in resistant plants, indicating that sequence variations may underlie the difference between resistant and susceptible phenotypes. Modelling of the PH4GP-Ht1 protein indicated that it has structural homology to the Arabidopsis NLR resistance gene ZAR1, and probably forms a similar homopentamer structure following activation. RNA sequencing data from an infection time course revealed that 1 week after inoculation there was a threefold reduction in fungal biomass in the PH4GP-Ht1 transgenic plants compared to wild-type plants. Furthermore, PH4GP-Ht1 transgenics had significantly more inoculation-responsive differentially expressed genes than wild-type plants, with enrichment seen in genes associated with both defence and photosynthesis. These results demonstrate that the NLR PH4GP-Ht1 is the causal gene underlying Ht1, which represents a different mode of action compared to the previously reported wall-associated kinase northern corn leaf blight resistance gene Htn1/Ht2/Ht3.

Zea mays RNA-seq estimated transcript abundances are strongly affected by read mapping bias

BackgroundGenetic variation for gene expression is a source of phenotypic variation for natural and agricultural species. The common approach to map and to quantify gene expression from genetically distinct individuals is to assign their RNA-seq reads to a single reference genome. However, RNA-seq reads from alleles dissimilar to this reference genome may fail to map correctly, causing transcript levels to be underestimated. Presently, the extent of this mapping problem is not clear, particularly in highly diverse species. We investigated if mapping bias occurred and if chromosomal features associated with mapping bias. Zea mays presents a model species to assess these questions, given it has genotypically distinct and well-studied genetic lines.ResultsIn Zea mays, the inbred B73 genome is the standard reference genome and template for RNA-seq read assignments. In the absence of mapping bias, B73 and a second inbred line, Mo17, would each have an approximately equal number of regulatory alleles that increase gene expression. Remarkably, Mo17 had 2–4 times fewer such positively acting alleles than did B73 when RNA-seq reads were aligned to the B73 reference genome. Reciprocally, over one-half of the B73 alleles that increased gene expression were not detected when reads were aligned to the Mo17 genome template. Genes at dissimilar chromosomal ends were strongly affected by mapping bias, and genes at more similar pericentromeric regions were less affected. Biased transcript estimates were higher in untranslated regions and lower in splice junctions. Bias occurred across software and alignment parameters.ConclusionsMapping bias very strongly affects gene transcript abundance estimates in maize, and bias varies across chromosomal features. Individual genome or transcriptome templates are likely necessary for accurate transcript estimation across genetically variable individuals in maize and other species.

A maize cytochrome b-c1 complex subunit protein ZmQCR7 controls variation in the hypersensitive response.

The gene GRMZM2G318346 which encodes a cytochrome b-c1 complex subunit 7 is associated with variation in strength of the hypersensitive response in maize. We previously identified a QTL at 3,545,354bp (B73 reference genome V2) on maize chromosome 5 associated with variation in the hypersensitive response (HR) conferred by the autoactive R-gene Rp1-D21 (Olukolu et al. in PLoS Genet 10:e1004562 2014). In this study, we show that a gene at this locus, GRMZM2G318346 which encodes a cytochrome b-c1 complex subunit seven (ZmQCR7), an important part of the mitochondrial electron transport chain, can suppress HR mediated by Rp1-D21 in a transient expression assay. ZmQCR7 alleles from two maize lines, W22 and B73 differ for the encoded proteins at just two sites, amino acid 27 (threonine and alanine in B73 and W22, respectively) and amino acid 109 (asparagine and serine), however, the B73 allele is much more effective at suppressing HR. We show that variation at amino acid 27 controlled this variation in HR-suppressing effects. We furthermore demonstrate that the B73 allele of ZmQCR7 can suppress HR induced by RPM1(D505V), another autoactive R-gene, and that Arabidopsis homologs of ZmQCR7 can also suppress NLR-induced HR. The implications of these findings are discussed.

Allele specific expression analysis identifies regulatory variation associated with stress-related genes in the Mexican highland maize landrace Palomero Toluqueño.

