Population structure of Miscanthus sacchariflorus reveals two major polyploidization events, tetraploid-mediated unidirectional introgression from diploid M. sinensis, and diversity centred around the Yellow Sea.

BACKGROUND AND AIMS: Miscanthus, a C4 perennial grass native to East Asia, is a promising biomass crop. Miscanthus sacchariflorus has a broad geographic range, is used to produce paper in China and is one of the parents (along with Miscanthus sinensis) of the important biomass species Miscanthus × giganteus. The largest study of M. sacchariflorus population genetics to date is reported here. METHODS: Collections included 764 individuals across East Asia. Samples were genotyped with 34 605 single nucleotide polymorphisms (SNPs) derived from restriction site-associated DNA sequencing (RAD-seq) and ten plastid microsatellites, and were subjected to ploidy analysis by flow cytometry. KEY RESULTS: Six major genetic groups within M. sacchariflorus were identified using SNP data: three diploid groups, comprising Yangtze (M. sacchariflorus ssp. lutarioriparius), N China and Korea/NE China/Russia; and three tetraploid groups, comprising N China/Korea/Russia, S Japan and N Japan. Miscanthus sacchariflorus ssp. lutarioriparius was derived from the N China group, with a substantial bottleneck. Japanese and mainland tetraploids originated from independent polyploidization events. Hybrids between diploid M. sacchariflorus and M. sinensis were identified in Korea, but without introgression into either parent species. In contrast, tetraploid M. sacchariflorus in southern Japan and Korea exhibited substantial hybridization and introgression with local diploid M. sinensis. CONCLUSIONS: Genetic data indicated that the land now under the Yellow Sea was a centre of diversity for M. sacchariflorus during the last glacial maximum, followed by a series of migrations as the climate became warmer and wetter. Overall, M. sacchariflorus has greater genetic diversity than M. sinensis, suggesting that breeding and selection within M. sacchariflorus will be important for the development of improved M. × giganteus. Ornamental M. sacchariflorus genotypes in Europe and North America represent a very narrow portion of the species' genetic diversity, and thus do not well represent the species as a whole.

Ecological characteristics and in situ genetic associations for yield-component traits of wild Miscanthus from eastern Russia.

Background and aims Miscanthus is a genus of perennial C4 grasses native to East Asia. It includes the emerging ligno-cellulosic biomass crop M. ×giganteus, a hybrid between M. sinensis and M. sacchariflorus. Biomass yield and cold tolerance are of particular interest in Miscanthus, given that this crop is more temperate adapted than its C4 relatives maize, sorghum and sugarcane. Methods A plant exploration was conducted in eastern Russia, at the northern extreme of the native range for Miscanthus, with collections including 174 clonal germplasm accessions (160 M. sacchariflorus and 14 M. sinensis) from 47 sites. Accessions were genotyped by restriction site-associated DNA sequencing (RAD-seq) and plastid microsatellites. Key Results Miscanthus sinensis was found in maritime climates near Vladivostok (43·6°N) and on southern Sakhalin Island (46·6°N). Miscanthus sacchariflorus was found inland at latitudes as high as 49·3°N, where M. sinensis was absent. Most M. sacchariflorus accessions were diploid, but approx. 2 % were tetraploids. Molecular markers revealed little population structure (Jost's D < 0·007 among diploid groups) but high genetic diversity (expected heterozygosity = 0·14) within the collection of Russian M. sacchariflorus. Genome-wide association (GWA) analysis for traits measured at the collection sites revealed three M. sacchariflorus single nucleotide polymorphisms (SNPs) significantly associated with the number of stems per unit area, one with height and one with basal stem diameter; three were near or within previously described sorghum quantitative trait loci for related traits. Conclusions This new Miscanthus germplasm collection from eastern Russia will be useful for breeding Miscanthus and sugarcane cultivars with improved adaptation to cold. Moreover, a strategy is proposed to facilitate the rapid utilization of new germplasm collections: by implementing low-cost SNP genotyping to conduct GWA studies of phenotypic data obtained at collection sites, plant breeders can be provided with actionable information on which accessions have desirable traits and alleles.

Contrasting allelic distribution of CO/Hd1 homologues in Miscanthus sinensis from the East Asian mainland and the Japanese archipelago.

