Cell-based screening methods for nuclear receptor ligands that use transgenic plant cells expressing a single human NR may have advantages over other eukaryotic systems which express multiple NRs. For example, signal-to-noise ratio might be improved because ligands would be less likely to bind to other NRs and/or less likely to cause confounding functional changes in plant cells. As a first step toward this aim we have expressed in plants truncated human estrogen receptor (ER) constructs linked to reporters, or selective markers such as luciferase, green fluorescent protein (GFP) and hygromycin. A variety of ligands for the ER (including estradiol and known phytoestrogens) have then been tested for their ability to over-express the linked marker gene(s) which could be measured (luciferase activity), visualized under fluorescent microscopy (GFP activity), or selected on antibiotic-containing media (Hygromycin B). Our results show a close association between the effects of ER ligands in the transgenic plant roots and their effects on native ERs in mammalian cells. With the stable expression of an ERalpha-GFP ligand detection system in the estradiol- mediated response in transgenic roots is inhibited by an ER partial agonist (tamoxifen) and an antagonist (fulvestrant) at concentrations relevant to their use in breast cancer. We conclude that it is possible to express human NRs in plants in a form that can report on exogenous or endogenous ER ligands and that these constructs have a pharmacology which is relevant to ligands for the native NRs in human cells.

The gene encodes isopentenyl pyrophosphate transferase, a key enzyme in cytokinin biosynthesis. We introduced under the control of the CaMV35S promoter into Asakura-sanshoo ( (L.) DC. f. Makino) via stable -mediated transformation. Three of 3-year-old transgenic Asakura-sanshoo lines Y5, Y16, and Y17 were used to evaluate the effects of expression on the morphological characteristics, leaf senescence, and essential oil composition. Introduced into Asakura-sanshoo stimulated an increase in cytokinin content and a decrease in auxin level. The increase in the cytokinin/auxin ratio affected the tree architecture in 3-year-old transgenic lines. The phenotypes of transgenic lines included reduced stem elongation, decreased leaf surface area, increased branching, and delayed leaf senescence. The expression of in Asakura-sanshoo also affected the leaf essential oil composition. The amount of oxygenated sesquiterpenoid compounds in Y5 and Y16 was 21.1 and 15.8 % higher, respectively, than that in wild type (WT). The amount of aromatic compounds in Y5 and Y16 was 2.9 and 24.6 % lower, respectively, than that in WT. These results show that expression in Asakura-sanshoo conferred desirable traits, including a dwarf growth habit, delayed senescence, and increased concentrations of some sesquiterpenoid compounds.

Full-size members of the ABCG (ATP-binding cassette, subfamily G) subfamily of ABC transporters have been found only in plants and fungi. The plant genes encoding full-size ABCGs identified so far appeared to be differentially regulated under various environmental constraints, plant growth regulators, and microbial elicitors, indicating a broad functional role of these proteins in plant responses to abiotic and biotic stress. Nevertheless, the structure and physiological function of full-size ABCGs in many plant species are still unknown. We have recently identified 16 genes encoding full-size ABCG proteins in cucumber and found that the transcripts of two of them, () and (), are most abundant in roots and are significantly affected by phytohormones and auxin herbicide. In this study, we analyzed the structure and phylogeny of all the full-size cucumber ABCG transporters and studied the organ expression profiles of the remaining 14 genes. In addition, we investigated the effect of different plant growth regulators and the diterpene sclareolide on expression in cucumber roots. Until now, the full-size plant ABCG transporters have been grouped into five different clusters. The new phylogenetic analysis of full-size ABCGs from model plants and cucumber clustered these proteins into six different subgroups. Interestingly, the expression profiles of cucumber genes assigned to the same clusters were not correlated, suggesting functional diversification or different regulatory mechanisms of the full-size cucumber ABCG proteins.

