Diverse Inbred Lines Research Articles

Phenotypic Plasticity Contributes to Maize Adaptation and Heterosis.

Plant phenotypic plasticity describes altered phenotypic performance of an individual when grown in different environments. Exploring genetic architecture underlying plant plasticity variation may help mitigate the detrimental effects of a rapidly changing climate on agriculture, but little research has been done in this area to date. In the present study, we established a population of 976 maize F1 hybrids by crossing 488 diverse inbred lines with two elite testers. Genome-wide association study identified hundreds of quantitative trait loci associated with phenotypic plasticity variation across diverse F1 hybrids, the majority of which contributed very little variance, in accordance with the polygenic nature of these traits. We identified several quantitative trait locus regions that may have been selected during the tropical-temperate adaptation process. We also observed heterosis in terms of phenotypic plasticity, in addition to the traditional genetic value differences measured between hybrid and inbred lines, and the pattern of which was affected by genetic background. Our results demonstrate a landscape of phenotypic plasticity in maize, which will aid in the understanding of its genetic architecture, its contribution to adaptation and heterosis, and how it may be exploited for future maize breeding in a rapidly changing environment.

Evaluation of Corn Germplasm Accessions for Resistance to Clavibacter nebraskensis, Causal Agent of Goss's Bacterial Wilt and Leaf Blight.

Goss's bacterial wilt and leaf blight of corn (Zea mays), caused by Clavibacter nebraskensis, is a reemerging disease in the Midwestern United States. From 2011 to 2013, field studies and a greenhouse study were conducted to assess the University of Illinois maize inbred collection for putative sources of resistance to Goss's bacterial wilt and leaf blight. This inbred collection consisted of over 2,000 diverse inbred corn lines that have been collected from all over the world. An initial field screen of over 1,000 inbred lines from the collection was conducted in Urbana, IL in 2011. These lines were inoculated with a C. nebraskensis cell suspension and rated for Goss's bacterial wilt and leaf blight severity using a 1-to-9 scale, with a score of 1 being most resistant. Means for Goss's bacterial wilt and leaf blight ratings ranged from 1 to 8.5. The initial screen identified over 150 lines that had high levels of resistance (severity score of ≤2.5). In total, 177 lines were used in the second stage of field screening. In the second stage, average Goss's bacterial wilt and leaf blight severity ranged from 1.1 to 7.4. Nine lines with high levels of resistance in 2011 and 2012 were advanced to the third stage of field screening. The mean Goss's bacterial wilt and leaf blight severity rating of the resistant lines in the last stage was 1.9, while the susceptible check had a mean score of 6.4. These nine lines were also used in the greenhouse to assess whether resistance varied based on inoculating roots, stems, or leaves. Disease severity was significantly (P ≤ 0.05) less when roots were inoculated compared with both leaf and stem inoculations, which were not significantly different from each other. Lines having high levels of field resistance were also found to be resistant in greenhouse screening regardless of inoculation method. Clustering of pedigree distance of the 34 resistant lines (severity score of ≤2.5) with known pedigree information found that 21 clustered with the Lancaster heterotic family, 4 were related to the Iowa Stiff Stalk Synthetic family, and 9 did not cluster with an identifiable heterotic family. These results show that the Lancaster family is an excellent source of Goss's wilt resistance, and that fewer sources of resistance were found in other families. The most resistant lines identified from this research are potential sources of resistance to Goss's bacterial wilt and leaf blight, and their lineage can be used in corn breeding programs to develop resistant hybrids.