The genus Miscanthus is a perennial C4 grass native to eastern Asia and is a promising candidate bioenergy crop for cool temperate areas. Flowering time is a crucial factor governing regional and seasonal adaptation; in addition, it is also a key target trait for extending the vegetative phase to improve biomass potential. Homologues of CONSTANS (CO)/Heading date 1(Hd1) were cloned from Miscanthus sinensis and named MsiHd1. Sequences of MsiHd1 homologues were compared among 24 wild M. sinensis accessions from Japan, 14 from China, and three from South Korea. Two to five MsiHd1 alleles in each accession were identified, suggesting that MsiHd1 consists of at least three loci in the Miscanthus genome. Verifying the open reading frame in MsiHd1, they were classified as putative functional alleles without mutations or non-functional alleles caused by indels. The Neighbor-Joining tree indicated that one of the multiple MsiHd1 loci is a pseudogene locus without any functional alleles. The pseudogene locus was named MsiHd1b, and the other loci were considered to be part of the MsiHd1a multi-locus family. Interestingly, in most Japanese accessions 50% or more of the MsiHd1a alleles were non-functional, whereas accessions from the East Asian mainland harboured only functional alleles. Five novel miniature inverted transposable elements (MITEs) (MsiMITE1-MsiMITE5) were observed in MsiHd1a/b. MsiMITE1, detected in exon 1 of MsiHd1a, was only observed in Japanese accessions and its revertant alleles derived from retransposition were predominantly in Chinese accessions. These differences in MsiHd1a show that the dependency on functional MsiHd1a alleles is different between accessions from the East Asian mainland and Japan.

Genetic structure of Miscanthus sinensis and Miscanthus sacchariflorus in Japan indicates a gradient of bidirectional but asymmetric introgression.

Unilateral introgression from diploids to tetraploids has been hypothesized to be an important evolutionary mechanism in plants. However, few examples have been definitively identified, perhaps because data of sufficient depth and breadth were difficult to obtain before the advent of affordable high-density genotyping. Throughout Japan, tetraploid Miscanthus sacchariflorus and diploid Miscanthus sinensis are common, and occasionally hybridize. In this study, 667 M. sinensis and 78 M. sacchariflorus genotypes from Japan were characterized using 20 704 SNPs and ten plastid microsatellites. Similarity of SNP genotypes between diploid and tetraploid M. sacchariflorus indicated that the tetraploids originated through autopolyploidy. Structure analysis indicated a gradient of introgression from diploid M. sinensis into tetraploid M. sacchariflorus throughout Japan; most tetraploids had some M. sinensis DNA. Among phenotypically M. sacchariflorus tetraploids, M. sinensis ancestry averaged 7% and ranged from 1-39%, with introgression greatest in southern Japan. Unexpectedly, rare (~1%) diploid M. sinensis individuals from northern Japan were found with 6-27% M. sacchariflorus ancestry. Population structure of M. sinensis in Japan included three groups, and was driven primarily by distance, and secondarily by geographic barriers such as mountains and straits. Miscanthus speciation is a complex and dynamic process. In contrast to limited introgression between diploid M. sacchariflorus and M. sinensis in northern China, selection for adaptation to a moderate maritime climate probably favoured cross-ploidy introgressants in southern Japan. These results will help guide the selection of Miscanthus accessions for the breeding of biomass cultivars.

A footprint of past climate change on the diversity and population structure of Miscanthus sinensis.