Cassava ( Crantz) is a tropical root crop and sensitive to low temperature. However, it is poorly to know how cassava can modify its metabolism and growth to adapt to cold stress. An investigation aimed at a better understanding of cold-tolerant mechanism of cassava plantlets was carried out with the approaches of physiology and proteomics in the present study. The principal component analysis of seven physiological characteristics showed that electrolyte leakage (EL), chlorophyll content, and malondialdehyde (MDA) may be the most important physiological indexes for determining cold-resistant abilities of cassava. The genome-wide proteomic analysis showed that 20 differential proteins had the same patterns in the apical expanded leaves of cassava SC8 and Col1046. They were mainly related to photosynthesis, carbon metabolism and energy metabolism, defense, protein synthesis, amino acid metabolism, signal transduction, structure, detoxifying and antioxidant, chaperones, and DNA-binding proteins, in which 40 % were related with photosynthesis. The remarkable variation in photosynthetic activity and expression level of peroxiredoxin is closely linked with expression levels of proteomic profiles. Moreover, analysis of differentially expressed proteins under cold stress is an important step toward further elucidation of mechanisms of cold stress resistance.

Next-generation sequencing technologies provide opportunities to ascertain the genetic basis of phenotypic differences, even in the closely related cultivars via detection of large amount of DNA polymorphisms. In this study, we performed whole-genome re-sequencing of two mei cultivars with contrasting tree architecture. 75.87 million 100 bp pair-end reads were generated, with 92 % coverage of the genome. Re-sequencing data of two former upright mei cultivars were applied for detecting DNA polymorphisms, since we were more interested in variations conferring weeping trait. Applying stringent parameters, 157,317 mutual single nucleotide polymorphisms (SNPs) and 15,064 mutual insertions-deletions (InDels) were detected and found unevenly distributed within and among the mei chromosomes, which lead to the discovery of 220 high-density, 463 low-density SNP regions together with 80 high-density InDel regions. Additionally, 322 large-effect SNPs and 433 large-effect InDels were detected, and 10.09 % of the SNPs were observed in coding regions. 5.25 % SNPs in coding regions resulted in non-synonymous changes. Ninety SNPs were chosen randomly for validation using high-resolution melt analysis. 93.3 % of the candidate SNPs contained the predicted SNPs. Pfam analysis was further conducted to better understand SNP effects on gene functions. DNA polymorphisms of two known QTL loci conferring weeping trait and their functional effect were also analyzed thoroughly. This study highlights promising functional markers for molecular breeding and a whole-genome genetic basis of weeping trait in mei.

Seabuckthorn ( L.), an upcoming superfood plant, has attracted researchers' attention worldwide for its medicinal, nutritional, and socio-economic value, along with its characteristic features to sustain extreme climatic conditions. We have studied microsatellite marker-based genetic and morphometric diversity in 93 collections of from different geographic sites representing two regions, namely Leh and Lahaul of the Indian Himalayas. Microsatellite markers were isolated using two different approaches, including screening of microsatellite-enriched genomic library, and in silico screening of in-house developed seabuckthorn EST database and whole transcriptome assembly. In Leh and Lahaul collections, 32 and 30 microsatellite markers were found polymorphic, respectively. All the markers developed for showed cross-species transferability to and Two to six alleles were recorded in the two sets of collections with an average of 3.71 and 3.53 alleles per locus in Leh and Lahaul collections, respectively. Mean polymorphic information content (PIC) values for microsatellite markers were 0.39 and 0.41 for Leh and Lahaul collections, respectively. The average expected heterozygosity was less than the observed heterozygosity. Wright's fixation index ( ) varied from (-)0.2045 to 1.0 and (-)0.1688 to 1.0 for Leh and Lahaul collections, respectively. Shannon's informative index () remained in the range of 0.6745 to 1.8621, and 0.6824 to 1.6308 for Leh and Lahaul collections, respectively. The UPGMA-based combined dendrogram showed clear demarcation between Leh and Lahaul collections, although a few ecotypes were regrouped with collections from the other region. No significant relationship was observed between the morphological distance matrix and molecular marker distance matrix. The findings of the present study may prove helpful in future breeding and conservation strategies aiming for seabuckthorn improvement.

