Gareth J. Norton

Genome-wide association mapping reveals a rich genetic architecture of complex traits in Oryza sativa

Asian rice, Oryza sativa is a cultivated, inbreeding species that feeds over half of the worlds population. Understanding the genetic basis of diverse physiological, developmental, and morphological traits provides the basis for improving yield, quality and sustainability of rice. Here we show the results of a genome-wide association study based on genotyping 44,100 SNP variants across 413 diverse accessions of O. sativa collected from 82 countries that were systematically phenotyped for 34 traits. Using cross-population-based mapping strategies, we identified dozens of common variants influencing numerous complex traits. Significant heterogeneity was observed in the genetic architecture associated with subpopulation structure and response to environment. This work establishes an open-source translational research platform for genome-wide association studies in rice that directly links molecular variation in genes and metabolic pathways with the germplasm resources needed to accelerate varietal development and crop improvement.

Grain Unloading of Arsenic Species in Rice

Rice (Oryza sativa) is the staple food for over half the worlds population yet may represent a significant dietary source of inorganic arsenic (As), a nonthreshold, class 1 human carcinogen. Rice grain As is dominated by the inorganic species, and the organic species dimethylarsinic acid (DMA). To investigate how As species are unloaded into grain rice, panicles were excised during grain filling and hydroponically pulsed with arsenite, arsenate, glutathione-complexed As, or DMA. Total As concentrations in flag leaf, grain, and husk, were quantified by inductively coupled plasma mass spectroscopy and As speciation in the fresh grain was determined by x-ray absorption near-edge spectroscopy. The roles of phloem and xylem transport were investigated by applying a ± stem-girdling treatment to a second set of panicles, limiting phloem transport to the grain in panicles pulsed with arsenite or DMA. The results demonstrate that DMA is translocated to the rice grain with over an order magnitude greater efficiency than inorganic species and is more mobile than arsenite in both the phloem and the xylem. Phloem transport accounted for 90% of arsenite, and 55% of DMA, transport to the grain. Synchrotron x-ray fluorescence mapping and fluorescence microtomography revealed marked differences in the pattern of As unloading into the grain between DMA and arsenite-challenged grain. Arsenite was retained in the ovular vascular trace and DMA dispersed throughout the external grain parts and into the endosperm. This study also demonstrates that DMA speciation is altered in planta, potentially through complexation with thiols.

Assessing the influence of compost and biochar amendments on the mobility and toxicity of metals and arsenic in a naturally contaminated mine soil.

Amending contaminated soils with organic wastes can influence trace element mobility and toxicity. Soluble concentrations of metals and arsenic were measured in pore water and aqueous soil extracts following the amendment of a heavily contaminated mine soil with compost and biochar (10% v:v) in a pot experiment. Speciation modelling and toxicity assays (Vibrio fischeri luminescence inhibition and Lolium perenne germination) were performed to discriminate mechanisms controlling metal mobility and assess toxicity risk thereafter. Biochar reduced free metal concentrations furthest but dissolved organic carbon primarily controlled metal mobility after compost amendment. Individually, both amendments induced considerable solubilisation of arsenic to pore water (>2500 μg l(-1)) related to pH and soluble phosphate but combining amendments most effectively reduced toxicity due to simultaneous reductions in extractable metals and increases in soluble nutrients (P). Thus the measure-monitor-model approach taken determined that combining the amendments was most effective at mitigating attendant toxicity risk.