Identification, Association of Natural Variation and Expression Analysis of ZmNAC9 Gene Response to Low Phosphorus in Maize Seedling Stage

Phosphorus (P) is an essential macroelement supporting maize productivity and low-P stress is a limiting factor of maize growth and yield. Improving maize plant tolerance to low P through molecular breeding is an effective alternative to increase crop productivity. In this study, a total of 111 diverse maize inbred lines were used to identify the favorable alleles and nucleotide diversity of candidate ZmNAC9, which plays an important role in response to low P and regulation in root architecture. A significant difference was found under low- and sufficient-P conditions for each of the 22 seedling traits, and a total of 41 polymorphic sites including 32 single nucleotide polymorphisms (SNPs) and 9 insertion and deletions (InDels) were detected in ZmNAC9 among 111 inbred lines. Among the 41 polymorphic studied sites, a total of 39 polymorphic sites were associated with 20 traits except for the dry weight of shoots and forks, of which six sites were highly significantly associated with a diverse number of low-P tolerant root trait index values by using a mixed linear model (MLM) at −log10 P = 3.61. In addition, 29 polymorphic sites under P-sufficient and 32 polymorphic sites under P-deficient conditions were significantly associated with a diverse number of seedling traits, of which five polymorphic sites (position S327, S513, S514, S520, and S827) were strongly significantly associated with multiple seedling traits under low-P and normal-P conditions. Among highly significant sites, most of the sites were associated with root traits under low-P, normal-P, and low-P trait index values. Linkage disequilibrium (LD) was strong at (r2 > 1.0) in 111 inbred lines. Furthermore, the effect of five significant sites was verified for haplotypes in 111 lines and the favorable allele S520 showed a positive effect on the dry weight of roots under the low-P condition. Furthermore, the expression pattern confirmed that ZmNAC9 was highly induced by low P in the roots of the P-tolerant 178 inbred line. Moreover, the subcellular localization of ZmNAC9 encoded by protein was located in the cytoplasm and nucleus. Haplotypes carrying more favorable alleles exhibited superior effects on phenotypic variation and could be helpful in developing molecular markers in maize molecular breeding programs. Taken together, the finding of this study might lead to further functions of ZmNAC9 and genes that might be involved in responses to low-P stress in maize.

Genetic Diversity and Population Structure of Maize Inbred Lines with Varying Levels of Resistance to Striga Hermonthica Using Agronomic Trait-Based and SNP Markers.

Striga hermonthica is a serious biotic stress limiting maize production in sub-Saharan Africa. The limited information on the patterns of genetic diversity among maize inbred lines derived from source germplasm with mixed genetic backgrounds limits the development of inbred lines, hybrids, and synthetics with durable resistance to S. hermonthica. This study was conducted to assess the level of genetic diversity in a panel of 150 diverse maize inbred lines using agronomic and molecular data and also to infer the population structure among the inbred lines. Ten Striga-resistance-related traits were used for the phenotypic characterization, and 16,735 high-quality single-nucleotide polymorphisms (SNPs), identified by genotyping-by-sequencing (GBS), were used for molecular diversity. The phenotypic and molecular hierarchical cluster analyses grouped the inbred lines into five clusters, respectively. However, the grouping patterns between the phenotypic and molecular hierarchical cluster analyses were inconsistent due to non-overlapping information between the phenotypic and molecular data. The correlation between the phenotypic and molecular diversity matrices was very low (0.001), which is in agreement with the inconsistencies observed between the clusters formed by the phenotypic and molecular diversity analyses. The joint phenotypic and genotypic diversity matrices grouped the inbred lines into three groups based on their reaction patterns to S. hermonthica, and this was able to exploit a broad estimate of the actual diversity among the inbred lines. The joint analysis shows an invaluable insight for measuring genetic diversity in the evaluated materials. The result indicates that wide genetic variability exists among the inbred lines and that the joint diversity analysis can be utilized to reliably assign the inbred lines into heterotic groups and also to enhance the level of resistance to Striga in new maize varieties.

