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Background Fiber quality is an important economic trait of cotton, and its improvement is a major goal of cotton breeding. To better understand the genetic mechanisms responsible for fiber quality traits, we conducted a genome-wide association study to identify and mine fiber-quality-related quantitative trait loci (QTLs) and genes. Results In total, 42 single nucleotide polymorphisms (SNPs) and 31 QTLs were identified as being significantly associated with five fiber quality traits. Twenty-five QTLs were identified in previous studies, and six novel QTLs were firstly identified in this study. In the QTL regions, 822 genes were identified and divided into four clusters based on their expression profiles. We also identified two pleiotropic SNPs. The SNP locus i52359Gb was associated with fiber elongation, strength, length and uniformity, while i11316Gh was associated with fiber strength and length. Moreover, these two SNPs were nonsynonymous and located in genes Gh_D09G2376 and Gh_D06G1908, respectively. RT-qPCR analysis revealed that these two genes were preferentially expressed at one or more stages of cotton fiber development, which was consistent with the RNA-seq data. Thus, Gh_D09G2376 and Gh_D06G1908 may be involved in fiber developmental processes. Conclusions The findings of this study provide insights into the genetic bases of fiber quality traits, and the identified QTLs or genes may be applicable in cotton breeding to improve fiber quality.

Gene flow patterns and the genetic structure of domesticated crops like cotton are not well understood. Furthermore, marker-assisted breeding of cotton has lagged far behind that of other major crops because the loci associated with cotton traits such as fiber yield and quality have scarcely been identified. In this study, we used 19 microsatellites to first determine the population genetic structure and patterns of gene flow of superior germplasm resources in upland cotton. We then used association analysis to identify which markers were associated with 15 agronomic traits (including ten yield and five fiber quality traits). The results showed that the upland cotton accessions have low levels of genetic diversity (polymorphism information content = 0.427), although extensive gene flow occurred among different ecological and geographic regions. Bayesian clustering analysis indicated that the cotton resources used in this study did not belong to obvious geographic populations, which may be the consequence of a single source of domestication followed by frequent genetic introgression mediated by human transference. A total of 82 maker–trait associations were examined in association analysis and the related ratios for phenotypic variations ranged from 3.04% to 47.14%. Interestingly, nine SSR markers were detected in more than one environmental condition. In addition, 14 SSR markers were co-associated with two or more different traits. It was noteworthy that NAU4860 and NAU5077 markers detected at least in two environments were simultaneously associated with three fiber quality traits (uniformity index, specific breaking strength and micronaire value). In conclusion, these findings provide new insights into the population structure and genetic exchange pattern of cultivated cotton accessions. The quantitative trait loci of domesticated cotton identified will also be very useful for improvement of yield and fiber quality of cotton in molecular breeding programs.

Precise DNA sizing can boost sequencing efficiency, reduce cost, improve data quality, and even allow sequencing of low-input samples, while current pervasive DNA sizing approaches are incapable of differentiating DNA fragments under 200 bp with high resolution (20 bp). In non-invasive prenatal testing (NIPT), the size distribution of cell-free fetal DNA in maternal plasma (main peak at 143 bp) is significantly different from that of maternal cell-free DNA (main peak at 166 bp). The current pervasive workflow of NIPT and DNA sizing is unable to take advantage of this 20 bp difference, resulting in sample rejection, test inaccuracy, and restricted clinical utility. Here we report a simple, automatable, high-resolution DNA size enrichment workflow, named MiniEnrich, on a magnetic nano-platform to exploit this 20 bp size difference and to enrich fetal DNA fragments from maternal blood. Two types of magnetic nanoparticles were developed, with one able to filter high-molecular-weight DNA with high resolution and the other able to recover the remaining DNA fragments under the size threshold of interest with95% yield. Using this method, the average fetal fraction was increased from 13% to 20% after the enrichment, as measured by plasma DNA sequencing. This approach provides a new tool for high-resolution DNA size enrichment under 200 bp, which may improve NIPT accuracy by rescuing rejected non-reportable clinical samples, and enable NIPT earlier in pregnancy. It also has the potential to improve non-invasive screening for fetal monogenic disorders, differentiate tumor-related DNA in liquid biopsy and find more applications in autoimmune disease diagnosis.

Three new compounds, namely massonside C (1), massonianoside F (2), and 3, 8-dimethyl- herbacetin-7-O-β-D-glucopyranoside (3), together with five known compounds (4-8), were isolated from the fresh needles of Pinus massoniana. Their structures were established by 1D, 2D NMR, HRMS and comparison with the literature data. The absolute configuration of 1 was confirmed by a combination of X-ray single crystal analysis. All isolated compounds were evaluated for the protective effect of human umbilical vein endothelial cells against oxidative damage.

A genome-wide associated study identified six novel QTLs for lint percentage. Two candidate genes underlying this trait were also detected. Increasing lint percentage (LP) is a core goal of cotton breeding. To better understand the genetic basis of LP, a genome-wide association study (GWAS) was conducted using 276 upland cotton accessions planted in multiple environments and genotyped with a CottonSNP63K array. After filtering, 10,660 high-quality single-nucleotide polymorphisms (SNPs) were retained. Population structure, principal component and neighbor-joining phylogenetic tree analyses divided the accessions into two subpopulations. These results along with linkage disequilibrium decay indicated accessions were not highly structured and exhibited weak relatedness. GWAS uncovered 23 polymorphic SNPs and 15 QTLs significantly associated with LP, with six new QTLs identified. Two candidate genes, Gh_D05G0313 and Gh_D05G1124, both contained one significant SNP, highly expressed during ovule and fiber development stages, implying that the two genes may act as the most promising regulators of LP. Furthermore, the phenotypic value of LP was found to be positively correlated with the number of favorable SNP alleles. These favorable alleles for LP identified in the study may be useful for improving lint yield.