Rice-arsenate interactions in hydroponics : whole genome transcriptional analysis

Rice (Oryza sativa) varieties that are arsenate-tolerant (Bala) and -sensitive (Azucena) were used to conduct a transcriptome analysis of the response of rice seedlings to sodium arsenate (AsV) in hydroponic solution. RNA extracted from the roots of three replicate experiments of plants grown for 1 week in phosphate-free nutrient with or without 13.3 μM AsV was used to challenge the Affymetrix (52K) GeneChip Rice Genome array. A total of 576 probe sets were significantly up-regulated at least 2-fold in both varieties, whereas 622 were down-regulated. Ontological classification is presented. As expected, a large number of transcription factors, stress proteins, and transporters demonstrated differential expression. Striking is the lack of response of classic oxidative stress-responsive genes or phytochelatin synthases/synthatases. However, the large number of responses from genes involved in glutathione synthesis, metabolism, and transport suggests that glutathione conjugation and arsenate methylation may be important biochemical responses to arsenate challenge. In this report, no attempt is made to dissect differences in the response of the tolerant and sensitive variety, but analysis in a companion article will link gene expression to the known tolerance loci available in the Bala×Azucena mapping population.

Variation in Rice Cadmium Related to Human Exposure

Cereal grains are the dominant source of cadmium in the human diet, with rice being to the fore. Here we explore the effect of geographic, genetic, and processing (milling) factors on rice grain cadmium and rice consumption rates that lead to dietary variance in cadmium intake. From a survey of 12 countries on four continents, cadmium levels in rice grain were the highest in Bangladesh and Sri Lanka, with both these countries also having high per capita rice intakes. For Bangladesh and Sri Lanka, there was high weekly intake of cadmium from rice, leading to intakes deemed unsafe by international and national regulators. While genetic variance, and to a lesser extent milling, provide strategies for reducing cadmium in rice, caution has to be used, as there is environmental regulation as well as genetic regulation of cadmium accumulation within rice grains. For countries that import rice, grain cadmium can be controlled by where that rice is sourced, but for countries with subsistence rice economies that have high levels of cadmium in rice grain, agronomic and breeding strategies are required to lower grain cadmium.

Environmental and genetic control of arsenic accumulation and speciation in rice grain: comparing a range of common cultivars grown in contaminated sites across Bangladesh, China, and India

The concentration of arsenic (As) in rice grains has been identified as a risk to human health. The high proportion of inorganic species of As (As(i)) is of particular concern as it is a nonthreshold, class 1 human carcinogen. To be able to breed rice with low grain As, an understanding of genetic variation and the effect of different environments on genetic variation is needed. In this study, 13 cultivars grown at two field sites each in Bangladesh, India, and China are evaluated for grain As. There was a significant site, genotype, and site by genotype interaction for total grain As. Correlations were observed only between sites in Bangladesh and India, not between countries or within the Chinese sites. For seven cultivars the As was speciated which revealed significant effects of site, genotype, and site by genotype interaction for percentage As(i). Breeding low grain As cultivars that will have consistently low grain As and low As(i), over multiple environments using traditional breeding approaches may be difficult, although CT9993-5-10-1-M, Lemont, Azucena, and Te-qing in general had low grain As across the field sites.

Genetic mapping of the rice ionome in leaves and grain: identification of QTLs for 17 elements including arsenic, cadmium, iron and selenium.

Research into the composition of cereal grains is motivated by increased interest in food quality. Here multi-element analysis is conducted on leaves and grain of the Bala x Azucena rice mapping population grown in the field. Quantitative trait loci (QTLs) for the concentration of 17 elements were detected, revealing 36 QTLs for leaves and 41 for grains. Epistasis was detected for most elements. There was very little correlation between leaf and grain element concentrations. For selenium, lead, phosphorus and magnesium QTLs were detected in the same location for both tissues. In general, there were no major QTL clusters, suggesting separate regulation of each element. QTLs for grain iron, zinc, molybdenum and selenium are potential targets for marker assisted selection to improve seed nutritional quality. An epistatic interaction for grain arsenic also looks promising to decrease the concentration of this carcinogenic element.