QTL mapping for resistance to ear rot caused by Fusarium graminearum using an IBM Syn10 DH population in maize

Gibberella ear rot (GER) caused by Fusarium graminearum (teleomorph Gibberella zeae), is one of the most devastating diseases in maize that not only severely reduces grain yield but also contaminates the kernels with potent mycotoxins which is harmful to livestock and humans. Identification of stable quantitative trait loci (QTL) and candidate genes conferring resistance is a promising strategy in resistance breeding programs for the disease. Here, the ten times intermated B73 × Mo17-doubled haploid (IBM Syn10 DH) population comprising a total of 298 diverse maize inbred lines were subjected to phenotypic evaluation in a 2-year field test across two environments, and the IBM Syn10 DH population were employed to detect QTL across multiple environments using an ultra-high-density bin map. Ten QTL were detected on chromosomes 1, 2, 3, 6, 7, 8, and 9, respectively. Of them, a major QTL on chromosome 7, named qGRfg7, could explain 33.26% of the phenotypic variation. Based on fine-mapping strategy, we narrowed down the interested region into 91 kb, flanked by two developed Indel markers. A predicted gene in the mapped region, encoding a lipopolysaccharide-binding protein (LBP), is suggested to be a candidate gene conferring resistance to GER disease. These results will facilitate the breeding process for GER resistance by marker-assisted selection (MAS) in maize.

Heterosis of leaf and rhizosphere microbiomes in field-grown maize.

Macroorganisms' genotypes shape their phenotypes, which in turn shape the habitat available to potential microbial symbionts. This influence of host genotype on microbiome composition has been demonstrated in many systems; however, most previous studies have either compared unrelated genotypes or delved into molecular mechanisms. As a result, it is currently unclear whether the heritability of host-associated microbiomes follows similar patterns to the heritability of other complex traits. We take a new approach to this question by comparing the microbiomes of diverse maize inbred lines and their F1 hybrid offspring, which we quantified in both rhizosphere and leaves of field-grown plants using 16S-v4 and ITS1 amplicon sequencing. We show that inbred lines and hybrids differ consistently in the composition of bacterial and fungal rhizosphere communities, as well as leaf-associated fungal communities. A wide range of microbiome features display heterosis within individual crosses, consistent with patterns for nonmicrobial maize phenotypes. For leaf microbiomes, these results were supported by the observation that broad-sense heritability in hybrids was substantially higher than narrow-sense heritability. Our results support our hypothesis that at least some heterotic host traits affect microbiome composition in maize.

Assessment of genetic diversity in inbred lines of maize (Zea mays L.) and its relationship with heterosis

The genetic divergence among thirteen genotypes of maize were estimated by using Mahalanobis D2 statistic for ten characters. The genotypes were grouped into three clusters. Cluster I comprised 11 parental genotypes (CLQR, VL111, R4093, R6429, S8481, S8200, CML490, HK1, WLS, CLQRC and G18), while Cluster II (CLQ25) and III (CG18) were mono-genotypic. The highest inter cluster distance was recorded between cluster II and III (4.88) followed by cluster I and III (4.67). Maximum genetic distance was observed between the parents WLS and G18, whereas minimum distance was observed between the parents VL111 and S8481 suggesting more variability in genetic makeup of the genotypes included in these clusters. Cluster I comprising of 11 genotypes have maximum mean values for number of kernels per row and number of kernels row per cob while minimum mean values for plant height and ear height. Cluster II containing only CLQ25 have maximum mean values for ear girth and grain yield per plant. The highest contribution in the manifestation of total genetic divergence was exhibited by grain yield per plant (28.63%) followed by ear girth (28.47%), ear length (19.23%), number of kernel per row (11.50), number of kernels row per cob (10.04). Out of 30 crosses, 11 crosses showed positive significant heterobeltiosis for grain yield per plant. The crosses CLQ25 × CLQR and WLS × R4093 belongs to high divergent class. It could be concluded that genetic diversity can be utilized as a reliable parameter for predicting heterosis in hybrids.

Genotypic and phenotypic characterization of a large, diverse population of maize near-isogenic lines.