Alternative oxidase (AOX) is a mitochondrial terminal oxidase which is responsible for an alternative route of electron transport in the respiratory chain. This nuclear-encoded enzyme is involved in a major path of survival under adverse conditions by transfer of electrons from ubiquinol instead of the main cytochrome pathway. AOX protects against unexpected inhibition of the cytochrome c oxidase pathway and plays an important role in stress tolerance. Two AOX subfamilies (AOX1 and AOX2) exist in higher plants and are usually encoded by small gene families. In this study, genome-wide searches and cloning were completed to identify and characterize AOX genes in cucumber ( L.). Our results revealed that cucumber possesses no AOX1 gene(s) and only a single AOX2 gene located on chromosome 4. Expression studies showed that AOX2 in wild-type cucumber is constitutively expressed at low levels and is upregulated by cold stress. AOX2 transcripts and protein were detected in leaves and flowers of wild-type plants, with higher levels in the three independently derived mosaic (MSC) mitochondrial mutants. Because cucumber possesses a single AOX gene and its expression increases under cold stress and in the MSC mutants, this plant is a unique and intriguing model to study AOX expression and regulation particularly in the context of mitochondria-to-nucleus retrograde signaling.

L. is an ancient genus of heterosporous lycopsids with a unique phylogenetic position. Repeated adaptations to environmental changes over time have contributed to occupying a variety of niches in . However, we know little about how they adapt to the environmental changes, and the sequence resources are very limited in public databases. is an amphibious plant in this genus, alternating frequently between terrestrial and aquatic environments. In this study, was applied to investigate the adaptations under terrestrial (TC) and submerged (ST) conditions using Illumina RNA-sequencing technology. Approximately 87 million high-quality reads were yielded and assembled into 31,619 unigenes with an average length of 1618 bp. Overall, 28,208 unigenes were annotated against the National Center of Biotechnology Information (NCBI), Non-redundant (Nr), Cluster of Orthologous Groups (COG), Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases. Moreover, we identified 1740 differentially expressed genes with 1146 up-regulated and 594 down-regulated genes under TC. GO annotation revealed that stress-relevant categories were remarkably enriched, and KEGG enrichment analysis showed that the phytohormone signalings and carbohydrate metabolism were significantly influenced. Furthermore, a total of 1646 transcription factors (TF) were identified and classified into 54 TF families; among them, 180 TFs were dynamic between terrestrial and submerged conditions. This study is the first report for to generate numerous sequences and establish general understandings about the adaptations in the changing environments. The dataset provides a foundation for novel gene discoveries, comparative genomics, functional genomics, and phylogenetics in .

The enzyme UDP-glucose dehydrogenase (UGD) competes with sucrose-phosphate synthase for the common photosynthesis product UDP-glucose. Sucrose-phosphate synthase is part of a pathway for the export of sucrose from source leaves to neighboring cells or the phloem. UGD is a central enzyme in a pathway for many nucleotide sugars used in local cell wall biosynthesis. Here, we identify a highly conserved phosphorylation site in UGD which is readily phosphorylated by MAP-kinase 3 in Arabidopsis. Phosphorylation occurs at a surface-exposed extra loop in all plant UGDs that is absent in UGDs from bacteria or animals. Phosphorylated sucrose-phosphate synthase is shifted to an inactive form which we did not measure for phosphorylated UGD. Plant UGDs have an extra loop which is phosphorylated by AtMPK3. Phosphorylation is not causing a reduction of UGD activity as found for the competitor enzymes and thus sets a preference for maintaining UDP-sugars at a constant level to prioritize cell wall biosynthesis.

