Stanisław Jeżowski

A framework genetic map for Miscanthus sinensis from RNAseq-based markers shows recent tetraploidy

BackgroundMiscanthus (subtribe Saccharinae, tribe Andropogoneae, family Poaceae) is a genus of temperate perennial C4 grasses whose high biomass production makes it, along with its close relatives sugarcane and sorghum, attractive as a biofuel feedstock. The base chromosome number of Miscanthus (x = 19) is different from that of other Saccharinae and approximately twice that of the related Sorghum bicolor (x = 10), suggesting large-scale duplications may have occurred in recent ancestors of Miscanthus. Owing to the complexity of the Miscanthus genome and the complications of self-incompatibility, a complete genetic map with a high density of markers has not yet been developed.ResultsWe used deep transcriptome sequencing (RNAseq) from two M. sinensis accessions to define 1536 single nucleotide variants (SNVs) for a GoldenGate™ genotyping array, and found that simple sequence repeat (SSR) markers defined in sugarcane are often informative in M. sinensis. A total of 658 SNP and 210 SSR markers were validated via segregation in a full sibling F1 mapping population. Using 221 progeny from this mapping population, we constructed a genetic map for M. sinensis that resolves into 19 linkage groups, the haploid chromosome number expected from cytological evidence. Comparative genomic analysis documents a genome-wide duplication in Miscanthus relative to Sorghum bicolor, with subsequent insertional fusion of a pair of chromosomes. The utility of the map is confirmed by the identification of two paralogous C4-pyruvate, phosphate dikinase (C4-PPDK) loci in Miscanthus, at positions syntenic to the single orthologous gene in Sorghum.ConclusionsThe genus Miscanthus experienced an ancestral tetraploidy and chromosome fusion prior to its diversification, but after its divergence from the closely related sugarcane clade. The recent timing of this tetraploidy complicates discovery and mapping of genetic markers for Miscanthus species, since alleles and fixed differences between paralogs are comparable. These difficulties can be overcome by careful analysis of segregation patterns in a mapping population and genotyping of doubled haploids. The genetic map for Miscanthus will be useful in biological discovery and breeding efforts to improve this emerging biofuel crop, and also provide a valuable resource for understanding genomic responses to tetraploidy and chromosome fusion.

Progress in upscaling Miscanthus biomass production for the European bio-economy with seed-based hybrids

Field trials in Europe with Miscanthus over the past 25 years have demonstrated that interspecies hybrids such as M. × giganteus (M × g) combine both high yield potentials and low inputs in a wide range of soils and climates. Miscanthus hybrids are expected to play a major role in the provision of perennial lignocellulosic biomass across much of Europe as part of a lower carbon economy. However, even with favourable policies in some European countries, uptake has been slow. M × g, as a sterile clone, can only be propagated vegetatively, which leads to high establishment costs and low multiplication rates. Consequently, a decade ago, a strategic decision to develop rapidly multiplied seeded hybrids was taken. To make progress on this goal, we have (1) harnessed the genetic diversity in Miscanthus by crossing and progeny testing thousands of parental combinations to select several candidate seed‐based hybrids adapted to European environments, (2) established field scale seed production methods with annual multiplication factors >1500×, (3) developed the agronomy for establishing large stands from seed sown plug plants to reduce establishment times by a year compared to M × g, (4) trialled a range of harvest techniques to improve compositional quality and logistics on a large scale, (5) performed spatial analyses of yield potential and land availability to identify regional opportunities across Europe and doubled the area within the bio‐climatic envelope, (6) considered on‐farm economic, practical and environmental benefits that can be attractive to growers. The technical barriers to adoption have now been overcome sufficiently such that Miscanthus is ready to use as a low‐carbon feedstock in the European bio‐economy.

Cytological analysis of hybrid embryos of intergeneric crosses between Salix viminalis and Populus species.

Intergeneric hybridisation between Salix viminalis L. as the female and four Populus species (Populus trichocarpa, P. tremula, P. × canadensis and P. simonii) as male pollen donors was performed by in vitro stigma pollination. To overcome postzygotic barriers, transfer of hybrid embryos to new medium is necessary. We carried out detailed ultrastructural analyses to establish: (i) at which stage of embryo development the first signs of programmed cell death (PCD) could be detected; and (ii) at which stage the lack of serious or irreversible changes guaranteed that advanced development of hybrid plants could occur after embryo rescue. Transmission electron microscopy and confocal laser scanning microscopy revealed the presence of both developing and degenerating embryos. Developing globular, heart-shaped, and early cotyledonary embryos contained cells of correct ultrastructure. The only sign of intergeneric hybridisation was a delay in development for a few days, in comparison with control embryos. The earliest indicators of embryo degeneration were noted at 9 days after pollination (DAP). The most common indicators were excessive embryo vacuolisation, which was characterised by a large number of vesicles and formation of small vacuoles, as well as enlarged central vacuoles. Extended plastid thylakoids, folding of the cell wall, and autophagosomes were observed. Our detailed investigation of PCD in hybrid embryos enabled us to conclude that the embryo rescue technique was most effective in intergeneric willow × poplar crosses if applied between 9 and 16 DAP.

Low-temperature leaf photosynthesis of a Miscanthus germplasm collection correlates positively to shoot growth rate and specific leaf area.