Cancer-induced bone pain is a severe and complex pain caused by metastases to bone in cancer patients. The aim of this study was to investigate the analgesic effect of scutellarin on cancer-induced bone pain in rat models by intrathecal injection of Walker 256 carcinoma cells. Mechanical allodynia was determined by paw withdrawal threshold in response to mechanical stimulus, and thermal hyperalgesia was indicated by paw withdrawal latency in response to noxious thermal stimulus. The paw withdrawal threshold and paw withdrawal latencies were significantly decreased after inoculation of tumor cells, whereas administration of scutellarin significantly attenuated tumor cell inoculation-induced mechanical and heat hyperalgesia. Tumor cell inoculation-induced tumor growth was also significantly abrogated by scutellarin. Ca2+/calmodulin-dependent protein kinase II is a multifunctional kinase with up-regulated activity in bone pain models. The activation of Ca2+/calmodulin-dependent protein kinase II triggers phosphorylation of cAMP-response element binding protein. Scutellarin significantly reduced the expression of phosphorylated-Ca2+/calmodulin-dependent protein kinase II and phosphorylated-cAMP-response element binding protein in cancer-induced bone pain rats. Collectively, our study demonstrated that scutellarin attenuated tumor cell inoculation-induced bone pain by down-regulating the expression of phosphorylated-Ca2+/calmodulin-dependent protein kinase II and phosphorylated-cAMP-response element binding protein. The suppressive effect of scutellarin on phosphorylated-Ca2+/calmodulin-dependent protein kinase II/phosphorylated-cAMP-response element binding protein activation may serve as a novel therapeutic strategy for CIBP management.

Objective To identify the key genes promoting the differentiation of melon non-embryogenic callus into embryogenic callus. Methods The transcriptome sequencing analysis was used to analyze the mRNA sequence in embryogenic callus (Z) and non-embryogenic callus (F); transcript mapping, gene expression analysis, cluster analysis, classification analysis and enrichment analysis were then used to detect the differentially expressed genes and enriched pathways. Results The correlation coefficient between sample Z and sample F was 0.929 after transcript mapping. The overall gene expression levels in sample Z were higher as compared with sample F. Furthermore, cluster analysis showed that the expression of genes involved in photosynthesis was increased in sample Z when comparing to F. Besides, the classification of differential Gene Ontology (GO) showed that many metabolic processes were affected with the metabolism enhanced in embryogenic callus. Interestingly, Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis further demonstrated the high metabolic activity and active secondary metabolite formation in the embryogenic callus. Conclusion The genes associated with photosynthesis, metabolic pathways and biosynthesis of secondary metabolites may promote the differentiation of callus into embryogenic callus.

The wall-associated kinases (WAKs) and WAK-like kinases (WAKLs) form a group of receptor-like kinases (RLKs) with extracellular domains tightly linked to the cell wall. The WAKs/WAKLs have been known to be involved in plant growth, development, and stress responses. However, the functions of WAKs/WAKLs are less well known in cotton. In this study, 58, 66, and 99 WAK/WAKL genes were identified in Gossypium arboreum, G. raimondii, and G. hirsutum, respectively. Phylogenetic analysis showed they were classified into five groups, with two groups specific to cotton. Collinearity analysis revealed that segmental and tandem duplications resulted in expansion of the WAK/WAKL gene family in cotton. Moreover, the Ka/Ks ratios indicated this family was exposed to purifying selection pressure during evolution. The structures of the GhWAK/WAKL genes and encoded proteins suggested the functions of WAKs/WAKLs in cotton were conserved. Transient expression of four WAK/WAKL-GFP fusion constructs in Arabidopsis protoplasts indicated that they were localized on the plasma membrane. The cis-elements in the GhWAK/WAKL promoters were responsive to multiple phytohormones and abiotic stresses. Expression profiling showed that GhWAK/WAKL genes were induced by various abiotic stresses. This study provides insights into the evolution of WAK/WAKL genes and presents fundamental information for further analysis in cotton.

Background: The third-generation genome editing system CRISPR/Cas had shown strong application prospects in crop genetic improvement. However, this technology largely depends on genetic transformation. Public concerns on GMO (genetically modified organisms) safety, as well as related regulations, have restricted the application. Therefore, establishing DNA-free transfection system is important to promote CRISPR/Cas genome editing in agriculture.Results：In this paper, cell penetrating peptides fusion protein (CPP-mCherry) was found to be effective on DNA-free transfection. DNA sequences of nine tandem arginine (R9), one cysteine (cys), reporter mCherry and histidine label were sequentially constructed into pET 45B+ expression vector and transformed into Escherichia coli BL21(DE3) strain. CPP-mCherry fusion protein can be induced by 1mM IPTG for at least 1 hour in 28 °C. CPP-mCherry fusion protein can be obtained by 200W ultrasonication, then purified by Ni column and MWCO dialysis. The Arabidopsis thaliana root tips and leaves, as well as Chinese cabbage microspores and 3-week-old microspore embryo can be used as transfected recipient. Concentration can be selected between 10-100μg/ml and incubated overnight at room temperature. R9-cys-mCherry protein can be translocated into the nucleus of microspore. The transfection efficiency of root tips reached 100% and of microspore and MDE was 8.13% and 94.79%, respectively. Conclusions: Here, a CPP mediated DNA-free transfection system was built in dicots. These results lay a technical foundation of DNA-free genome editing.

Tillering is an important biomass yield component trait in switchgrass (Panicum virgatum L.). Teosinte branched 1 (tb1)/Branched 1 (BRC1) gene is a known regulator for tillering/branching in several plant species; however, its role on tillering in switchgrass remains unknown. Here, we report physiological and molecular characterization of mutants created by CRISPR/Cas9. We successfully obtained nonchimeric Pvtb1a and Pvtb1b mutants from chimeric T0 mutants using nodal culture. The biallelic Pvtb1a-Pvtb1b mutant plants produced significantly more tillers and higher fresh weight biomass than the wild-type plants. The increased tiller number in the mutant plants resulted primarily from hastened outgrowth of lower axillary buds. Increased tillers were also observed in transgene-free BC1 monoallelic mutants for either Pvtb1a-Pvtb1b or Pvtb1b gene alone, suggesting Pvtb1 genes act in a dosage-dependent manner. Transcriptome analysis showed 831 genes were differentially expressed in the Pvtb1a-Pvtb1b double knockdown mutant. Gene Ontology analysis revealed downregulation of Pvtb1 genes affected multiple biological processes, including transcription, flower development, cell differentiation, and stress/defense responses in edited plants. This study demonstrates that Pvtb1 genes play a pivotal role in tiller production as a negative regulator in switchgrass and provides opportunities for further research aiming to elucidate the molecular pathway regulating tillering in switchgrass.