BackgroundGene regulatory variation has been proposed to play an important role in the adaptation of plants to environmental stress. In the central highlands of Mexico, farmer selection has generated a unique group of maize landraces adapted to the challenges of the highland niche. In this study, gene expression in Mexican highland maize and a reference maize breeding line were compared to identify evidence of regulatory variation in stress-related genes. It was hypothesised that local adaptation in Mexican highland maize would be associated with a transcriptional signature observable even under benign conditions.MethodsAllele specific expression analysis was performed using the seedling-leaf transcriptome of an F1 individual generated from the cross between the highland adapted Mexican landrace Palomero Toluqueño and the reference line B73, grown under benign conditions. Results were compared with a published dataset describing the transcriptional response of B73 seedlings to cold, heat, salt and UV treatments.ResultsA total of 2,386 genes were identified to show allele specific expression. Of these, 277 showed an expression difference between Palomero Toluqueño and B73 alleles under benign conditions that anticipated the response of B73 cold, heat, salt and/or UV treatments, and, as such, were considered to display a prior stress response. Prior stress response candidates included genes associated with plant hormone signaling and a number of transcription factors. Construction of a gene co-expression network revealed further signaling and stress-related genes to be among the potential targets of the transcription factors candidates.DiscussionPrior activation of responses may represent the best strategy when stresses are severe but predictable. Expression differences observed here between Palomero Toluqueño and B73 alleles indicate the presence of cis-acting regulatory variation linked to stress-related genes in Palomero Toluqueño. Considered alongside gene annotation and population data, allele specific expression analysis of plants grown under benign conditions provides an attractive strategy to identify functional variation potentially linked to local adaptation.

Genetic Analysis of Teosinte Alleles for Kernel Composition Traits in Maize.

Teosinte (Zea mays ssp. parviglumis) is the wild ancestor of modern maize (Zea mays ssp. mays). Teosinte contains greater genetic diversity compared with maize inbreds and landraces, but its use is limited by insufficient genetic resources to evaluate its value. A population of teosinte near isogenic lines (NILs) was previously developed to broaden the resources for genetic diversity of maize, and to discover novel alleles for agronomic and domestication traits. The 961 teosinte NILs were developed by backcrossing 10 geographically diverse parviglumis accessions into the B73 (reference genome inbred) background. The NILs were grown in two replications in 2009 and 2010 in Columbia, MO and Aurora, NY, respectively, and near infrared reflectance spectroscopy and nuclear magnetic resonance calibrations were developed and used to rapidly predict total kernel starch, protein, and oil content on a dry matter basis in bulk whole grains of teosinte NILs. Our joint-linkage quantitative trait locus (QTL) mapping analysis identified two starch, three protein, and six oil QTL, which collectively explained 18, 23, and 45% of the total variation, respectively. A range of strong additive allelic effects for kernel starch, protein, and oil content were identified relative to the B73 allele. Our results support our hypothesis that teosinte harbors stronger alleles for kernel composition traits than maize, and that teosinte can be exploited for the improvement of kernel composition traits in modern maize germplasm.

Fine mapping of a quantitative resistance gene for gray leaf spot of maize (Zea mays L.) derived from teosinte (Z. mays ssp. parviglumis).

In this study we mapped the QTL Qgls8 for gray leaf spot (GLS) resistance in maize to a ~130 kb region on chromosome 8 including five predicted genes. In previous work, using near isogenic line (NIL) populations in which segments of the teosinte (Zea mays ssp. parviglumis) genome had been introgressed into the background of the maize line B73, we had identified a QTL on chromosome 8, here called Qgls8, for gray leaf spot (GLS)resistance. We identified alternate teosinte alleles at this QTL, one conferring increased GLS resistance and one increased susceptibility relative to the B73 allele. Using segregating populations derived from NIL parents carrying these contrasting alleles, we were able to delimit the QTL region to a ~130kb (based on the B73 genome) which encompassed five predicted genes.

English

For photosynthesizing plants, studies on the effect of different levels of Nitrogen on chlorophyll could let us know about the overall physiological status of the plant under different environmental conditions. Objective of our study was to understand if there was any difference among the ten recombinant inbred lines (RILs) of IBM population and their parents B73 and Mo17 for chlorophyll content and dry matter accumulation in their response to the N stress. Five IBM genotypes that carry predominantly B73 alleles and five IBM genotypes that carry Mo17 alleles at 5 quantitative trait loci (QTL) regions associated with root traits and N use efficiency from the published Maize Genome Database were evaluated in this study along with their parents. Plants were grown in the Guterman Green House (Cornell University, USA). Two nitrogen treatments (solution with high N contained 65.79 g Ca (NO3)2 4H2O in 100L of 1X solution making 5.0mM NO3 and solution with low N contained 2.63 g Ca (NO3)2 4H2O in 100L of 1X solution making 0.2 mM NO3) were given. Plant traits like leaf dry weight, stalk dry weight and root dry weight were observed. Chlorophyll content was estimated to measure the effect of different N levels on photosynthetic activity. Genotype with high in B73 composition had relative advantage over Mo17 in chlorophyll content, and dry weight of roots under low N condition. The highest root-shoot ratio under low and high N was observed in IBM056 and IBM153 respectively. Except for IBM153 and IBM337, all other genotypes showed reduced LFS at high N. The result showing lower root-shoot ratio and leaf fraction of shoot (LFS) under the high N treatment suggests that shoot growth increases more than root growth in response to increased N application and that within the shoot, stem growth increases more than leaf growth. Results support the conclusion that under low N condition, shoot growth is retarded than root growth. Keywords: Nitrogen, Maize, Chlorophyll, Dry matter accumulation, Photosynthesis.