In plants, the small protein related to ubiquitin (RUB) modifies cullin (CUL) proteins in ubiquitin E3 ligases to allow for efficient transfer of ubiquitin to substrate proteins for degradation by the 26S proteasome. At the molecular level, the conjugation of RUB to individual CUL proteins is transient in nature, which aids in the stability of the cullins and adaptor proteins. Many changes in cellular processes occur within the plant upon exposure to light, including well-documented changes in the stability of individual proteins. However, overall activity of E3 ligases between dark- and light-grown seedlings has not been assessed in plants. In order to understand more about the activity of the protein degradation pathway, overall levels of RUB-modified CULs were measured in seedlings growing in different light conditions. We found that light influenced the global levels of RUBylation on CULs, but not uniformly. Blue light had little effect on both Cul1 and Cul3 RUBylation levels. However, red light directed the increase in Cul3 RUBylation levels, but not Cul1. This red-light regulation of Cul3 was at least partially dependent on the activation of the phytochrome B signaling pathway. The results indicate that the RUBylation levels on individual CULs change in response to different light conditions, which enable plants to fine-tune their growth and development to the various light environments.

is an axenically cultivable fungus, which colonizes a broad range of plant species including the model plant . Root colonization by this endophyte leads to enhanced plant fitness and performance and promotes resistance against different biotic and abiotic stresses. The involvement of MPK6 in this mutualistic interaction had been previously shown with an mutant, which failed to respond to colonization. Here, we demonstrate that roots are significantly less colonized by compared to wild-type roots and the foliar application of plant hormones, ethylene, or jasmonic acid, restores the colonization rate at least to the wild-type level. Further, hormone-treated plants show typical -induced growth promotion effects. Moreover, expression levels of several genes related to plant defense and hormone signaling are significantly changed at different colonization phases. Our results demonstrate that the successful root colonization by depends on efficient suppression of plant immune responses. In , this process relies on intact hormone signaling in which MPK6 seems to play a pivotal role.

Peanut (), a major source of vegetable oil in many Asian countries, has become an integral part of human diet globally due to its high nutritional properties and option to consume in different forms. In order to meet the demand of vegetable oil, many peanut breeding programs of China have intensified their efforts in increasing oil content in newly bred varieties for reducing the import of edible oils in China. In this context, transcriptome sequencing data generated on 49 peanut cultivars were analyzed to identify candidate genes and develop molecular markers for seed oil content across multiple environments. Transcriptome analysis identified 5458 differentially expressed genes (DEGs) including 2243 positive DEGs and 3215 negative DEGs involved in oil synthesis process. Genome-wide association study identified 48 significant insertion/deletion (InDel) markers associated with seed oil content across five environments. A comparative genomics and transcriptomics analysis detected a total of 147 common gene clusters located in 17 chromosomes. Interestingly, an InDel cluster associated with seed oil content on A03 chromosome was detected in three different environments. Candidate genes identified on A03 form a haplotype, in which variable alleles were found to be different in oil content in an independent population. This locus is important for understanding the genetic control of peanut oil content and may be useful for marker-assisted selection in peanut breeding programs.

Common wild rice ( Griff.) is invaluable genetic resource for rice resistance breeding. Whole-genome re-sequencing was conducted to systematically analyze the variations in two new inbred lines (Huaye 3 and Huaye 4) developed from a common wild rice. A total of 4,841,127 SNPs, 1,170,479 InDels, 24,080 structural variations (SVs), and 298 copy number variations (CNVs) were identified in three materials. Approximately 16.24 and 5.64% of the total SNPs and InDels of Huaye 3 and Huaye 4 were located in genic regions, respectively. Together, 12,486 and 15,925 large-effect SNPs, and 12,417 and 14,513 large-effect InDels, which affect the integrity of the encoded protein, were identified in Huaye 3 and Huaye 4, respectively. The distribution map of 194 and 245 NBS-LRR encoding homologs was constructed across 12 rice chromosomes. Further, GO enrichment analysis of the homologs with identical genotype variations in Huaye 3 and Huaye 4 revealed 67, 82, and 58 homologs involved in cell death, response to stress, and both terms, respectively. Comparative analysis displayed that 550 out of 652 SNPs and 129 out of 147 InDels were present in a widely used blast-susceptible rice variety (LTH). Protein-protein interaction analysis revealed a strong interaction between NBS-LRR candidates and several known R genes. One homolog of disease resistance protein (RPM1) was involved in the plant-pathogen interaction pathway. Artificial inoculation of disease/insect displayed resistance phenotypes against rice blast and brown planthopper in two lines. The results will provide allele-specific markers for rice molecular breeding.