A new oribatid mite species of the family Pediculochelidae, Paralycus nortoni sp. nov., is described and illustrated based on adult specimens collected from barks of dead Pinus massoniana Lamb. infested by Monochamus alternatus in Fuzhou city, China. This new species described here is the fourth species of the genus Paralycus in China. An updated key to the species of the genus Paralycus is provided., Acarologia, 60, 481-487

Tillering is an important biomass yield component trait in switchgrass (Panicum virgatum L.).Teosinte branched 1(tb1)/Branched 1(BRC1) gene is a known regulator for tillering/branching in several plant species; however, its role on tillering in switchgrass remains unknown. Here, we report physiological and molecular characterization of mutants created by CRISPR/Cas9. We successfully obtained non-chimericPvtb1aandPvtb1bmutants from chimeric T0 mutants using nodal culture. The biallelicPvtb1a-Pvtb1bmutant plants produced significantly more tillers and higher fresh weight biomass than the wild-type plants. The increased tiller production in the mutant plants resulted primarily from hastened outgrowth of lower axillary buds. Increased tillers were also observed in transgene-free T1 monoallelic mutants for eitherPvtb1a-Pvtb1borPvtb1bgene alone, suggestingPvtb1genes act in a dosage-dependent manner. Transcriptome analysis showed 831 genes were differentially expressed in thePvtb1a-Pvtb1bdouble knockdown mutant. Gene Ontology analysis revealed downregulation ofPvtb1genes affected multiple biological processes, including transcription, flower development, cell differentiation, and stress/defense responses in edited plants. This study demonstrates thatPvtb1genes play a pivotal role in tiller production as a negative regulator in switchgrass and provides opportunities for further research aiming to elucidate the molecular pathway regulating tillering in switchgrass.HighlightSolid non-chimeric mutants were successfully isolated from CRISPR/Cas9-induced chimeric mutants using nodal culture.Teosinte branched 1(tb1) genes are involved in various pathways to regulate tillering in switchgrass.

Heparin is the most widely studied glycosaminoglycan. It plays an important role in regulating several normal physiological and pathological processes, including inhibition of thrombocytopenia, lipid regulation, metabolism and electrostatic attraction with various proteins. Hence, it is crucial to develop selective and sensitive assays for monitoring heparin levels in biological systems. In the present study, we designed a facile synthesis method to prepare near-infrared emitting denatured bovine serum albumin-templated copper nanoclusters (dBSA-Cu NCs) for the trace detection of heparin. In aqueous solution, the bovine serum albumin (BSA) was denatured by heating at a temperature of 70 °C to prepare dBSA, which was employed as the template for fabricating the dBSA-Cu NCs. Then, Cu2+ ions were directly reduced to dBSA-Cu NCs by hydrazine hydrate at room temperature. The synthetic process was very simple to control due to the lack of any complicated procedure, such as heating or adjusting pH. In addition, the fluorescence intensity of dBSA-Cu NCs at 642 nm was quenched by heparin samples. Hence, the dBSA-Cu NCs can be used as a probe for the trace detection of heparin with a very low limit of detection (LOD) of 0.26 ng mL−1 in a linear range from 1.25 ng mL−1 to 250 ng mL−1. Furthermore, the as-developed dBSA-Cu NCs were also successfully applied to determine heparin in human plasma samples with quantitative spike recoveries from 95% to 104%.

Generally, cesium ion (Cs+) which concentration of above the millimolar order is toxic to microorganisms. In this study we report the isolation and genomic characteristics of Bacillus sp. Cs-700, which can grow in media supplemented with 700 mM Cs+ and was one of the most Cs+ tolerating bacteria been reported. Bacillus sp. Cs-700 was isolated from the South China Sea, and comprised one circular chromosome of 4,297,839 bp (40.32 mol% G + C content), containing 4366 predicted protein-coding sequences. We predict that numerous genes related to high cesium tolerating, and the genome of Bacillus sp. Cs-700 will be helpful for further insights into the mechanism of bacterial resistance to excess Cs+ conditions and the bioremediation of radiocesium-polluting environments.

Summary Upland cotton (Gossypium hirsutum) is the world's largest source of natural fibre and dominates the global textile industry. Hybrid cotton varieties exhibit strong heterosis that confers high fibre yields, yet the genome‐wide effects of artificial selection that have influenced Upland cotton during its breeding history are poorly understood. Here, we resequenced Upland cotton genomes and constructed a variation map of an intact breeding pedigree comprising seven elite and 19 backbone parents. Compared to wild accessions, the 26 pedigree accessions underwent strong artificial selection during domestication that has resulted in reduced genetic diversity but stronger linkage disequilibrium and higher extents of selective sweeps. In contrast to the backbone parents, the elite parents have acquired significantly improved agronomic traits, with an especially pronounced increase in the lint percentage. Notably, identify by descent (IBD) tracking revealed that the elite parents inherited abundant beneficial trait segments and loci from the backbone parents and our combined analyses led to the identification of a core genomic segment which was inherited in the elite lines from the parents Zhong 7263 and Ejing 1 and that was strongly associated with lint percentage. Additionally, SNP correlation analysis of this core segment showed that a non‐synonymous SNP (A‐to‐G) site in a gene encoding the cell wall‐associated receptor‐like kinase 3 (GhWAKL3) protein was highly correlated with increased lint percentage. Our results substantially increase the valuable genomics resources available for future genetic and functional genomics studies of cotton and reveal insights that will facilitate yield increases in the molecular breeding of cotton.

Biofuel production in the midwestern United States has largely focused on corn (Zea mays L.) grain for ethanol production and more recently, corn stover for lignocellulosic ethanol. Tropical corn refers to corn adapted from tropical regions. Growing tropical germplasm in temperate environments is not attractive for grain yield, but shows promise for biomass production through taller and more vigorous plants with thick stems and long leaves. The comparison between tropical corn populations and their temperate adapted counterparts with a focus on biomass production has not yet been explored under Iowa growing conditions. This study refines the accuracy of the leaf collar method, and characterizes the adaptation of the populations by evaluating crop development and biomass production. Therefore, field trials were established in central Iowa during the 2014 and 2015 growing seasons. Adapted and non-adapted versions of three populations, Tuxpeno, Suwan, and Tuson, were planted at three dates to evaluate their phenology and potential for biomass production under midwestern U.S. conditions. Plant height correlated well with vegetative development and total biomass. Adapted tropical corn had advanced grain development, while non-adapted tropical corn had advanced vegetative development and greater biomass yield. Non-adapted tropical corn flowered later, had 22% greater total biomass on average (4.6 Mg ha–¹ greater than adapted), and had 43% greater non-grain biomass on average, up to 20 Mg ha–¹ (6 Mg ha–¹ greater than adapted), while adapted populations yielded more grain. Non-adapted tropical corn has a high potential as feedstock for Gen2 biofuels in central Iowa.