BACKGROUND AND AIMS: Miscanthus is a perennial C4 grass that is a leading potential feedstock crop for the emerging bioenergy industry in North America, Europe and China. However, only a single, sterile genotype of M. × giganteus (M×g), a nothospecies derived from diploid M. sinensis (Msi) and tetraploid M. sacchariflorus (Msa), is currently available to farmers for biomass production. To facilitate breeding of Miscanthus, this study characterized genetic diversity and population structure of Msi in its native range of East Asia. METHODS: A total of 767 accessions were studied, including 617 Msi from most of its native range in China, Japan and South Korea, and 77 ornamental cultivars and 43 naturalized individuals from the USA. Accessions were evaluated with 21 207 restriction site-associated DNA sequencing single nucleotide polymorphism (SNP) markers, 424 GoldenGate SNPs and ten plastid microsatellite markers. KEY RESULTS: Six genetic clusters of Msi from geographically distinct regions in Asia were identified. Genetic data indicated that (1) south-eastern China was the origin of Msi populations found in temperate eastern Asia, which is consistent with this area probably having been a refugium during the last glacial maximum (LGM); (2) Msi migrated directly from south-eastern China to Japan before migrating to the same latitudes in China and Korea, which is consistent with the known sequence of warming post-LGM; (3) ornamental Msi cultivars were derived from the southern Japan population, and US naturalized populations were derived from a sub-set of the ornamental cultivars; and (4) many ornamental cultivars previously described as Msi have hybrid ancestry from Msa and Msi, whereas US naturalized populations of Msi do not. CONCLUSIONS: Population structure of Msi was driven by patterns of warming since the LGM, and secondarily by geographical barriers. This study will facilitate germplasm conservation, association analyses and identification of potential heterotic groups for the improvement of Miscanthus as a bioenergy crop.

Glionitrin A, a new diketopiperazine disulfide, activates ATM-ATR-Chk1/2 via 53BP1 phosphorylation in DU145 cells and shows antitumor effect in xenograft model.

In a recent study, we isolated the diketopiperazine disulfide glionitrin A from the co-culture broth of a mine drainage-derived fungus (Aspergillus fumigatus KMC901) and bacterium (Sphingomonas KMK001). Here, we investigated the antitumor activity of glionitrin A and its underlying molecular mechanisms in human prostate cancer DU145 cells. Glionitrin A showed significant cytotoxicity, promoting cell cycle arrest and apoptosis. Glionitrin A-treated cells exhibited elevated levels of phospho-histone 2AX (Ser139), a marker of DNA damage, and accumulated in both S phase and G2/M phase due to the activation of checkpoints associated with the ataxia-telangiectasia-mutated and ataxia-telangiectasia-mutated-Rad3-related Chk1/2 pathway downstream of p53-binding protein 1 phosphorylation at Ser1778. In addition, glionitrin A induced apoptosis through both caspase-dependent and -independent pathways. Glionitrin A activated caspase-8, -9 and -3 and also released endonuclease G from the mitochondria to the nucleus in a dose-dependent manner. Our in vivo study performed in nude mice bearing xenografts of DU145 cells showed that glionitrin A dramatically reduced the tumor volume by an average of 38.2% (5 mg/kg, per os (p.o.)) and 71.3% (10 mg/kg, p.o.) at 27 d after the beginning of treatment. Taken together, these findings provide a detailed description of the mechanism underlying the biological activity of the new natural product glionitrin A, which has the potential to be developed as an anti-prostate cancer agent.

Fabrication and characterization of quantum dots-bound hydrogels with fluorescent and temperature-sensitive functionalities.

In this work, dual-functional composite particles possessing fluorescence and temperature-sensitive functionalities were developed in the form of QD-bound hydrogels for biomedical applications. First, the surface of silica nanoparticles (SNPs) was functionalized with olefin silanes, followed by hydrogel encapsulation through a radical polymerization. The encapsulated hydrogels were poly(N- isopropylacrylamide-co-acrylic acid) P(NIPAM-co-AAc) copolymer, showing the sensitive volume changes corresponding to the alternating temperature changes between 25 degrees C and 45 degrees C. At an optimal pH5, the hydrogel-encapsulated SNPs (SiO2@hydrogel) were effectively anchored by amino quantum dots (amino-QDs) through electrostatic (attractive) interactions between carboxylate groups of hydrogels and amine groups of QDs. QD-bound hydrogels with co-monomer ratio of [NIPAM:AAc = 83:17 wt%] exhibited the higher PL intensity than other samples with [NIPAM:AAc = 96:4 wt% or 91:9 wt%], indicating that higher fraction of carboxylate groups by AAc induced the effective bounding with QDs possessing positively charged amine groups.

AraDQ: an automated digital phenotyping software for quantifying disease symptoms of flood-inoculated Arabidopsis seedlings.

