The Metabolic Profile of Young, Watered Chickpea Plants Can Be Used as a Biomarker to Predict Seed Number under Terminal Drought.

Chickpea is the second-most-cultivated legume globally, with India and Australia being the two largest producers. In both of these locations, the crop is sown on residual summer soil moisture and left to grow on progressively depleting water content, finally maturing under terminal drought conditions. The metabolic profile of plants is commonly, correlatively associated with performance or stress responses, e.g., the accumulation of osmoprotective metabolites during cold stress. In animals and humans, metabolites are also prognostically used to predict the likelihood of an event (usually a disease) before it occurs, e.g., blood cholesterol and heart disease. We sought to discover metabolic biomarkers in chickpea that could be used to predict grain yield traits under terminal drought, from the leaf tissue of young, watered, healthy plants. The metabolic profile (GC-MS and enzyme assays) of field-grown chickpea leaves was analysed over two growing seasons, and then predictive modelling was applied to associate the most strongly correlated metabolites with the final seed number plant-1. Pinitol (negatively), sucrose (negatively) and GABA (positively) were significantly correlated with seed number in both years of study. The feature selection algorithm of the model selected a larger range of metabolites including carbohydrates, sugar alcohols and GABA. The correlation between the predicted seed number and actual seed number was R2 adj = 0.62, demonstrating that the metabolic profile could be used to predict a complex trait with a high degree of accuracy. A previously unknown association between D-pinitol and hundred-kernel weight was also discovered and may provide a single metabolic marker with which to predict large seeded chickpea varieties from new crosses. The use of metabolic biomarkers could be used by breeders to identify superior-performing genotypes before maturity is reached.

Differential expression of starch and sucrose metabolic genes linked to varying biomass yield in Miscanthus hybrids.

BACKGROUND: Miscanthus is a commercial lignocellulosic biomass crop owing to its high biomass productivity and low chemical input requirements. Within an interspecific Miscanthus cross, progeny with high biomass yield were shown to have low concentrations of starch and sucrose but high concentrations of fructose. We performed a transcriptional RNA-seq analysis between selected Miscanthus hybrids with contrasting values for these phenotypes to clarify how these phenotypes are genetically controlled. RESULTS: We observed that genes directly involved in the synthesis and degradation of starch and sucrose were down-regulated in high-yielding Miscanthus hybrids. At the same time, glycolysis and export of triose phosphates were up-regulated in high-yielding Miscanthus hybrids. These differentially expressed genes and biological functions were regulated by a well-connected network of less than 25 co-regulated transcription factors. CONCLUSIONS: Our results evidence a direct relationship between high expression of essential enzymatic genes in the starch and sucrose pathways and co-expression with their transcriptional regulators, with high starch concentrations and lower biomass production. The strong interconnectivity between gene expression and regulators, chemotype and agronomic traits opens the door to use the expression of well-characterised genes associated with carbohydrate metabolism, particularly in the starch and sucrose pathway, for the early selection of high biomass-yielding genotypes from large Miscanthus populations.

Investigating the potential of novel non-woven fabrics for efficient pollination control in plant breeding.

Plant breeding is achieved through the controlled self- or cross-pollination of individuals and typically involves isolation of floral parts from selected parental plants. Paper, cellulose or synthetic materials are used to avoid self pollination or cross contamination. Low seed set limits the rate of breeding progress and increases costs. We hypothesized that a novel 'non-woven' fabric optimal for both pollination and seed set in multiple plant species could be developed. After determining the baseline pollen characteristics and usage requirements we established iterative three phase development and biological testing. This determined (1) that white fabric gave superior seed return and informed the (2) development of three non-woven materials using different fibre and layering techniques. We tested their performance in selfing and hybridisation experiments recording differences in performance by material type within species. Finally we (3) developed further advanced fabrics with increased air permeability and tested biological performance. An interaction between material type and species was observed and environmental decoupling investigated, showing that the non-woven fabrics had superior water vapour transmission and temperature regulation compared to controls. Overall, non-woven fabrics outperformed existing materials for both pollination and seed set and we found that different materials can optimize species-specific, rather than species-generic performance.

Predicting future biomass yield in Miscanthus using the carbohydrate metabolic profile as a biomarker.

In perennial energy crop breeding programmes, it can take several years before a mature yield is reached when potential new varieties can be scored. Modern plant breeding technologies have focussed on molecular markers, but for many crop species, this technology is unavailable. Therefore, prematurity predictors of harvestable yield would accelerate the release of new varieties. Metabolic biomarkers are routinely used in medicine, but they have been largely overlooked as predictive tools in plant science. We aimed to identify biomarkers of productivity in the bioenergy crop, Miscanthus, that could be used prognostically to predict future yields. This study identified a metabolic profile reflecting productivity in Miscanthus by correlating the summer carbohydrate composition of multiple genotypes with final yield 6 months later. Consistent and strong, significant correlations were observed between carbohydrate metrics and biomass traits at two separate field sites over 2 years. Machine-learning feature selection was used to optimize carbohydrate metrics for support vector regression models, which were able to predict interyear biomass traits with a correlation (R) of >0.67 between predicted and actual values. To identify a causal basis for the relationships between the glycome profile and biomass, a 13C-labelling experiment compared carbohydrate partitioning between high- and low-yielding genotypes. A lower yielding and slower growing genotype partitioned a greater percentage of the 13C pulse into starch compared to a faster growing genotype where a greater percentage was located in the structural biomass. These results supported a link between plant performance and carbon flow through two rival pathways (starch vs. sucrose), with higher yielding plants exhibiting greater partitioning into structural biomass, via sucrose metabolism, rather than starch. Our results demonstrate that the plant metabolome can be used prognostically to anticipate future yields and this is a method that could be used to accelerate selection in perennial energy crop breeding programmes.