Genetic mapping shows intraspecific variation and transgressive segregation for caterpillar-induced aphid resistance in maize.

Plants in nature have inducible defences that sometimes lead to targeted resistance against particular herbivores, but susceptibility to others. The metabolic diversity and genetic resources available for maize (Zea mays) make this a suitable system for a mechanistic study of within-species variation in such plant-mediated interactions between herbivores. Beet armyworms (Spodoptera exigua) and corn leaf aphids (Rhopalosiphum maidis) are two naturally occurring maize herbivores with different feeding habits. Whereas chewing herbivore-induced methylation of 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one glucoside (DIMBOA-Glc) to form 2-hydroxy-4,7-dimethoxy-1,4-benzoxazin-3-one glucoside (HDMBOA-Glc) promotes caterpillar resistance, lower DIMBOA-Glc levels favour aphid reproduction. Thus, caterpillar-induced DIMBOA-Glc methyltransferase activity in maize is predicted to promote aphid growth. To test this hypothesis, the impact of S. exigua feeding on R. maidis progeny production was assessed using seventeen genetically diverse maize inbred lines. Whereas aphid progeny production was increased by prior caterpillar feeding on lines B73, Ki11, Ki3 and Tx303, it decreased on lines Ky21, CML103, Mo18W and W22. Genetic mapping of this trait in a population of B73 × Ky21 recombinant inbred lines identified significant quantitative trait loci on maize chromosomes 1, 7 and 10. There is a transgressive segregation for aphid resistance, with the Ky21 alleles on chromosomes 1 and 7 and the B73 allele on chromosome 10 increasing aphid progeny production. The chromosome 1 QTL coincides with a cluster of three maize genes encoding benzoxazinoid O-methyltransferases that convert DIMBOA-Glc to HDMBOA-Glc. Gene expression studies and benzoxazinoid measurements indicate that S. exigua -induced responses in this pathway differentially affect R. maidis resistance in B73 and Ky21.

Additive effects of two quantitative trait loci that confer Rhopalosiphum maidis (corn leaf aphid) resistance in maize inbred line Mo17.

Plants show considerable within-species variation in their resistance to insect herbivores. In the case of Zea mays (cultivated maize), Rhopalosiphum maidis (corn leaf aphids) produce approximately twenty times more progeny on inbred line B73 than on inbred line Mo17. Genetic mapping of this difference in maize aphid resistance identified quantitative trait loci (QTL) on chromosomes 4 and 6, with the Mo17 allele reducing aphid reproduction in each case. The chromosome 4 QTL mapping interval includes several genes involved in the biosynthesis of DIMBOA (2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one), a maize defensive metabolite that also is required for callose accumulation in response to aphid feeding. Consistent with the known association of callose with plant defence against aphids, R. maidis reproduction on B73×Mo17 recombinant inbred lines was negatively correlated with both DIMBOA content and callose formation. Further genetic mapping, as well as experiments with near-isogenic lines, confirmed that the Mo17 allele causes increased DIMBOA accumulation relative to the B73 allele. The chromosome 6 aphid resistance QTL functions independently of DIMBOA accumulation and has an effect that is additive to that of the chromosome 4 QTL. Thus, at least two separate defence mechanisms account for the higher level of R. maidis resistance in Mo17 compared with B73.

A pair of homoeolog ClpP5 genes underlies a virescent yellow-like mutant and its modifier in maize.