BACKGROUND: Plant scientists have largely relied on pathogen growth assays and/or transcript analysis of stress-responsive genes for quantification of disease severity and susceptibility. These methods are destructive to plants, labor-intensive, and time-consuming, thereby limiting their application in real-time, large-scale studies. Image-based plant phenotyping is an alternative approach that enables automated measurement of various symptoms. However, most of the currently available plant image analysis tools require specific hardware platform and vendor specific software packages, and thus, are not suited for researchers who are not primarily focused on plant phenotyping. In this study, we aimed to develop a digital phenotyping tool to enhance the speed, accuracy, and reliability of disease quantification in Arabidopsis. RESULTS: Here, we present the Arabidopsis Disease Quantification (AraDQ) image analysis tool for examination of flood-inoculated Arabidopsis seedlings grown on plates containing plant growth media. It is a cross-platform application program with a user-friendly graphical interface that contains highly accurate deep neural networks for object detection and segmentation. The only prerequisite is that the input image should contain a fixed-sized 24-color balance card placed next to the objects of interest on a white background to ensure reliable and reproducible results, regardless of the image acquisition method. The image processing pipeline automatically calculates 10 different colors and morphological parameters for individual seedlings in the given image, and disease-associated phenotypic changes can be easily assessed by comparing plant images captured before and after infection. We conducted two case studies involving bacterial and plant mutants with reduced virulence and disease resistance capabilities, respectively, and thereby demonstrated that AraDQ can capture subtle changes in plant color and morphology with a high level of sensitivity. CONCLUSIONS: AraDQ offers a simple, fast, and accurate approach for image-based quantification of plant disease symptoms using various parameters. Its fully automated pipeline neither requires prior image processing nor costly hardware setups, allowing easy implementation of the software by researchers interested in digital phenotyping of diseased plants.

Antisense-overexpression of the MsCOMT gene induces changes in lignin and total phenol contents in transgenic tobacco plants.

Initially, we isolated the caffeic acid O-methyltransferase (COMT) gene from Miscanthus sinensis (accession number HM062766.1). Next, we produced transgenic tobacco plants with down-regulated COMT gene expression to study its control of total phenol and lignin content and to perform morphological analysis. These transgenic plants were found to have reduced PAL and ascorbate peroxidases expression, which are related to the phenylpropanoid pathway and antioxidant activity. The MsCOMT-down-regulated plants had decreased total lignin in the leaves and stem compared with control plants. Reduced flavonol concentrations were confirmed in MsCOMT-down-regulated transgenic plants. We also observed a morphological difference, with reduced plant cell number in transgenic plants harboring antisense MsCOMT. The transgenic tobacco plants with down-regulated COMT gene expression demonstrate that COMT plays a crucial role related to controlling lignin and phenol content in plants. Also, COMT activity may be related to flavonoid production in the plant lignin pathway.

Cellular Morphology and Transcriptome Comparative Analysis of Astragalus membranaceus Bunge Sprouts Cultured In Vitro under Different LED Light.

Astragalus membranaceus, the major components of which are saponins, flavonoids, and polysaccharides, has been established to have excellent pharmacological activity. After ginseng, it is the second most used medicinal plant. To examine the utility of A. membranaceus as a sprout crop for plant factory cultivation, we sought to establish a functional substance control model by comparing the transcriptomes of sprouts grown in sterile, in vitro culture using LED light sources. Having sown the seeds of A. membranaceus, these were exposed to white LED light (continuous spectrum), red LED light (632 nm, 1.58 µmol/m2/s), or blue LED light (465 nm, 1.44 µmol/m2/s) and grown for 6 weeks; after which, the samples were collected for transcriptome analysis. Scanning electron microscopy analysis of cell morphology in plants exposed to the three light sources revealed that leaf cell size was largest in those plants exposed to red light, where the thickest stem was observed in plants exposed to white light. The total number of genes in A. membranaceus spouts determined via de novo assembly was 45,667. Analysis of differentially expressed genes revealed that for the comparisons of blue LED vs. red LED, blue LED vs. white LED, and red LED vs. white LED, the numbers of upregulated genes were 132, 148, and 144, respectively. Binding, DNA integration, transport, phosphorylation, DNA biosynthetic process, membrane, and plant-type secondary cell wall biogenesis were the most enriched in the comparative analysis of blue LED vs. red LED, whereas Binding, RNA-templated DNA biosynthetic process, DNA metabolic process, and DNA integration were the most enriched in the comparative analysis of blue vs. white LED, and DNA integration and resolution of meiotic recombination intermediates were the most enrichment in the comparison between red LED vs. white LED. The GO term associated with flavonoid biosynthesis, implying the functionality of A. membranaceus, was the flavonoid biosynthetic process, which was enriched in the white LED vs. red LED comparison. The findings of this study thus indicate that different LED light sources can differentially influence the transcriptome expression pattern of A. membranaceus sprouts, which can provide a basis for establishing a flavonoid biosynthesis regulation model and thus, the cultivation of high-functional Astragalus sprouts.