Summary The CRISPR/Cas9 system has become a powerful tool for targeted mutagenesis. Switchgrass (Panicum virgatum L.) is a high yielding perennial grass species that has been designated as a model biomass crop by the U.S. Department of Energy. The self‐infertility and high ploidy level make it difficult to study gene function or improve germplasm. To overcome these constraints, we explored the feasibility of using CRISPR/Cas9 for targeted mutagenesis in a tetraploid cultivar ‘Alamo’ switchgrass. We first developed a transient assay by which a non‐functional green‐fluorescent protein gene containing a 1‐bp frameshift insertion in its 5′ coding region was successfully mutated by a Cas9/sgRNA complex resulting in its restored function. Agrobacterium‐mediated stable transformation of embryogenic calli derived from mature caryopses averaged a 3.0% transformation efficiency targeting the genes of teosinte branched 1(tb1)a and b and phosphoglycerate mutase (PGM). With a single construct containing two sgRNAs targeting different regions of tb1a and tb1b genes, primary transformants (T0) containing CRISPR/Cas9‐induced mutations were obtained at frequencies of 95.5% (tb1a) and 11% (tb1b), respectively, with T0 mutants exhibiting increased tiller production. Meanwhile, a mutation frequency of 13.7% was obtained for the PGM gene with a CRISPR/Cas9 construct containing a single sgRNA. Among the PGM T0 mutants, six are heterozygous and one is homozygous for a 1‐bp deletion in the target region with no apparent phenotypical alterations. We show that CRISPR/Cas9 system can generate targeted mutagenesis effectively and obtain targeted homozygous mutants in T0 generation in switchgrass, circumventing the need of inbreeding.

The Midwestern U.S. landscape is one of the most highly altered and intensively managed ecosystems in the country. The predominant crops grown are maize (Zea mays L.) and soybean [Glycine max (L.) Merr]. They are typically grown as monocrops in a simple yearly rotation or with multiple years of maize (2 to 3) followed by a single year of soybean. This system is highly productive because the crops and management systems have been well adapted to the regional growing conditions through substantial public and private investment. Furthermore, markets and supporting infrastructure are highly developed for both crops. As maize and soybean production have intensified, a number of concerns have arisen due to the unintended environmental impacts on the ecosystem. Many areas across the Midwest are experiencing negative impacts on water quality, soil degradation, and increased flood risk due to changes in regional hydrology. The water quality impacts extend even further downstream. We propose the development of an innovative system for growing maize and soybean with perennial groundcover to recover ecosystem services historically provided naturally by predominantly perennial native plant communities. Reincorporating perennial plants into annual cropping systems has the potential of restoring ecosystem services without negatively impacting grain crop production and offers the prospect of increasing grain crop productivity through improving the biological functioning of the system.

Cotton is one of the most important economic crops worldwide. As the global demands rising, cotton yield improvement is the most important goal of cotton breeding. Hybrids have great potential for increasing yield, however, the genetic mechanism of hybrids is still not clear. To investigate the genetic basis of cotton hybrids, we resequenced 9053 and sGK9708 with 62.13x coverage depth, the parents of the elite hybrid cotton CCRI63 that has obvious heterosis in lint percentage (LP) and boll weight (BW). Based on the cotton reference genome (TM-1), 1,287,661 single nucleotide polymorphisms (SNPs) and 152,479 insertions/deletions (InDels) were identified in 9053, and 1,482,784 SNPs and 152,985 InDels in sGK9708. Among them, 8649 SNPs and 629 InDels in the gene coding regions showed polymorphism between parents. Moreover, these variations involved 5092 genes, and 3835 of these genes were divided into 10 clusters based on the gene expression profiles. The genes in Cluster 3 and 7 were specifically expressed in the ovule and fiber development stage, suggesting that they might relate to LP and BW. We further co-localized the polymorphic SNPs and InDels with the reported quantitative trait loci (QTLs) of LP and BW, and identified 68 genes containing the polymorphic SNPs or InDels within these QTL intervals and as being related to fiber development. This suggested that the outstanding traits of CCRI63 such as LP and BW might be generated by accumulating the favorable variations from the parents. The results generated herein provide a genetic basis for cotton hybrids and genetic markers for marker-assisted selection breeding of cotton.

Feedstock quality mainly depends upon the biomass composition and bioenergy conversion system being used. Higher cellulose and hemicellulose concentrations are desirable for biochemical conversion, whereas higher lignin is favored for thermochemical conversion. The efficiency of these conversion systems is influenced by the presence of high nitrogen and ash concentrations. Switchgrass ( Panicum virgatum L.) varieties are classified into two ecotypes based on their habitat preferences, i.e., upland and lowland. The objectives of this study were to quantify the chemical composition of switchgrass varieties as influenced by harvest management, and to determine if ecotypic differences exist among them. A field study was conducted near Ames, IA during 2012 and 2013. Upland (‘Cave-in-Rock’, ‘Trailblazer’ and ‘Blackwell’) and lowland switchgrass varieties (‘Kanlow’ and ‘Alamo’) were grown in a randomized block design with six replications. Six biomass harvests were collected at approximately 2-week intervals each year. In both years, delaying harvest increased cellulose, hemicellulose and lignin concentrations while decreasing nitrogen and ash concentrations in all varieties. On average, Kanlow had the highest cellulose and hemicellulose concentration (354 and 321 g kg −1 DM respectively), and Cave-in-Rock had the highest lignin concentration (33 g kg −1 DM). The lowest nitrogen and ash concentrations were observed in Kanlow (14 and 95 g kg −1 DM respectively). In general, our results indicate that delaying harvest until fall improves feedstock quality, and ecotypic differences do exist between varieties for important feedstock quality traits. These findings also demonstrate potential for developing improved switchgrass cultivars as bioenergy feedstock by intermating lowland and upland ecotypes.

As the dominant and unique species in Lake Qinghai, the naked carp Gymnocypris przewalskii (Kessler) is characterized by upstream migration to spawn during late spring and summer in each year and back lake to live during the remaining seasons, but little is known about its migratory pattern. The migratory pattern of the naked carp is significant in understanding its living habitat and in protecting this precious fish. Otoliths are paired biogenic carbonates in the inner ear of teleost fish and grow layer by layer throughout the fish life. Owing to its metabolically inert without involvement in metabolic processes after deposition, chemical compositions of daily and annual growth increments in otoliths record the water elements of the ambient waters where the fish encountered during its life history. Therefore, microchemistry variation of otoliths has been used widely to trace migration pattern of fish and to reconstruct water chemistry and environmental conditions in the past.