Genome-wide association (GWA) studies can identify quantitative trait loci (QTL) putatively underlying traits of interest, and nested association mapping (NAM) can further assess allelic series. Near-isogenic lines (NILs) can be used to characterize, dissect and validate QTL, but the development of NILs is costly. Previous studies have utilized limited numbers of NILs and introgression donors. We characterized a panel of 1270 maize NILs derived from crosses between 18 diverse inbred lines and the recurrent inbred parent B73, referred to as the nested NILs (nNILs). The nNILs were phenotyped for flowering time, height and resistance to three foliar diseases, and genotyped with genotyping-by-sequencing. Across traits, broad-sense heritability (0.4-0.8) was relatively high. The 896 genotyped nNILs contain 2638 introgressions, which span the entire genome with substantial overlap within and among allele donors. GWA with the whole panel identified 29 QTL for height and disease resistance with allelic variation across donors. To date, this is the largest and most diverse publicly available panel of maize NILs to be phenotypically and genotypically characterized. The nNILs are a valuable resource for the maize community, providing an extensive collection of introgressions from the founders of the maize NAM population in a B73 background combined with data on six agronomically important traits and from genotyping-by-sequencing. We demonstrate that the nNILs can be used for QTL mapping and allelic testing. The majority of nNILs had four or fewer introgressions, and could readily be used for future fine mapping studies.

Assessing the host genetic background effects on type 2 diabetes and obesity development in response to mixed-oral bacteria and high-fat diet using the collaborative cross mouse model.

BackgroundHost genetic background and sex, play central roles in defining the pathogenesis of type 2 diabetes (T2D), obesity and infectious diseases. Our previous studies demonstrated the utilization of genetically highly diverse inbred mouse lines, namely collaborative cross (CC), for dissecting host susceptibility for the development of T2D and obesity, showing significant variations following high‐fat (42% fat) diet (HFD). Here, we aimed to assessing the host genetic background and sex effects on T2D and obesity development in response to oral‐mixed bacterial infection and HFD using the CC lines.Materials and MethodsStudy cohort consists of 97 mice from 2 CC lines (both sexes), maintained on either HFD or Standard diet (CHD) for 12 weeks. At week 5 a group of mice from each diet were infected with Porphyromonas gingivalis (Pg) and Fusobacterium nucleatum (Fn) bacteria (control groups without infection). Body weight (BW) and glucose tolerance ability were assessed at the end time point of the experiment.ResultsThe CC lines varied (P < .05) at their BW gain and glucose tolerance ability (with sex effect) in response to diets and/or infection, showing opposite responses despite sharing the same environmental conditions. The combination of diet and infection enhances BW accumulation for IL1912, while restraints it for IL72. As for glucose tolerance ability, only females (both lines) were deteriorated in response to infection.ConclusionsThis study emphasizes the power of the CC mouse population for the characterization of host genetic makeup for defining the susceptibility of the individual to development of obesity and/or impaired glucose tolerance.

Genetic divergence studies in Bottle gourd [Lagenaria siceraria (Mol.) Standl.

The present investigation was carried out at Horticultural Research Station, Mondouri, Bidhan Chandra Krishi Viswavidyalaya, Mohanpur, Nadia, West Bengal, India during spring summer seasons of 2015, 2016 and 2017 in a Randomized Block Design with three replications. Genetic divergence was assessed among twenty one genotypes of bottle gourd for sixteen quantitative characters using Mahalanobis’ D2 statistics. The genotypes were grouped into six clusters. Fruit yield per plant (53.81%) contributed maximum towards divergence followed ascorbic acid content of the pulp (28.10%), seed number per fruit (4.29%) and TSS of the pulp (4.29%). Highest inter cluster distance was observed between cluster I and VI followed by cluster V and VI. Highest cluster mean values were observed for most of the traits with the genotypes present in cluster I. Based on superior mean performance for fruit yield per plant, genetic distances and clustering pattern, six promising and diverse inbred lines or varieties of bottle gourd viz., RJBGC-140, RJBGC-118, BOGVAR-2 and Pusa Samridhi (from cluster I) and Arka Bahar (from cluster VI) and Pusa Sandesh (from cluster V) selected as parents for the exploitation of hybrid vigour in bottle gourd.

Complex Genetic System Involved in Fusarium Ear Rot Resistance in Maize as Revealed by GWAS, Bulked Sample Analysis, and Genomic Prediction.