Ginseng-derived exosome-like nanovesicles extracted by sucrose gradient ultracentrifugation to inhibit osteoclast differentiation.

Plant-derived extracellular nanovesicles contain RNA and proteins with unique and diverse pharmacological mechanisms. The extracellular nanovesicles encapsulating plant extracts resemble exosomes as they have a round, lipid bilayer morphology. Ginseng is anti-inflammatory, anti-cancer, immunostimulant, and osteogenic/anti-osteoporotic. Here, we confirmed that ginseng-derived extracellular nanovesicles (GDNs) inhibit osteoclast differentiation and elucidated the associated molecular mechanisms. We isolated GDNs by centrifugation with a sucrose gradient. We measured their dynamic light scattering and zeta potentials and examined their morphology by transmission electron microscopy. We used bone marrow-derived macrophages (BMMs) to determine the potential cytotoxicity of GDNs and establish their ability to inhibit osteoclast differentiation. The GDNs treatment maintained high BMM viability and proliferation whilst impeding osteoclastogenesis. Tartrate-resistant acid phosphatase and F-actin staining revealed that GDNs at concentrations >1 µg mL-1 strongly hindered osteoclast differentiation. Moreover, they substantially suppressed the RANKL-induced IκBα, c-JUN n-terminal kinase, and extracellular signal-regulated kinase signaling pathways and the genes regulating osteoclast maturation. The GDNs contained elevated proportions of Rb1 and Rg1 ginsenosides and were more effective than either of them alone or in combination at inhibiting osteoclast differentiation. In vivo bone analysis via microcomputerized tomography, bone volume/total volume ratios, and bone mineral density and bone cavity measurements demonstrated the inhibitory effect of GDNs against osteoclast differentiation in lipopolysaccharide-induced bone resorption mouse models. The results of this work suggest that GDNs are anti-osteoporotic by inhibiting osteoclast differentiation and are, therefore, promising for use in the clinical prevention and treatment of bone loss diseases.

Development of a Temperature-Responsive Hydrogel Incorporating PVA into NIPAAm for Controllable Drug Release in Skin Regeneration.

Melanoma, a highly malignant and aggressive form of skin cancer, poses a significant global health threat, with limited treatment options and potential side effects. In this study, we developed a temperature-responsive hydrogel for skin regeneration with a controllable drug release. The hydrogel was fabricated using an interpenetrating polymer network (IPN) of N-isopropylacrylamide (NIPAAm) and poly(vinyl alcohol) (PVA). PVA was chosen for its adhesive properties, biocompatibility, and ability to address hydrophobicity issues associated with NIPAAm. The hydrogel was loaded with doxorubicin (DOX), an anticancer drug, for the treatment of melanoma. The NIPAAm-PVA (N-P) hydrogel demonstrated temperature-responsive behavior with a lower critical solution temperature (LCST) around 34 °C. The addition of PVA led to increased porosity and faster drug release. In vitro biocompatibility tests showed nontoxicity and supported cell proliferation. The N-P hydrogel exhibited effective anticancer effects on melanoma cells due to its rapid drug release behavior. This N-P hydrogel system shows great promise for controlled drug delivery and potential applications in skin regeneration and cancer treatment. Further research, including in vivo studies, will be essential to advance this hydrogel system toward clinical translation and impactful advancements in regenerative medicine and cancer therapeutics.

Impact of genotype-calling methodologies on genome-wide association and genomic prediction in polyploids.