Background and Aims The C4 perennial grass miscanthus has been found to be less sensitive to cold than most other C4 species, but still emerges later in spring than C3 species. Genotypic differences in miscanthus were investigated to identify genotypes with a high cold tolerance at low temperatures and quick recovery upon rising temperatures to enable them to exploit the early growing season in maritime cold climates. Suitable methods for field screening of cold tolerance in miscanthus were also identified. Methods Fourteen genotypes of M. sacchariflorus, M. sinensis, M. tinctorius and M. × giganteus were selected and grown under warm (24 °C) and cold (14 °C) conditions in a controlled environment. Dark-adapted chlorophyll fluorescence, specific leaf area (SLA) and net photosynthetic rate at a photosynthetically active radiation (PAR) of 1000 μmol m–2 s–1 (A1000) were measured. Photosynthetic light and CO2 response curves were obtained from 11 of the genotypes, and shoot growth rate was measured under field conditions. Key Results A positive linear relationship was found between SLA and light-saturated photosynthesis (Asat) across genotypes, and also between shoot growth rate under cool field conditions and A1000 at 14 °C in a climate chamber. When lowering the temperature from 24 to 14 °C, one M. sacchariflorus exhibited significantly higher Asat and maximum photosynthetic rate in the CO2 response curve (Vmax) than other genotypes at 14 °C, except M. × giganteus ‘Hornum’. Several genotypes returned to their pre-chilling A1000 values when the temperature was increased to 24 °C after 24 d growth at 14 °C. Conclusions One M. sacchariflorus genotype had similar or higher photosynthetic capacity than M. × giganteus, and may be used for cultivation together with M. × giganteus or for breeding new interspecies hybrids with improved traits for temperate climates. Two easily measured variables, SLA and shoot growth rate, may be useful for genotype screening of productivity and cold tolerance.

Establishment, Growth, and Yield Potential of the Perennial Grass Miscanthus × Giganteus on Degraded Coal Mine Soils

Miscanthus × giganteus is a giant C4 grass native to Asia. Unlike most C4 species, it is relatively cold tolerant due to adaptations across a wide range of altitudes. These grasses are characterized by high productivity and low input requirements, making them excellent candidates for bioenergy feedstock production. The aim of this study was to investigate the potential for growing Miscanthus on extremely marginal soils, degraded by open lignite (brown coal) mining. Field experiments were established within three blocks situated on waste heaps originating from the lignite mine. Analyses were conducted over the first 3 years following Miscanthus cultivation, focusing on the effect of organic and mineral fertilization on crop growth, development and yield in this extreme environment. The following levels of fertilization were implemented between the blocks: the control plot with no fertilization (D0), a plot with sewage sludge (D1), a plot with an identical amount of sewage sludge plus one dose of mineral fertilizer (D2) and a plot with an identical amount of sewage sludge plus a double dose of mineral fertilizer (D3). Crop development and characteristics (plant height, tillering, and biomass yield [dry matter]) were measured throughout the study period and analyzed using Analysis of Variance (ANOVA). Significant differences were apparent between plant development and 3rd year biomass production over the course of the study (0.964 kg plant-1 for DO compared to 1.503 kg plant-1 for D1). Soil analyses conducted over the course of the experiment showed that organic carbon levels within the soil increased significantly following the cultivation of Miscanthus, and overall, pH decreased. With the exception of iron, macronutrient concentrations remained stable throughout. The promising yields and positive effects of Miscanthus on the degraded soil suggests that long term plantations on land otherwise unsuitable for agriculture may prove to be of great environmental and economic significance.

Gas exchange and yield in Miscanthus species for three years at two locations in Poland

Głowacka, K., Jeżowski, S. and Kaczmarek, Z. 2013. Gas exchange and yield in Miscanthus species for three years at two locations in Poland. Can. J. Plant Sci. 93: 627-637. The influence of gas exchange parameters on yields of six Miscanthus clones were evaluated in two locations over three years. Five out of six genotypes had significant increases in the photosynthetic rate and intrinsic water use efficiency between the third and first or third and second years, while intercellular CO2 concentration and stomatal conductance decreased. In addition, the multiple regression analyses of yields on parameters showed that the type of the correlation coefficients for transpiration, stomatal conductance and intercellular CO2 concentration varied throughout the years. The strongest correlation (0.86) between yield and photosynthetic rate was found for 3-year-old plants.

Moisture Loss Rate in Grass Cut at Anthesis: Variation Among Selected Traditional Species

The objective of this study was to quantify the variation in moisture loss of different grass species after cutting at anthesis. The study was conducted over 3 years at Poznan, Poland. Grasses included in the experiments consisted of traditional grasses belonging to the species: Festuca—cv. Emeraude and Jordane, Dactylis—cv. Ambassador and Donata, Festulolium—cv. Felina and Hykor, as well as Phalaris—cv. Chiftain and Bamse. After cutting at anthesis, the drying rate of biomass was measured over time both in the natural environment (a field) and in a more stable environment (a barn). On this basis, significant differences were found between the tested grass cultivars in terms of the moisture loss rate of biomass. The study also showed that the year of the experiment and the environment in which drying took place had a significant effect on drying rate.