Soil salinization, which is primarily due to excessive Na+ levels, is a major abiotic stress adversely affecting plant growth and development. The Na+/H+ antiporter (NHX) is a transmembrane protein mediating the transport of Na+ or K+ and H+ across the membrane to modulate the ionic balance of plants in response to salt stress. Research regarding NHXs has mainly focused on the vacuolar-type NHX family members. However, the biological functions of the endosomal-type NHXs remain relatively uncharacterized. In this study, 22 NHX family members were identified in Gossypium hirsutum. A phylogenetic analysis divided the GhNHX genes into two categories, with 18 and 4 in the vacuolar and endosomal groups, respectively. The chromosomal distribution of the NHX genes revealed the significant impact of genome-wide duplication during the polyploidization process on the number of GhNHX genes. Analyses of gene structures and conserved motifs indicated that GhNHX genes in the same phylogenetic cluster are conserved. Additionally, the salt-induced expression patterns confirmed that the expression levels of most of the GhNHX genes are affected by salinity. Specifically, in the endosomal group, GhNHX4A expression was substantially up-regulated by salt stress. A yeast functional complementation test proved that GhNHX4A can partially restore the salt tolerance of the salt-sensitive yeast mutant AXT3. Silencing GhNHX4A expression decreased the resistance of cotton to salt stress because of an increase in the accumulation of Na+ in stems and a decrease in the accumulation of K+ in roots. The results of this study may provide the basis for an in-depth characterization of the regulatory functions of NHX genes related to cotton salt tolerance, especially the endosomal-type GhNHX4A. Furthermore, the presented data may be useful for selecting appropriate candidate genes for the breeding of new salt-tolerant cotton varieties.

Sucrose transporters (SUTs) play key roles in allocating the translocation of assimilates from source to sink tissues. Although the characteristics and biological roles of SUTs have been intensively investigated in higher plants, this gene family has not been functionally characterized in cotton. In this study, we performed a comprehensive analysis of SUT genes in the tetraploid cotton Gossypium hirsutum. A total of 18 G. hirsutum SUT genes were identified and classified into three groups based on their evolutionary relationships. Up to eight SUT genes in G. hirsutum were placed in the dicot-specific SUT1 group, while four and six SUT genes were, respectively, clustered into SUT4 and SUT2 groups together with members from both dicot and monocot species. The G. hirsutum SUT genes within the same group displayed similar exon/intron characteristics, and homologous genes in G. hirsutum At and Dt subgenomes, G. arboreum, and G. raimondii exhibited one-to-one relationships. Additionally, the duplicated genes in the diploid and polyploid cotton species have evolved through purifying selection, suggesting the strong conservation of SUT loci in these species. Expression analysis in different tissues indicated that SUT genes might play significant roles in cotton fiber elongation. Moreover, analyses of cis-acting regulatory elements in promoter regions and expression profiling under different abiotic stress and exogenous phytohormone treatments implied that SUT genes, especially GhSUT6A/D, might participate in plant responses to diverse abiotic stresses and phytohormones. Our findings provide valuable information for future studies on the evolution and function of SUT genes in cotton.

The SWEET (sugars will eventually be exported transporters) proteins are sugar efflux transporters containing the MtN3_saliva domain, which affects plant development as well as responses to biotic and abiotic stresses. These proteins have not been functionally characterized in the tetraploid cotton, Gossypium hirsutum, which is a widely cultivated cotton species. In this study, we comprehensively analyzed the cotton SWEET gene family. A total of 55 putative G. hirsutum SWEET genes were identified. The GhSWEET genes were classified into four clades based on a phylogenetic analysis and on the examination of gene structural features. Moreover, chromosomal localization and an analysis of homologous genes in Gossypium arboreum, Gossypium raimondii, and G. hirsutum suggested that a whole-genome duplication, several tandem duplications, and a polyploidy event contributed to the expansion of the cotton SWEET gene family, especially in Clade III and IV. Analyses of cis-acting regulatory elements in the promoter regions, expression profiles, and artificial selection revealed that the GhSWEET genes were likely involved in cotton developmental processes and responses to diverse stresses. These findings may clarify the evolution of G. hirsutum SWEET gene family and may provide a foundation for future functional studies of SWEET proteins regarding cotton development and responses to abiotic stresses.

High rate of platelet adhesion and limited biocompatibility were regarded as the chief weaknesses of vascular tissue engineering scaffolds. The objective of this research was to overcome these drawbacks by ultilizing coaxial electrospinning to combine CDPS with PLA. CDPS and PLA were located at the inner and external layer respectively so that a “core-sheath” structure was formed. The properties of scaffolds were tested by methods such as mechanical testing, MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] assay and subcutaneous transplantation. Compared to natural tissues, CDPS/PLA coaxial scaffolds showed excellent biomechanic properties and hemocompatibility so that CDPS owned good potentiality in vascular tissue engineering.

Switchgrass is native to the tallgrass prairie of North America. It is self-incompatible and has varied ploidy levels from diploid (2x) to dodecaploid (12x) with tetraploid and octoploid being the most common. The high yielding potential and the ability to grow well in marginal lands make switchgrass an ideal species as a dedicated biomass producer for lignocellulosic ethanol production. Genetic transformation is an important tool for studying gene function and for germplasm improvement in switchgrass, the genome of which has been sequenced recently. This paper intends to provide a comprehensive review on plant regeneration and genetic transformation in switchgrass. We first reviewed the effect of explants, basal medium and plant growth regulators on plant regeneration in switchgrass, which is a prerequisite for genetic transformation. We then reviewed the progresses on genetic transformation with either the biolistic or Agrobacterium-mediated method in switchgrass, and discussed various techniques employed to improve the transformation efficiency. Finally we reviewed the recent progresses on the use of genetic transformation in improving biomass quality such as the reduction of lignin, and in increasing biomass yield in switchgrass. We also provided a future perspective on the use of new genome editing technologies in switchgrass and its potential impact on regulatory processes.