Gene-background interaction is a commonly observed phenomenon in many species, but the molecular mechanisms of such an interaction is less well understood. Here we report the cloning of a maize mutant gene and its modifier. A recessive mutant with a virescent yellow-like (vyl) phenotype was identified in an ethyl methanesulfonate-mutagenized population derived from the maize inbred line B73. Homozygous mutant maize plants exhibited a yellow leaf phenotype after emergence but gradually recovered and became indistinguishable from wild-type plants after approximately 2 weeks. Taking the positional cloning approach, the Chr.9_ClpP5 gene, one of the proteolytic subunits of the chloroplast Clp protease complex, was identified and validated as the candidate gene for vyl. When introgressed by backcross into the maize inbred line PH09B, the mutant phenotype of vyl lasted much longer in the greenhouse and was lethal in the field, implying the presence of a modifier(s) for vyl. A major modifier locus was identified on chromosome 1, and a paralogous ClpP5 gene was isolated and confirmed as the candidate for the vyl-modifier. Expression of Chr.1_ClpP5 is induced significantly in B73 by the vyl mutation, while the expression of Chr.1_ClpP5 in PH09B is not responsive to the vyl mutation. Moreover, expression and sequence analysis suggests that the PH09B Chr.1_ClpP5 allele is functionally weaker than the B73 allele. We propose that functional redundancy between duplicated paralogous genes is the molecular mechanism for the interaction between vyl and its modifier.

Allelic Effect Variation at Key Photoperiod Response Quantitative Trait Loci in Maize

Tropical maize (Zea mays L.) represents a valuable genetic resource containing unique alleles not present in elite temperate maize. The strong delay in flowering in response to long daylength photoperiods exhibited by most tropical maize hinders its incorporation into temperate maize breeding programs. We tested the hypothesis that diverse tropical inbreds carry alleles with similar effects at four key photoperiod response quantitative trait loci (QTL) previously identified in maize. Four tropical maize inbreds were each crossed and backcrossed twice to the temperate recurrent parent B73 to establish four sets of introgression lines. Evaluation of these lines under long daylengths demonstrated that all four QTL have significant effects on flowering time or height in these lines, but the functional allelic effects varied substantially across the tropical donor lines. At the most important photoperiod response QTL on chromosome 10, one tropical line allele even promoted earlier flowering relative to the B73 allele. Significant allelic effect differences among tropical founders were also demonstrated directly in an F2 population derived from the cross of Ki14 and CML254. The chromosome 10 photoperiod response QTL position was validated in a set of heterogeneous inbred families evaluated in field tests and in controlled environments.

Allozyme-Specific Modification of a Maize Seed Chitinase by a Protein Secreted by the Fungal Pathogen Stenocarpella maydis

Stenocarpella maydis causes both dry-ear rot and stalk rot of maize. Maize inbred lines have varying levels of resistance to ear rot caused by S. maydis. The genetic basis of resistance appears to rely on multiple genetic factors, none of which are known. The commonly used stiff-stalk inbred line B73 has been shown to be strongly susceptible to ear rot caused by S. maydis. Here, we report that the ChitA protein alloform from B73, ChitA-F, encoded by a known allele of the chiA gene, is susceptible to modification by a protein (Stm-cmp) secreted by S. maydis. We also identify a new allele of chiA (from inbred line LH82) which encodes ChitA-S, an alloform of ChitA that is resistant to Stm-cmp modification. Chitinase zymogram analysis of seed from a commercial field showed the presence of both ChitA alloforms in healthy ears, and showed that ChitA-F but not ChitA-S was modified in ears rotted by S. maydis. The ChitA-F protein was purified from inbred line B73 and ChitA-S from LH82. ChitA-F was modified more efficiently than ChitA-S by S. maydis protein extracts in vitro. The chiA gene from LH82 was cloned and sequenced. It is a novel allele that encodes six polymorphisms relative to the known allele from B73. This is the first demonstration that the susceptibility to modification of a fungal targeted plant chitinase differs among inbred lines. These findings suggest that the LH82 chiA allele may be a specific genetic determinant that contributes to resistance to ear rot caused by S. maydis whereas the B73 allele may contribute to susceptibility.

Resistance loci affecting distinct stages of fungal pathogenesis: use of introgression lines for QTL mapping and characterization in the maize--Setosphaeria turcica pathosystem.