Discovery and analysis of genetic variants underlying agriculturally important traits are key to molecular breeding of crops. Reduced representation approaches have provided cost-efficient genotyping using next-generation sequencing. However, accurate genotype calling from next-generation sequencing data is challenging, particularly in polyploid species due to their genome complexity. Recently developed Bayesian statistical methods implemented in available software packages, polyRAD, EBG, and updog, incorporate error rates and population parameters to accurately estimate allelic dosage across any ploidy. We used empirical and simulated data to evaluate the three Bayesian algorithms and demonstrated their impact on the power of genome-wide association study (GWAS) analysis and the accuracy of genomic prediction. We further incorporated uncertainty in allelic dosage estimation by testing continuous genotype calls and comparing their performance to discrete genotypes in GWAS and genomic prediction. We tested the genotype-calling methods using data from two autotetraploid species, Miscanthus sacchariflorus and Vaccinium corymbosum, and performed GWAS and genomic prediction. In the empirical study, the tested Bayesian genotype-calling algorithms differed in their downstream effects on GWAS and genomic prediction, with some showing advantages over others. Through subsequent simulation studies, we observed that at low read depth, polyRAD was advantageous in its effect on GWAS power and limit of false positives. Additionally, we found that continuous genotypes increased the accuracy of genomic prediction, by reducing genotyping error, particularly at low sequencing depth. Our results indicate that by using the Bayesian algorithm implemented in polyRAD and continuous genotypes, we can accurately and cost-efficiently implement GWAS and genomic prediction in polyploid crops.

Dibenzocyclooctadiene lignans, gomisins J and N inhibit the Wnt/ß-catenin signaling pathway in HCT116 cells.

Here, we report that gomisin J and gomisin N, dibenzocyclooctadiene type lignans isolated from Schisandra chinensis, inhibit Wnt/ß-catenin signaling in HCT116 cells. Gomisins J and N appear to inhibit Wnt/ß-catenin signaling by disrupting the interaction between ß-catenin and its specific target DNA sequences (TCF binding elements, TBE) rather than by altering the expression of the ß-catenin protein. Gomisins J and N inhibit HCT116 cell proliferation by arresting the cell cycle at the G0/G1 phase. The G0/G1 phase arrest induced by gomisins J and N appears to be caused by a decrease in the expression of Cyclin D1, a representative target gene of the Wnt/ß-catenin signaling pathway, as well as Cdk2, Cdk4, and E2F-1. Therefore, gomisins J and N, the novel Wnt/ß-catenin inhibitors discovered in this study, may serve as potential agents for the prevention and treatment of human colorectal cancers.

Role of social capital in adolescents' online gaming: A longitudinal study focused on the moderating effect of social capital between gaming time and psychosocial factors.

Adolescents often create social relationships with their gaming peers who take on the role of offline friends and peer groups. Through collaboration and competition in the games, the social relationships of adolescents are becoming broader and thicker. Although this is a common phenomenon in online games, few studies have focused on the formation and roles of social capital among adolescent gamers. In particular, longitudinal research that examines the role of social capital in terms of influencing gaming time on adolescent gamers' psychosocial factors has been minimal. This study was designed to fill this gap to see the long-term effect of social capital among adolescent gamers. Specifically, by using the three-year longitudinal data involving 403 adolescents, we analyzed the effect of gaming time on psychological factors (i.e., loneliness, depression, self-esteem, and life satisfaction) with the moderating role of social capital. Results showed that social capital played a crucial moderating role. In the higher social capital group, gaming time enhanced the degree of self-esteem and life satisfaction. However, a vicious circle was found in the lower social capital group: Gaming time increased the degree of depression but decreased self-esteem, which in turn led to increase in gaming time. These results indicate that games work as an important tool for social capital cultivation among adolescent gamers, which imply successful cultivation of social capital is a key to positive gaming effects. Theoretical and practical implications are discussed.

Measurement of Environmentally Influenced Variations in Anthocyanin Accumulations in Brassica rapa subsp. Chinensis (Bok Choy) Using Hyperspectral Imaging.