Isolariciresinol-9'-O-α-L-arabinofuranoside (MWS‑19) isolated from Pinus massoniana Lamb. Fresh pine needles is the major ingredient of the Songling Xuemaikang capsule therapy used for hypertension. The present study aimed to investigate the effects and underlying mechanisms of MWS‑19 on hydrogen peroxide (H2O2)‑induced apoptosis in human umbilical vein endothelial cells (HUVECs). To investigate the effect of MWS‑19 on apoptosis in HUVECs, an oxidative stress‑induced apoptosis model was established in HUVECs using H2O2, and the present study performed Hoechst 33258 staining and a Cell Counting Kit‑8 (CCK‑8) assay. Furthermore, western blot analysis was also performed to investigate the underlying mechanism of the effects of MWS‑19 on the model. The results demonstrated that MWS‑19 reversed the effects of H2O2 on cell apoptosis at a concentration range of 15.6‑250 µg/ml, with dose‑dependent increases in cell growth. Hoechst staining indicated that 500 µM H2O2 induced HUVEC apoptosis, and MWS‑19 markedly protected HUVECs against apoptosis at 31.3, 62.5 and 125 µg/ml. Furthermore, the protein expression of phosphatidylinositol 3‑kinase (PI3K), phosphorylated‑Akt and Bcl‑2‑associated agonist of cell death (Bad) were increased, and reduced caspase‑3 activation was observed, following treatment with MWS‑19 in H2O2‑treated HUVECs. Additionally, the PI3K inhibitor wortmannin attenuated PI3K/Akt/Bad signaling induced by MWS‑19 treatment and neutralized the effect of MWS‑19 on the growth of HUVECs. In conclusion, the results of the present study indicate that MWS‑19 may protect against H2O2‑induced HUVEC apoptosis via the PI3K/Akt/Bad signaling pathway. MWS‑19 may serve an important role in the prevention of oxidative damage in vascular endothelial cells in hypertension patients.

Freezing stress substantially reduces crop yields and limits plant distribution. The identification of genes critical for cold acclimation is thus of great importance. C-repeat binding factors (CBFs) are transcription factors that play key regulatory roles in the cold acclimation process, which dramatically increases freezing tolerance in plants. We report here that B. distachyon can successfully cold acclimate and we identified a CBF gene family consisting of eight genes in a tandem array and are designated as BdCBF1-8. Expression analysis indicated that all the eight BdCBF genes are induced by cold. Freezing tolerance experiments showed that the knockdown of BdCBF3 gene in RNAi cbf3 mutant plants results in a significant reduction in survival after an exposure to freezing temperatures. RNA-seq transcriptomic analysis was conducted using the wild type and cbf3 mutant plants under both normal and cold conditions. We identified 460, 3213, 2839 and 1871 differentially expressed genes exhibiting different expression levels by pairwise comparisons of cbf3 (23°C) vs. WT (23°C), WT (23°C) vs. WT (4°C), cbf3 (23°C) vs. cbf3 (4°C), and cbf3 (4°C) vs. WT (4°C), respectively. These differentially expressed genes were enriched in several biological pathways. Combined analyses of differentially expressed genes in some of the enriched pathways provide insights into mechanisms of plant response to cold in the BdCBF3-dependent, -independent or -compensatory categories.

Diabetic cystopathy is a common complication of voiding disorders in diabetes mellitus. Neuropathy and bladder remodeling underlie the lack of efficacy of pharmacological and surgical treatments. Previous studies have shown that decreased levels of nerve growth factor (NGF) are closely associated with disease progression. Besides, application of human umbilical cord mesenchymal stem cells (hUC-MSCs) is also considered a promising therapeutic strategy for treatment of diabetic neuropathy. In our study, we determine the therapeutic efficacy and mechanisms of hUC-MSCs which transfected with NGF geen in ameliorating diabetic cystopathy for the first time. We transducted hUC-MSCs with NGF-expressing lentivirus so that the hUC-MSCs can express NGF efficiently, then the NGF-expressing hUC-MSCs were intrathecally administrated in L6-S1 spinal cord of diabetic rats 3 days after induced by streptozotocin. Nine weeks later, the level of neurotrophins and voiding function of bladder were detected. Results show that improvements in voiding function were related to the neurotrophins and cytokines released by the intrathecally transplanted hUC-MSCs. In addition, the hUC-MSCs also differentiated into neurons and astrocytes within the spinal cord in rats. These two mechanisms play a combined role in neural regeneration and the amelioration of the symptoms of diabetic cystopathy.

All-trans retinoic acid (t-RA) is a natural component and representative physiologically active metabolite of vitamin A, having multiple physiologic functions. The objective of this study was to evaluate the effect of t-RA on goat oocyte maturation and cumulus cell apoptosis during in vitro maturation (IVM). Immature goat cumulus-oocyte complexes (COCs) were matured in vitro in the absence or presence of t-RA at concentrations of 10 nmol/L, 100 nmol/L and 1000 nmol/L. Oocyte maturation and embryo development were assessed by polar body formation and parthenogenetic activation, respectively. Oocyte survival was checked by Trypan blue staining. Apoptosis of cumulus cells was analyzed by terminal deoxynucleotidyl transferase nick end labeling staining and quantitative real-time PCR. In comparison with the control group, 100 nmol/L and 10 nmol/L t-RA significantly improved goat nuclear oocyte maturation and survival (P

Sustainable development of the bioenergy industry will depend upon the amount and quality of bioenergy feedstock produced. Switchgrass (Panicum virgatum L.) is a model lignocellulosic bioenergy crop but critical information is lacking for improved management, growth, and development simulation model calibration. A field study was conducted near Ames, IA during 2012–2013 with the objective to evaluate upland (“Cave-in-Rock”, ‘Trailblazer’ and ‘Blackwell’) and lowland (“Kanlow” and “Alamo”) switchgrass ecotypes for harvest timing on morphology (i.e., phenology, leaf area index (LAI), and biomass yield). The experiment used a randomized complete block design, with three upland and two lowland varieties harvested at six dates annually. In both years, delaying harvest to later maturity increased biomass yield; lowland cultivars produced greater biomass yield (6.15 tons ha−1) than upland ecotypes (5.10 tons ha−1). Lowland ecotypes had delayed reproductive development compared with upland ecotypes. At the end of both growing seasons, upland ecotypes had greater mean stage count (MSC) than lowland ecotypes. “Cave-in-Rock” had greatest MSC and LAI, but did not produce the greatest biomass. Relationships were nonlinear between MSC and biomass yield, with significant cultivar–year interaction. The relationship between biomass yield and MSC will be useful for improving switchgrass, including cultivar selection, fertilizer application, and optimum harvest time.

Reduced-growth, glyphosate-resistant Kentucky bluegrass (Poa pratensis L.) has recently been developed. There has been no comparative analysis of the newly developed transgenic Kentucky bluegrass lines to conventional cultivars or tissue culture-derived lines to determine vegetative and flower characteristic differences and similarities. This study measured the vegetative and reproductive characteristics of the transgenic lines, tissue culture-derived control lines, and reference cultivars during 2004 and 2005, in Ames, IA. The study was conducted in a noncompetitive, nonmown, spaced plant setting. Data collection of vegetative characteristics included rhizome spread and total plant area. Data collection for flower characteristics included flower phenology and morphological characteristic measurements, including panicle length, number of spikelets per panicle, and flag leaf characteristics. Transgenic lines as a group exhibited 29 and 32% less rhizome spread than the reference cultivars and tissue culture-derived control lines, respectively, in 2004 and 37 and 40% less rhizome spread than reference cultivars and tissue culture-derived control lines, respectively, in 2005. Flower emergence times for all transgenic lines were comparable to their respective tissue culture-derived control lines and reference cultivars.