Fusarium ear rot (FER) caused by Fusarium verticillioides is one of the most prevalent maize diseases in China and worldwide. Resistance to FER is a complex trait controlled by multiple genes highly affected by environment. In this paper, genome-wide association study (GWAS), bulked sample analysis (BSA), and genomic prediction were performed for understanding FER resistance using 509 diverse inbred lines, which were genotyped by 37,801 high-quality single-nucleotide polymorphisms (SNPs). Ear rot evaluation was performed using artificial inoculation in four environments in China: Xinxiang, Henan, and Shunyi, Beijing, during 2017 and 2018. Significant phenotypic and genetic variation for FER severity was observed, and FER resistance was significantly correlated among the four environments with a generalized heritability of 0.78. GWAS identified 23 SNPs that were associated with FER resistance, 2 of which (1_226233417 on chromosome 1 and 10_14501044 on chromosome 10) were associated at threshold of 2.65 × 10-7 [-log(0.01/37,801)]. Using BSA, resistance quantitative trait loci were identified on chromosomes 3, 4, 7, 9, and 10 at the 90% confidence level and on chromosomes 3 and 10 at the 95% confidence level. A key region, bin 10.03, was detected by both GWAS and BSA. Genomic prediction for FER resistance showed that the prediction accuracy by trait-related markers was higher than that by randomly selected markers under different levels of marker density. Marker-assisted selection using genomic prediction could be an efficient strategy for genetic improvement for complex traits like FER resistance.

Genome-wide association study reveals that different pathways contribute to grain quality variation in sorghum (Sorghum bicolor)

BackgroundIn sorghum (Sorghum bicolor), one paramount breeding objective is to increase grain quality. The nutritional quality and end use value of sorghum grains are primarily influenced by the proportions of tannins, starch and proteins, but the genetic basis of these grain quality traits remains largely unknown. This study aimed to dissect the natural variation of sorghum grain quality traits and identify the underpinning genetic loci by genome-wide association study.ResultsLevels of starch, tannins and 17 amino acids were quantified in 196 diverse sorghum inbred lines, and 44 traits based on known metabolic pathways and biochemical interactions amongst the 17 amino acids calculated. A Genome-wide association study (GWAS) with 3,512,517 SNPs from re-sequencing data identified 14, 15 and 711 significant SNPs which represented 14, 14, 492 genetic loci associated with levels of tannins, starch and amino acids in sorghum grains, respectively. Amongst these significant SNPs, two SNPs were associated with tannin content on chromosome 4 and colocalized with three previously identified loci for Tannin1, and orthologs of Zm1 and TT16 genes. One SNP associated with starch content colocalized with sucrose phosphate synthase gene. Furthermore, homologues of opaque1 and opaque2 genes associated with amino acid content were identified. Using the KEGG pathway database, six and three candidate genes of tannins and starch were mapped into 12 and 3 metabolism pathways, respectively. Thirty-four candidate genes were mapped into 16 biosynthetic and catabolic pathways of amino acids. We finally reconstructed the biosynthetic pathways for aspartate and branched-chain amino acids based on 15 candidate genes identified in this study.ConclusionPromising candidate genes associated with grain quality traits have been identified in the present study. Some of them colocalized with previously identified genetic regions, but novel candidate genes involved in various metabolic pathways which influence grain quality traits have been dissected. Our study acts as an entry point for further validation studies to elucidate the complex mechanisms controlling grain quality traits such as tannins, starch and amino acids in sorghum.

Diversity analysis of provitamin A maize inbred lines using single nucleotide polymorphism markers

ABSTRACTThe success of a breeding programme can be determined by the level of genetic diversity that exists among breeding materials. This study was carried out to (i) determine the level of genetic diversity that exist among the collection of tropical provitamin A maize inbred lines and (ii) evaluate the genetic structure and divergence of provitamin A maize inbred lines. Forty-six advanced provitamin A inbred lines obtained from the International Maize and Wheat Improvement Centre (CIMMYT) and International Institute of Tropical Agriculture (IITA) were genotyped using 3047 single nucleotide polymorphism (SNP) markers. Genotyping was done following an Illumina Infinium HD Assay Ultra protocol. Eight six percent of the SNPs were polymorphic with the mean polymorphic information content of 0.36. Cluster analysis displayed two distinct clusters. The average pairwise genetic distance among the inbred lines was 0.60. The average gene diversity was 0.359. Variation was partitioned into among individuals (78%), among populations (12%) and within individuals (10%). Overall results suggest the inbred lines are genetically diverse. The key implication of this study is that selection should be done from genetically diverse inbred lines in order to exploit heterosis when developing hybrids.