Dietary supplements of anthocyanin-rich vegetables have been known to increase potential health benefits for humans. The optimization of environmental conditions to increase the level of anthocyanin accumulations in vegetables during the cultivation periods is particularly important in terms of the improvement of agricultural values in the indoor farm using artificial light and climate controlling systems. This study reports on the measurement of variations in anthocyanin accumulations in leaf tissues of four different cultivars in Brassica rapa var. chinensis (bok choy) grown under the different environmental conditions of the indoor farm using hyperspectral imaging. Anthocyanin accumulations estimated by hyperspectral imaging were compared with the measured anthocyanin accumulation obtained by destructive analysis. Between hyperspectral imaging and destructive analysis values, no significant differences in anthocyanin accumulation were observed across four bok choy cultivars grown under the anthocyanin stimulation environmental condition, whereas the estimated anthocyanin accumulations displayed cultivar-dependent significant differences, suggesting that hyperspectral imaging can be employed to measure variations in anthocyanin accumulations of different bok choy cultivars. Increased accumulation of anthocyanin under the stimulation condition for anthocyanin accumulation was observed in "purple magic" and "red stem" by both hyperspectral imaging and destructive analysis. In the different growth stages, no significant differences in anthocyanin accumulation were found in each cultivar by both hyperspectral imaging and destructive analysis. These results suggest that hyperspectral imaging can provide comparable analytic capability with destructive analysis to measure variations in anthocyanin accumulation that occurred under the different light and temperature conditions of the indoor farm. Leaf image analysis measuring the percentage of purple color area in the total leaf area displayed successful classification of anthocyanin accumulation in four bok choy cultivars in comparison to hyperspectral imaging and destructive analysis, but it also showed limitation to reflect the level of color saturation caused by anthocyanin accumulation under different environmental conditions in "red stem," "white stem," and "green stem." Finally, our hyperspectral imaging system was modified to be applied onto the high-throughput plant phenotyping system, and its test to analyze the variation of anthocyanin accumulation in four cultivars showed comparable results with the result of the destructive analysis.

Managing flowering time in Miscanthus and sugarcane to facilitate intra- and intergeneric crosses.

Miscanthus is a close relative of Saccharum and a potentially valuable genetic resource for improving sugarcane. Differences in flowering time within and between Miscanthus and Saccharum hinders intra- and interspecific hybridizations. A series of greenhouse experiments were conducted over three years to determine how to synchronize flowering time of Saccharum and Miscanthus genotypes. We found that day length was an important factor influencing when Miscanthus and Saccharum flowered. Sugarcane could be induced to flower in a central Illinois greenhouse using supplemental lighting to reduce the rate at which days shortened during the autumn and winter to 1 min d-1, which allowed us to synchronize the flowering of some sugarcane genotypes with Miscanthus genotypes primarily from low latitudes. In a complementary growth chamber experiment, we evaluated 33 Miscanthus genotypes, including 28 M. sinensis, 2 M. floridulus, and 3 M. ×giganteus collected from 20.9° S to 44.9° N for response to three day lengths (10 h, 12.5 h, and 15 h). High latitude-adapted M. sinensis flowered mainly under 15 h days, but unexpectedly, short days resulted in short, stocky plants that did not flower; in some cases, flag leaves developed under short days but heading did not occur. In contrast, for M. sinensis and M. floridulus from low latitudes, shorter day lengths typically resulted in earlier flowering, and for some low latitude genotypes, 15 h days resulted in no flowering. However, the highest ratio of reproductive shoots to total number of culms was typically observed for 12.5 h or 15 h days. Latitude of origin was significantly associated with culm length, and the shorter the days, the stronger the relationship. Nearly all entries achieved maximal culm length under the 15 h treatment, but the nearer to the equator an accession originated, the less of a difference in culm length between the short-day treatments and the 15 h day treatment. Under short days, short culms for high-latitude accessions was achieved by different physiological mechanisms for M. sinensis genetic groups from the mainland in comparison to those from Japan; for mainland accessions, the mechanism was reduced internode length, whereas for Japanese accessions the phyllochron under short days was greater than under long days. Thus, for M. sinensis, short days typically hastened floral induction, consistent with the expectations for a facultative short-day plant. However, for high latitude accessions of M. sinensis, days less than 12.5 h also signaled that plants should prepare for winter by producing many short culms with limited elongation and development; moreover, this response was also epistatic to flowering. Thus, to flower M. sinensis that originates from high latitudes synchronously with sugarcane, the former needs day lengths >12.5 h (perhaps as high as 15 h), whereas that the latter needs day lengths <12.5 h.