Big bluestem ( Andropogon gerardii Vitman) and little bluestem [ Schizachyrium scoparium (Michaux) Nash.] are native to the North America and are important forage grasses and ornamental grasses. Both grasses are proposed as ideal biomass producers for cellulosic ethanol production. To apply genetic transformation, which is an important tool for incorporating desirable agronomic traits into plants to both species, however requires an efficient and reproducible regeneration protocol. We used mature caryopses from big and little bluestem as explants and tested the effect of various combinations of 2, 4-dichlorophenoxyacetic acid (2, 4-D) (1, 2, 3, 4 or 5 mg l −1 ) and kinetin (KT) (0, 0.1 or 0.2 mg l −1 ) on embryogenic callus induction with LS as the basal medium. The highest percentage of embryogenic calli induction occurred on medium containing 2, 4-D alone at 2 mg l −1 for ‘Bison’ and on medium containing 4 mg l −1 2, 4-D alone for ‘Bonilla’ big bluestem. For little bluestem, the highest percentage of embryogenic callus induction occurred on medium containing 3 mg l −1 2, 4-D plus 0.1 mg l −1 kinetin, suggesting that addition of KT is beneficial. Shoot regeneration took place on LS basal medium without any plant growth regulator for both species, although the addition of KT increased both regeneration frequency and the number of shoots produced per callus. Rooting of shoots reaching about 2 cm long occurred readily with or without α-naphthaleneacetic acid (NAA). Rooted plantlets were all successfully established in the soil.

Indiangrass [ Sorghastrum nutans (L.) Nash.] is native to the North America and is an important component of the original tall grass prairie. It is also an important ornamental and forage grass. Recently, it has been proposed as an ideal biomass producer for cellulosic ethanol production. Genetic transformation is an important tool for introducing important agronomic traits into plants, but an efficient and reproducible in vitro regeneration protocol is a prerequisite for successful genetic transformation. In this report, we used mature caryopses as explants and tested the effect of various combinations of 2,4-dichlorophenoxyacetic acid (2,4-D) (1–5) and kinetin (KT) (0, 0.1, and 0.2) on embryogenic callus induction using LS basal medium. Caryopses cultured on media supplemented with 2,4-D alone generally outperformed those cultured on media supplemented with both 2,4-D and kinetin for embryogenic callus induction. The best treatment is LS basal medium supplemented with 3 mg l −1 2,4-D. LS basal medium supplemented with KT of 0, 0.5, 1, 2 or 5 mg l −1 were tested for regeneration efficiency which was shown to increase as the KT concentration increased. The quality of the shoots produced on the medium containing KT at 5 mg l −1 , which produced the highest regeneration frequency appeared to be lower as leaves become vitrified. Shoots were moved to a rooting medium containing either 0 or 0.1 mg l −1 α-naphthaleneacetic acid (NAA). Rooted plantlets were then transferred to soil-containing pots and were placed in a mist room for 1 week before they are transferred to a normal greenhouse where they all survived. The reported regeneration protocol is very efficient and highly reproducible in spite of the heterogeneous nature of the tested cultivar; thus it should be suitable for genetic transformation.

Osmopriming with PEG has potential to improve seed germination, seedling emergence, and establishment, especially under stress conditions. This research investigated germination performance, seedling establishment, and effects of osmopriming with PEG on physiology in sorghum seedlings and their association with post-priming stress tolerance under various soil moisture stress conditions. Results showed that seed priming increased the environmental range suitable for sorghum germination and has potential to provide more uniform and synchronous emergence. Physiologically, seed priming strengthened the antioxidant activities of APX, CAT, POD, and SOD, as well as compatible solutes including free amino acid, reducing sugar, proline, soluble sugar, and soluble protein contents. As a result, seed priming reduced lipid peroxidation and stabilized the cell membrane, resulting in increased stress tolerance under drought or excessive soil moisture environments. Overall, results suggested that seed priming with PEG was effective in improving seed germination and seedling establishment of sorghum under adverse soil moisture conditions. Osmopriming effectively strengthened the antioxidant system and increased osmotic adjustment, likely resulting in increased stress tolerance.

Winter hardiness is a quantitative trait and the lack of it limits geographic distribution of ryegrass. Improving winter hardiness is an important breeding goal in ryegrass breeding programs. An understanding of the genetic basis for the component traits of winter hardiness would allow more efficient selection. A three-generation interspecific population of an annual × perennial ryegrass consisting of 152 progenies was used to map quantitative trait loci (QTL) that control winter hardiness-related traits including fall growth (FG), freezing tolerance (FT), and winter survival (WS) over 2 years. A total of 39 QTL were identified for the three traits from both the female parental (MFA) and the male parental (MFB) maps, of which 13 were for FG, 6 for FT, and 20 for WS. The proportion of phenotypic variation explained by individual QTL ranged from 10.4 to 22.1%. Both FG and FT were positively correlated with WS. Common QTL were detected between FG, FT, and WS. The QTL associated with WS on linkage groups (LGs) 4 and 5, and the QTL for FT on LG 5 were consistently identified over years and maps. These consistent QTL might serve as potential tools for marker-assisted selection to improve ryegrass winter hardiness.

Annual (Lolium multiflorum Lam.) and perennial (Lolium perenne L.) ryegrasses are two important forage and turfgrass species. Improving the digestibility of forage by decreasing fiber content is a major goal in forage crop breeding programs. An annual · perennial ryegrass interspecific hybrid population was used to map quantitative trait loci (QTLs) for fiber components, neutral detergent fiber (NDF), acid detergent fiber (ADF), and acid detergent lignin (ADL), and crude protein (CP). Samples were harvested three times in August and September 2003 and August 2004, respectively. Simple interval mapping was used to detect QTLs from both the male and female parental maps previ- ously developed for the population. Fiber com- ponents were all correlated positively with each other and were negatively correlated with CP. The largest correlations were between NDF and ADF with r = 0.86, 0.72, and 0.82 for each of the three harvests. All four traits showed interme- diate broad-sense heritability values ranging from 0.35 to 0.72. A total of 63 QTLs were de- tected for the four traits measured over the three harvests from both the female and male maps. Coincident QTLs were detected on linkage groups (LGs) 2, 6, and 7 for NDF, LGs 1, 2, and 7 for ADF, LGs 6 and 7 for ADL, and LG 2 for CP, respectively. Coincident QTLs were also detected on LGs 2, 6, and 7 for NDF and ADF, providing evidence of the genetic basis of the observed high level of phenotypic correlation. The QTLs on LGs 2, 6, and possibly 7 for fiber components were co-located on the same LG as several lignin biosynthetic genes from perennial ryegrass.