Genome-Wide Analyses and Prediction of Resistance to MLN in Large Tropical Maize Germplasm.

Maize lethal necrosis (MLN), caused by co-infection of maize chlorotic mottle virus and sugarcane mosaic virus, can lead up to 100% yield loss. Identification and validation of genomic regions can facilitate marker assisted breeding for resistance to MLN. Our objectives were to identify marker-trait associations using genome wide association study and assess the potential of genomic prediction for MLN resistance in a large panel of diverse maize lines. A set of 1400 diverse maize tropical inbred lines were evaluated for their response to MLN under artificial inoculation by measuring disease severity or incidence and area under disease progress curve (AUDPC). All lines were genotyped with genotyping by sequencing (GBS) SNPs. The phenotypic variation was significant for all traits and the heritability estimates were moderate to high. GWAS revealed 32 significantly associated SNPs for MLN resistance (at p < 1.0 × 10−6). For disease severity, these significantly associated SNPs individually explained 3–5% of the total phenotypic variance, whereas for AUDPC they explained 3–12% of the total proportion of phenotypic variance. Most of significant SNPs were consistent with the previous studies and assists to validate and fine map the big quantitative trait locus (QTL) regions into few markers’ specific regions. A set of putative candidate genes associated with the significant markers were identified and their functions revealed to be directly or indirectly involved in plant defense responses. Genomic prediction revealed reasonable prediction accuracies. The prediction accuracies significantly increased with increasing marker densities and training population size. These results support that MLN is a complex trait controlled by few major and many minor effect genes.

The comparison of several methods for calculating the degree of heritability and calculating the number of genes in maize (Zea mays L.). I. Agronomic traits

The objective of present study was to compare of several methods for estimating the degree of heritability and calculating the number of genes using generation mean analysis of maize (Zea mays L.). The experiment was conducted at the field of Field Crop Dept. College of Agric / Univ. of Baghdad, for many seasons, spring and fall seasons 2009, 2010, spring 2011 and fall 2013.Six diverse inbred lines were crossed to produce F1,F2,BC1 and BC2 for four superior crosses.Broad-sense and narrow sense heritability estimates based on variance of different generations. The results showed that the four formulas used to estimate the heritability were different in estimating the values of heritability of different traits, as well as the different crosses used in the study. The broad sense heritability was differed according to the components of the formulas used to estimate. The number of genes that controlled the trait differed according to the different method of estimation and according to the studied trait, and to the all crosses. The highest number of genes (for all formulas) for plant yield, ranging from 15.85 (cross 4) to 155.7 (cross 2), because the yield is complex quantitative trait of several components, so the number of genes they control are the sum of genes that control the components. The inbreeding depression differed according to the parents involved in the crosses, and according to the studied trait, the inbreeding depression ranged from -7 for number of branches/tassel of cross 1 and leaf area of cross 3 to 68 for grain yield of cross 2. We can conclude that the best formula to estimate heritability depends on experimental design and methods of breeding.