Flow cytometric fluorescence pulse width analysis of etoposide-induced nuclear enlargement in HCT116 cells.

Fluorescence pulse width can provide size information on the fluorescence-emitting particle, such as the nuclei of propidium iodide-stained cells. To analyze nuclear size in the present study, rather than perform the simple doublet discrimination approach usually employed in flow cytometric DNA content analyses, we assessed the pulse width of the propidium iodide fluorescence signal. The anti-cancer drug etoposide is reportedly cytostatic, can induce a strong G2/M arrest, and results in nuclear enlargement. Based on these characteristics, we used etoposide-treated HCT116 cells as our experimental model system. The fluorescence pulse widths (FL2-W) of etoposide-treated (10 microM, 48 h) cells were distributed at higher positions than those of vehicle control, so the peak FL2-W value of etoposide-treated cells appeared at 400 while those of vehicle control cells appeared at 200 and 270. These results were consistent with our microscopic observations. This etoposide-induced increase in FL2-W was more apparent in G2/M phase than other cell cycle phases, suggesting that etoposide-induced nuclear enlargement preferentially occurred in G2/M phase cells rather than in G0/G1 or S phase cells.

Inhibitory Mechanisms of Water Extract of Oplopanax elatus on Lipopolysaccharide-Induced Inflammatory Responses in RAW 264.7 Murine Macrophage Cells.

OBJECTIVE: To study the anti-inflammatory action and cellular mechanism of Oplopanax elatus. METHODS: A hot water extract of OE (WOE) was prepared and a major constituent, syringin, was successfully isolated. Its content in WOE was found to be 214.0 µg/g dried plant (w/w). Their anti-inflammatory activities were examined using RAW 264.7 macrophages and a mouse model of croton oil-induced ear edema. RESULTS: In lipopolysaccharide (LPS)-treated RAW 264.7 cells, a mouse macrophage cell line, WOE was found to significantly and strongly inhibit cyclooxygenase-2 (COX-2)-induced prostaglandin E2 (PGE2) production [half maximal inhibitory concentration (IC50)=135.2 µg/mL] and inducible nitric oxide synthase (iNOS)-induced NO production (IC50=242.9 µg/mL). In the same condition, WOE was revealed to inhibit NO production by down-regulating iNOS expression, mainly by interrupting mitogen activated protein kinases (MAPKs)/activator protein-1 (AP-1) pathway. The activation of all three major MAPKs, p38 MAPK, extracellular signal-regulated kinase (ERK), and c-Jun N-terminal kinase, was inhibited by WOE (50-300 µg/mL). On the other hand, WOE reduced PGE2 production by inhibiting COX-2 enzyme activity, but did not affect COX-2 expression levels. In addition, WOE inhibited the production of proinflammatory cytokines such as interleukin-6 and tumor necrosis factor-α. In croton oil-induced ear edema in mice, oral administration of WOE (50-300 mg/kg) dose-dependently inhibited edematic inflammation. CONCLUSION: Water extract of OE exhibited multiple anti-inflammatory action mechanisms and may have potential for treating inflammatory disorders.

Genome biology of the paleotetraploid perennial biomass crop Miscanthus.

Miscanthus is a perennial wild grass that is of global importance for paper production, roofing, horticultural plantings, and an emerging highly productive temperate biomass crop. We report a chromosome-scale assembly of the paleotetraploid M. sinensis genome, providing a resource for Miscanthus that links its chromosomes to the related diploid Sorghum and complex polyploid sugarcanes. The asymmetric distribution of transposons across the two homoeologous subgenomes proves Miscanthus paleo-allotetraploidy and identifies several balanced reciprocal homoeologous exchanges. Analysis of M. sinensis and M. sacchariflorus populations demonstrates extensive interspecific admixture and hybridization, and documents the origin of the highly productive triploid bioenergy crop M. × giganteus. Transcriptional profiling of leaves, stem, and rhizomes over growing seasons provides insight into rhizome development and nutrient recycling, processes critical for sustainable biomass accumulation in a perennial temperate grass. The Miscanthus genome expands the power of comparative genomics to understand traits of importance to Andropogoneae grasses.