The dehydration-responsive element binding proteins (DREB1)/C-repeat (CRT) binding factors (CBF) function as transcription factors and bind to the DRE/CRT cis-acting element (core motif: G/ACCGAC) commonly present in cold-regulated (COR) genes and subsequently upregulate the expression of such genes in Arabidopsis. We identified a DREB1A/CBF3-like gene, designated LpCBF3, from perennial ryegrass (Lolium perenne L.) by using RT-PCR and RACE (rapid amplification of cDNA end). The LpCBF3 gene contains all the conserved domains known to exist in other CBF genes. A comprehensive phylogenetic analysis using known and computationally identified CBF homologs in this study revealed that all monocot CBF genes are separately clustered from eudicot CBF genes and the LpCBF3 is the ortholog of rice OsDREB1A/CBF3 gene. Similar to other DREB1A/CBF3 homologs, expression of the LpCBF3 is induced by cold stress, but not by abscisic acid (ABA), drought, or salinity. Overexpression of the LpCBF3 cDNA in Arabidopsis induced expression of the Arabidopsis DREB1A/CBF3 target COR genes, COR15a and RD29A, without cold acclimation. Ion leakage in leaves of the overexpression transgenic plants was significantly reduced, an indication of enhanced freezing tolerance. Our data demonstrated that LpCBF3 not only resembles DREB/CBF genes of Arabidopsis, but is also capable of functioning as a transcriptional regulator in Arabidopsis, a species distant to the grass family.

Colonial bentgrass (Agrostis capillaris L.) is a cool-season grass, native to temperate Asia and Europe. It has good tolerance to low temperatures and partial shade and is well suited to golf course fairways and tees. Little information is available regarding levels and patterns of genetic variation among populations of colonial bentgrass, which would be useful for breeding programs. To study the genetic relationships among 27 colonial bentgrass accessions obtained from the US National Plant Germplasm System (NPGS), randomly amplified polymorphic DNA (RAPD) markers were scored and analyzed. Out of 80 primers screened, 16 were selected for further analysis, which yielded a total of 120 polymorphic bands used to differentiate the accessions. Dice's similarity coefficients for pair-wise comparisons ranged from 0.23 to 0.84 based on the RAPD data. Since there was no similarity coefficient value close to 1 between any two accessions, there was no apparent duplication among the sampled accessions. A dendrogram constructed on the basis of the Unweighted Pair Group Method with Arithmetic average (UPGMA) clustering algorithm clearly separated 26 of the accessions into three clusters with one accession distinct from the rest. The least similar pair of accessions was PI 204397 from Turkey and PI 628720 from Bulgaria, and the most similar pair was PI 509437 from Romania and PI 491264 from Finland. Clustering patterns based on principal components analysis (PCA) corresponded well with the dendrogram. A high cophenetic correlation (r = 0.82) was found between the RAPD data matrix and cophenetic matrix. The accession PI 628720, from Bulgaria, did not cluster with any other accessions.

Kentucky bluegrass (Poa pratensis L.) is an important turf and forage grass species with a facultative apomictic breeding behavior. In this study, mature seed and leaf tissue from 38 accessions of a USDA core collection of Kentucky bluegrass were analyzed with flow cytometry to characterize the reproductive mode and DNA content for each accession. Major reproductive pathways for each accession were determined based upon the presence and the position of the peaks observed and the known methods of reproduction for Kentucky bluegrass. While the majority of the accessions exhibited facultative apomictic reproductive behavior with a combination of reduced, zygotic and unreduced, parthenogenic embryo production, obligate sexual or obligate apomictic accessions were also found to be present in this core collection. In addition, reduced, parthenogenic and unreduced zygotic embryos were also detected in several accessions. Flow cytometric analysis of somatic tissue revealed a large range of DNA variation within this core collection. We also examined the sensitivity of flow cytometry in analyzing bulked samples containing a large number of plants with varied DNA content and determined that flow cytometry can effectively detect a plant having a different DNA content within a 15-plant bulk sample. Overall the combination of mature seed and somatic tissue analysis generated important information for the Kentucky bluegrass core collection and can be an effective and affordable tool to characterize even greater numbers of Kentucky bluegrass accessions.

Brassinosteroids (BRs) play important roles in plant growth, development and responses to a range of environmental cues. Although the mechanism of how BRs regulate growth and development is well-understood in Arabidopsis, the effect of BRs on stress tolerance, particularly drought tolerance remains unknown. We isolated a BRI1 (BRASSINOSTEROID INSENSITIVE 1) homologous gene, BdBRI1 from Brachypodium distachyon, a model for temperate grasses and cereals, created and characterized RNA interference (RNAi) knockdown mutants for BdBRI1 in Brachypodium. The loss-of-function BdBRI1-RNAi mutants exhibited reduced plant height, shortened internodes, narrow and short leaf, and reduced expression of BR signaling genes, BdBES1, BdBZR1, BdBLE2, and enhanced expression of BR biosynthesis genes BdD2, BdCPD and BdDWF4. More importantly, BdBRI1 RNAi mutants exhibited enhanced drought tolerance, accompanied by highly elevated expression of drought-responsive genes, BdP5CS, BdCOR47/BdRD17, together with BdERD1 and BdRD26, two putative targets of the transcription factors BES1 and BZR1 that are key components of the BR signaling pathway. Our results suggest that BR signaling and biosynthesis are largely conserved among Arabidopsis, rice and Brachypodium, and that BR signaling plays an important role in drought tolerance by directly regulating expression of key drought-responsive genes. The effect of BR biosynthesis or crosstalks between BR and other hormones or components of stress signaling pathways on drought tolerance is discussed.

Common bermudagrass, Cynodon dactylon, is a widely used warm-season turf and forage species in the temperate and tropical regions of the world. We have been able to transform the species using Agrobacterium-mediated approach. In seven experiments reported here, a total of 67 plates of calluses and suspensions were infected with Agrobacterium tumefaciens strains, and nine hygromycin B resistant calluses were obtained after selection. Among them two green independent transgenic plants were recovered. The plants growing in pots looked relatively compact at the beginning, but the ploidy level of the plants, as determined by nuclear DNA content, was not altered.