Vacuolar invertase genes SbVIN1 and SbVIN2 are differently associated with stem and grain traits in sorghum (Sorghum bicolor)

In higher plants, vacuolar invertases play essential roles in sugar metabolism, organ development, and sink strength. In sorghum (Sorghum bicolor), two vacuolar invertase genes, SbVIN1 (Sobic. 004G004800) and SbVIN2 (Sobic. 006G160700) have been reported, but their enzymatic properties and functional differences are largely unknown. We combined molecular, biochemical and genomic approaches to investigate their roles in sorghum stem and grain traits. SbVIN1 and SbVIN2 showed different expression levels in internodes, leaves, and panicles, indicating that their importance in each organ was different. In an in vitro sucrose hydrolysis assay, proteins of both genes heterologously expressed in Pichia pastoris displayed similar enzyme properties including the same optimum reaction pH (5) and similar Km for sucroe (49 mmol L−1 and 45 mmol L−1 for SbVIN1 and SbVIN2, respectively). The optimum reaction temperatures of SbVIN1 and SbVIN2 were 45 °C and 65 °C, respectively. SbVIN2 showed higher tolerance to high temperature than SbVIN1. We characterized the sequence variation of these two vacuolar invertase genes in a panel of 216 diverse inbred lines of sweet and grain sorghum and performed gene-based association analysis. SbVIN1 showed significant associations with stem traits including stem length, stem diameter, internode number, stem fresh weight, and Brix, as well as grain traits including hundred-grain weight and grain width. Significantly associated variation sites were mainly in 5′ upstream and intron regions. SbVIN2 only associated with grain width and stem water-soluble carbohydrates (WSCs) content. We conclude that the vacuolar invertase genes SbVIN1 and SbVIN2 are differently associated with stem and grain traits in sorghum.

Genetic and environmental variation impact the cuticular hydrocarbon metabolome on the stigmatic surfaces of maize

BackgroundSimple non-isoprenoid hydrocarbons accumulate in discrete regions of the biosphere, including within bacteria and algae as a carbon and/or energy store, and the cuticles of plants and insects, where they may protect against environmental stresses. The extracellular cuticular surfaces of the stigmatic silks of maize are rich in linear hydrocarbons and therefore provide a convenient system to study the biological origins and functions of these unique metabolites.ResultsTo test the hypotheses that genetics and environment influence the accumulation of surface hydrocarbons on silks and to examine the breadth of metabolome compositions across diverse germplasm, cuticular hydrocarbons were analyzed on husk-encased silks and silks that emerged from the husk leaves from 32 genetically diverse maize inbred lines, most of which are commonly utilized in genetics experiments. Total hydrocarbon accumulation varied ~ 10-fold among inbred lines, and up to 5-fold between emerged and husk-encased silks. Alkenes accounted for 5-60% of the total hydrocarbon metabolome, and the majority of alkenes were monoenes with a double bond at either the 7th or 9th carbon atom of the alkyl chain. Total hydrocarbon accumulation was impacted to similar degrees by genotype and husk encasement status, whereas genotype predominantly impacted alkene composition. Only minor differences in the metabolome were observed on silks that were emerged into the external environment for 3- versus 6-days. The environmental influence on the metabolome was further investigated by growing inbred lines in 2 years, one of which was warmer and wetter. Inbred lines grown in the drier year accumulated up to 2-fold more hydrocarbons and up to a 22% higher relative abundance of alkenes. In summary, the surface hydrocarbon metabolome of silks is primarily governed by genotype and husk encasement status, with smaller impacts of environment and genotype-by-environment interactions.ConclusionsThis study reveals that the composition of the cuticular hydrocarbon metabolome on silks is affected significantly by genetic factors, and is therefore amenable to dissection using quantitative genetic approaches. Such studies will clarify the genetic mechanisms responsible for the accumulation of these metabolites, enabling detailed functional investigations of the diverse and complex protective roles of silk surface lipids against environmental stresses.

Natural variation in a calreticulin gene causes reduced resistance to Ca2+ deficiency-induced tipburn in Chinese cabbage (Brassica rapa ssp. pekinensis).