BackgroundStudies on host-pathogen interactions in a range of pathosystems have revealed an array of mechanisms by which plants reduce the efficiency of pathogenesis. While R-gene mediated resistance confers highly effective defense responses against pathogen invasion, quantitative resistance is associated with intermediate levels of resistance that reduces disease progress. To test the hypothesis that specific loci affect distinct stages of fungal pathogenesis, a set of maize introgression lines was used for mapping and characterization of quantitative trait loci (QTL) conditioning resistance to Setosphaeria turcica, the causal agent of northern leaf blight (NLB). To better understand the nature of quantitative resistance, the identified QTL were further tested for three secondary hypotheses: (1) that disease QTL differ by host developmental stage; (2) that their performance changes across environments; and (3) that they condition broad-spectrum resistance.ResultsAmong a set of 82 introgression lines, seven lines were confirmed as more resistant or susceptible than B73. Two NLB QTL were validated in BC4F2 segregating populations and advanced introgression lines. These loci, designated qNLB1.02 and qNLB1.06, were investigated in detail by comparing the introgression lines with B73 for a series of macroscopic and microscopic disease components targeting different stages of NLB development. Repeated greenhouse and field trials revealed that qNLB1.06Tx303 (the Tx303 allele at bin 1.06) reduces the efficiency of fungal penetration, while qNLB1.02B73 (the B73 allele at bin 1.02) enhances the accumulation of callose and phenolics surrounding infection sites, reduces hyphal growth into the vascular bundle and impairs the subsequent necrotrophic colonization in the leaves. The QTL were equally effective in both juvenile and adult plants; qNLB1.06Tx303 showed greater effectiveness in the field than in the greenhouse. In addition to NLB resistance, qNLB1.02B73 was associated with resistance to Stewart's wilt and common rust, while qNLB1.06Tx303 conferred resistance to Stewart's wilt. The non-specific resistance may be attributed to pleiotropy or linkage.ConclusionsOur research has led to successful identification of two reliably-expressed QTL that can potentially be utilized to protect maize from S. turcica in different environments. This approach to identifying and dissecting quantitative resistance in plants will facilitate the application of quantitative resistance in crop protection.

Natural variation in maize architecture is mediated by allelic differences at the PINOID co‐ortholog barren inflorescence2

We characterized allelic variation at barren inflorescence2 (bif2), a maize co-ortholog of the Arabidopsis PINOID protein kinase (PID), and tested for trait associations with bif2 in both an association mapping population of 277 diverse maize inbreds and in the inter-mated B73 x Mo17 (IBM) linkage population. Results from the quantitative analyses were compared with previous reports of bif2 phenotypes in mutagenesis studies. All three approaches (association, linkage, and mutagenesis) detect a significant effect of bif2 on tassel architecture. Association mapping implicates bif2 in an unexpectedly wide range of traits including plant height, node number, leaf length, and flowering time. Linkage mapping finds a significant interaction effect for node number between bif2 and other loci, in keeping with previous reports that bif2;spi1 and Bif2;Bif1 double mutants produce fewer phytomers. The Mo17 allele is associated with a reduced tassel branch zone and shows lower expression than the B73 allele in hybrid B73-Mo17 F(1) inflorescences, consistent with the complete absence of tassel branches in the bif2 knockout mutant. Overall, these data suggest that allelic variation at bif2 affects maize architecture by modulating auxin transport during vegetative and inflorescence development.

Quantitative Genetic Dissection of Shoot Architecture Traits in Maize: Towards a Functional Genomics Approach

Quantitative trait loci (QTL) affecting the total number of leaves made before flowering and the number of leaves below the uppermost ear (NLBE) were mapped and characterized using the intermated B73 × Mo17 recombinant inbred lines (IBMRILs) of maize (Zea mays L.). B73 and Mo17 typically make 20 and 17 leaves, 14 and 11 of which are below the ear. Total number of leaves and the number of leaves below the uppermost ear are ∼80% heritable in the IBMRILs, which show strongly transgressive phenotypic ranges of 15 to 24 and 10 to 18 leaves for these traits. B73 alleles at loci in chromosome bins 1.06, 3.06, 4.08, 8.04, 8.05, 9.07, and 10.04 increase leaf numbers, with all but the 3.06 QTL affecting both of these highly correlated traits (r = 0.86, p < 0.0001). Conservative QTL confidence intervals were computed and projected onto the draft maize genome sequence, revealing very narrow localizations (∼1 Mb) for four of the seven loci. More than 40% of the heritable variation for both traits is explained by an additive model, squarely accounting for the dramatic parental differences, but leaving the basis of the strong transgression unexplained. In addition, error rate control and confidence interval methods tailored for composite interval mapping are introduced, and their potential for improving QTL reporting is discussed.