Could Miscanthus replace maize as the preferred substrate for anaerobic digestion in the United Kingdom? Future breeding strategies.

Fodder maize is the most commonly used crop for biogas production owing to its high yields, high concentrations of starch and good digestibility. However, environmental concerns and possible future conflict with land for food production may limit its long-term use. The bioenergy grass, Miscanthus, is a high-yielding perennial that can grow on marginal land and, with 'greener' environmental credentials, may offer an alternative. To compete with maize, the concentration of non-structural carbohydrates (NSC) and digestibility may need to be improved. Non-structural carbohydrates were quantified in 38 diverse genotypes of Miscanthus in green-cut biomass in July and October. The aim was to determine whether NSC abundance could be a target for breeding programmes or whether genotypes already exist that could rival maize for use in anaerobic digestion systems. The saccharification potential and measures of N P and K were also studied. The highest concentrations of NSC were in July, reaching a maximum of 20% DW. However, the maximum yield was in October with 300-400 g NSC plant-1 owing to higher biomass. The digestibility of the cell wall was higher in July than in October, but the increase in biomass meant yields of digestible sugars were still higher in October. Nutrient concentrations were at least twofold higher in July compared to November, and the abundance of potassium showed the greatest degree of variation between genotypes. The projected maximum yield of NSC was 1.3 t ha-1 with significant variation to target for breeding. Starch accumulated in the highest concentrations and continued to increase into autumn in some genotypes. Therefore, starch, rather than sugars, would be a better target for breeding improvement. If harvest date was brought forward to autumn, nutrient losses in non-flowering genotypes would be comparable to an early spring harvest.

High yielding biomass genotypes of willow (Salix spp.) show differences in below ground biomass allocation.

Willows (Salix spp.) grown as short rotation coppice (SRC) are viewed as a sustainable source of biomass with a positive greenhouse gas (GHG) balance due to their potential to fix and accumulate carbon (C) below ground. However, exploiting this potential has been limited by the paucity of data available on below ground biomass allocation and the extent to which it varies between genotypes. Furthermore, it is likely that allocation can be altered considerably by environment. To investigate the role of genotype and environment on allocation, four willow genotypes were grown at two replicated field sites in southeast England and west Wales, UK. Above and below ground biomass was intensively measured over two two-year rotations. Significant genotypic differences in biomass allocation were identified, with below ground allocation differing by up to 10% between genotypes. Importantly, the genotype with the highest below ground biomass also had the highest above ground yield. Furthermore, leaf area was found to be a good predictor of below ground biomass. Growth environment significantly impacted allocation; the willow genotypes grown in west Wales had up to 94% more biomass below ground by the end of the second rotation. A single investigation into fine roots showed the same pattern with double the volume of fine roots present. This greater below ground allocation may be attributed primarily to higher wind speeds, plus differences in humidity and soil characteristics. These results demonstrate that the capacity exists to breed plants with both high yields and high potential for C accumulation.

Leaves of the Arabidopsis maltose exporter1 mutant exhibit a metabolic profile with features of cold acclimation in the warm.

BACKGROUND: Arabidopsis plants accumulate maltose from starch breakdown during cold acclimation. The Arabidopsis mutant, maltose excess1-1, accumulates large amounts of maltose in the plastid even in the warm, due to a deficient plastid envelope maltose transporter. We therefore investigated whether the elevated maltose level in mex1-1 in the warm could result in changes in metabolism and physiology typical of WT plants grown in the cold. PRINCIPAL FINDINGS: Grown at 21 °C, mex1-1 plants were much smaller, with fewer leaves, and elevated carbohydrates and amino acids compared to WT. However, after transfer to 4 °C the total soluble sugar pool and amino acid concentration was in equal abundance in both genotypes, although the most abundant sugar in mex1-1 was still maltose whereas sucrose was in greatest abundance in WT. The chlorophyll a/b ratio in WT was much lower in the cold than in the warm, but in mex1-1 it was low in both warm and cold. After prolonged growth at 4 °C, the shoot biomass, rosette diameter and number of leaves at bolting were similar in mex1-1 and WT. CONCLUSIONS: The mex1-1 mutation in warm-grown plants confers aspects of cold acclimation, including elevated levels of sugars and amino acids and low chlorophyll a/b ratio. This may in turn compromise growth of mex1-1 in the warm relative to WT. We suggest that elevated maltose in the plastid could be responsible for key aspects of cold acclimation.

A nuclear-localized protein, KOLD SENSITIV-1, affects the expression of cold-responsive genes during prolonged chilling in Arabidopsis.

Plants respond to cold by transcriptional and metabolic responses which underlie tolerance and acclimation mechanisms, but details at the molecular level are incomplete. Here we describe KOLD SENSITIV-1 (KOS1), a new gene required for responses to cold. KOS1 protein is predicted to have coiled-coil, Structural Maintenance of Chromosomes and nuclear-targeting domains. GFP-labeled KOS1 localizes to the nucleus. Null mutants could not be isolated but two independent knockdown T-DNA mutants were obtained. Growth and development of kos1 knockdown mutant plants was comparable to wild type when grown at 21°C. However, when grown at 4°C these mutants exhibited accelerated leaf yellowing and smaller rosette size than wild type. Quantitative RT-PCR revealed that in the cold kos1 mutants had reduced expression of cold-responsive transcripts COR15A, COR15B, BAM3 and AMY3. Metabolite profiling revealed that ascorbate levels were lower in the mutants in the cold relative to wild type. KOS1 therefore represents a new gene that influences the regulation of transcript and metabolite levels in response to prolonged chilling temperatures.