Improved resolution in the position of drought-related QTLs in a single mapping population of rice by meta-analysis

BackgroundMeta-analysis of QTLs combines the results of several QTL detection studies and provides narrow confidence intervals for meta-QTLs, permitting easier positional candidate gene identification. It is usually applied to multiple mapping populations, but can be applied to one. Here, a meta-analysis of drought related QTLs in the Bala × Azucena mapping population compiles data from 13 experiments and 25 independent screens providing 1,650 individual QTLs separated into 5 trait categories; drought avoidance, plant height, plant biomass, leaf morphology and root traits. A heat map of the overlapping 1 LOD confidence intervals provides an overview of the distribution of QTLs. The programme BioMercator is then used to conduct a formal meta-analysis at example QTL clusters to illustrate the value of meta-analysis of QTLs in this population.ResultsThe heat map graphically illustrates the genetic complexity of drought related traits in rice. QTLs can be linked to their physical position on the rice genome using Additional file 1 provided. Formal meta-analysis on chromosome 1, where clusters of QTLs for all trait categories appear close, established that the sd1 semi-dwarfing gene coincided with a plant height meta-QTL, that the drought avoidance meta-QTL was not likely to be associated with this gene, and that this meta-QTL was not pleiotropic with close meta-QTLs for leaf morphology and root traits. On chromosome 5, evidence suggests that a drought avoidance meta-QTL was pleiotropic with leaf morphology and plant biomass meta-QTLs, but not with meta-QTLs for root traits and plant height 10 cM lower down. A region of dense root QTL activity graphically visible on chromosome 9 was dissected into three meta-QTLs within a space of 35 cM. The confidence intervals for meta-QTLs obtained ranged from 5.1 to 14.5 cM with an average of 9.4 cM, which is approximately 180 genes in rice.ConclusionThe meta-analysis is valuable in providing improved ability to dissect the complex genetic structure of traits, and distinguish between pleiotropy and close linkage. It also provides relatively small target regions for the identification of positional candidate genes.

Phloem transport of arsenic species from flag leaf to grain during grain filling

• Strategies to reduce arsenic (As) in rice grain, below concentrations that represent a serious human health concern, require that the mechanisms of As accumulation within grain be established. Therefore, retranslocation of As species from flag leaves into filling rice grain was investigated. • Arsenic species were delivered through cut flag leaves during grain fill. Spatial unloading within grains was investigated using synchrotron X-ray fluorescence (SXRF) microtomography. Additionally, the effect of germanic acid (a silicic acid analog) on grain As accumulation in arsenite-treated panicles was examined. • Dimethylarsinic acid (DMA) and monomethylarsonic acid (MMA) were extremely efficiently retranslocated from flag leaves to rice grain; arsenate was poorly retranslocated, and was rapidly reduced to arsenite within flag leaves; arsenite displayed no retranslocation. Within grains, DMA rapidly dispersed while MMA and inorganic As remained close to the entry point. Germanic acid addition did not affect grain As in arsenite-treated panicles. Three-dimensional SXRF microtomography gave further information on arsenite localization in the ovular vascular trace (OVT) of rice grains. • These results demonstrate that inorganic As is poorly remobilized, while organic species are readily remobilized, from leaves to grain. Stem translocation of inorganic As may not rely solely on silicic acid transporters.

Organic Matter-Solid Phase Interactions Are Critical for Predicting Arsenic Release and Plant Uptake in Bangladesh Paddy Soils

Agroecological zones within Bangladesh with low levels of arsenic in groundwater and soils produce rice that is high in arsenic with respect to other producing regions of the globe. Little is known about arsenic cycling in these soils and the labile fractions relevant for plant uptake when flooded. Soil porewater dynamics of field soils (n = 39) were recreated under standardized laboratory conditions to investigate the mobility and interplay of arsenic, Fe, Si, C, and other elements, in relation to rice grain element composition, using the dynamic sampling technique diffusive gradients in thin films (DGT). Based on a simple model using only labile DGT measured arsenic and dissolved organic carbon (DOC), concentrations of arsenic in Aman (Monsoon season) rice grain were predicted reliably. DOC was the strongest determinant of arsenic solid-solution phase partitioning, while arsenic release to the soil porewater was shown to be decoupled from that of Fe. This study demonstrates the dual importance of organic matter (OM), in terms of enhancing arsenic release from soils, while reducing bioavailability by sequestering arsenic in solution.