Tipburn is an irreversible physiological disorder of Chinese cabbage that decreases crop value. Because of a strong environmental component, tipburn-resistant cultivars are the only solution, although tipburn resistance genes are unknown in Chinese cabbage. We studied three populations of Chinese cabbage over four growing seasons under field conditions: (a) 194 diverse inbred lines, (b) a doubled haploid (DH100) population, and (c) an F2 population. The 194 lines were genotyped using single nucleotide polymorphism markers, and genome-wide-association mapping showed that 24 gQTLs were significantly associated with tipburn disease index. Analysis of the DH100 and F2 populations identified a shared tipburn-associated locus, gqbTRA06, that was found to cover the region defined by one of the 24 gQTLs. Of 35 genes predicted in the 0.14-Mb quantitative trait locus region, Bra018575 (calreticulin family protein, BrCRT2) showed higher expression levels during disease development. We cloned the two BrCRT2 alleles from tipburn-resistant (BrCRT2R ) and tipburn-susceptible (BrCRT2S ) lines and identified a 51-bp deletion in BrCRT2S . Overexpression of BrCRT2R increased Ca2+ storage in the Arabidopsis crt2 mutant and also reduced cell death in leaf tips and margins under Ca2+ -depleted conditions. Our results suggest that BrCRT2 is a possible candidate gene for controlling tipburn in Chinese cabbage.

Genotyping-by-sequencing and SNP-arrays are complementary for detecting quantitative trait loci by tagging different haplotypes in association studies

BackgroundSingle Nucleotide Polymorphism (SNP) array and re-sequencing technologies have different properties (e.g. calling rate, minor allele frequency profile) and drawbacks (e.g. ascertainment bias). This lead us to study their complementarity and the consequences of using them separately or combined in diversity analyses and Genome-Wide Association Studies (GWAS). We performed GWAS on three traits (grain yield, plant height and male flowering time) measured in 22 environments on a panel of 247 F1 hybrids obtained by crossing 247 diverse dent maize inbred lines with a same flint line. The 247 lines were genotyped using three genotyping technologies (Genotyping-By-Sequencing, Illumina Infinium 50 K and Affymetrix Axiom 600 K arrays).ResultsThe effects of ascertainment bias of the 50 K and 600 K arrays were negligible for deciphering global genetic trends of diversity and for estimating relatedness in this panel. We developed an original approach based on linkage disequilibrium (LD) extent in order to determine whether SNPs significantly associated with a trait and that are physically linked should be considered as a single Quantitative Trait Locus (QTL) or several independent QTLs. Using this approach, we showed that the combination of the three technologies, which have different SNP distributions and densities, allowed us to detect more QTLs (gain in power) and potentially refine the localization of the causal polymorphisms (gain in resolution).ConclusionsConceptually different technologies are complementary for detecting QTLs by tagging different haplotypes in association studies. Considering LD, marker density and the combination of different technologies (SNP-arrays and re-sequencing), the genotypic data available were most likely enough to well represent polymorphisms in the centromeric regions, whereas using more markers would be beneficial for telomeric regions.

Multiple Maize Reference Genomes Impact the Identification of Variants by Genome-Wide Association Study in a Diverse Inbred Panel.

Use of a single reference genome for genome-wide association studies (GWAS) limits the gene space represented to that of a single accession. This limitation can complicate identification and characterization of genes located within presence-absence variations (PAVs). In this study, we present the draft de novo genome assembly of 'PHJ89', an 'Oh43'-type inbred line of maize ( L.). From three separate reference genome assemblies ('B73', 'PH207', and PHJ89) that represent the predominant germplasm groups of maize, we generated three separate whole-seedling gene expression profiles and single nucleotide polymorphism (SNP) matrices from a panel of 942 diverse inbred lines. We identified 34,447 (B73), 39,672 (PH207), and 37,436 (PHJ89) transcripts that are not present in the respective reference genome assemblies. Genome-wide association studies were conducted in the 942 inbred panel with both the SNP and expression data values to map (SCMV) resistance. Highlighting the impact of alternative reference genomes in gene discovery, the GWAS results for SCMV resistance with expression values as a surrogate measure of PAV resulted in robust detection of the physical location of a known resistance gene when the B73 reference that contains the gene was used, but not the PH207 reference. This study provides the valuable resource of the Oh43-type PHJ89 genome assembly as well as SNP and expression data for 942 individuals generated from three different reference genomes.