Intra-species genetic variability assessment is an effective tool in formulating genetic improvement and germplasm conservation strategies. Thunb. is a semidomesticated medicinal herb consumed widely in traditional diet in North-Eastern India. In the present study, an effort has been made to assess the genetic diversity of Thunb. from Brahmaputra valley of North-East India. A total of 545 genotypes from 18 populations of Thunb. from four different regions, i.e. North-East, North-West, South-East and South-West, with respect to river Brahmaputra were collected and population genetic diversity and structure were analysed using ISSR molecular markers. Population genetic structure analysis using unweighted pair group method with averages (UPGMA)-based hierarchical cluster analysis, principal coordinate analysis (PCoA) and model-based clustering in STRUCTURE program revealed that the population of Thunb. grouped according to regional distribution and forms four genetically distinct clusters. The analysis of molecular variance showed that differentiation among regions was significant with 60% genetic variation among region, 3% genetic variation among population within region and 37% genetic variation within population. We found wide variation in Nei's gene diversity (Hj) ranging from 0.07782 in Margherita population to 0.13634 in Barapani population. Furthermore, Nei's gene diversity within population (Nei's Hs) and total gene diversity (Ht) were found to be 0.1081 and 0.1769 respectively. The genetic differentiation among 18 population was high (Fst = 0.3894;  < 0.001) with relatively restricted gene flow (Nm = 0.6564). Based on the result of this study, we suggest ex situ conservation could be an appropriate measure to adequately capture the total genetic diversity of Thunb. populations of North-East India by selecting few individuals from different populations.

By cultivating tipburn-susceptible plants in modified Hoagland's medium containing of gradient exogenous calcium (Ca), we have shown that Ca deficiency is one of the main causes of tipburn in Chinese cabbage ( L. ssp. ). The effect of endogenous plant Ca concentrations on tipburn was also studied in a doubled haploid (DH) population consisting of 100 individuals, but no correlation was found. We then examined the expression of 12 Ca transporter genes that function in cytosolic Ca homeostasis in both tipburn-susceptible and tipburn-resistant plants under normal and tipburn-inducing conditions. Expression patterns for most of these genes differed between the two types of plants. Salicylic acid (SA) accumulated in response to conditions of calcium deficiency in our study, and both total SA and SA β-glucoside (SAG) in tipburn-susceptible plants was ∼3-fold higher than it was in resistant plants following Ca deficiency treatment. Also, the changes observed in SA levels correlated well with cell death patterns revealed by trypan blue staining. Therefore, we speculate that the cytoplasmic Ca fluctuation-induced downstream signaling events, as well as SA signaling or other biological events, are involved in the plant defense response to tipburn in Chinese cabbage.

Production of oilseed rape/canola () is increasingly threatened by dry conditions while the demand for vegetable oil is increasing. is a genetically diverse ancestor of , and is readily crossed with . Recently, we reported promising levels of drought tolerance in a wild type of which could be a source of drought tolerance for . We analysed global gene expression by messenger RNA sequencing in seedlings of the drought-tolerant and a drought-sensitive genotype of under simulated drought stress and control conditions. A subset of stress-response genes were validated by reverse transcription quantitative PCR. Gene ontology enrichment analysis and pathway enrichment analysis revealed major differences between the two genotypes in the mode and onset of stress responses in the first 12 h of treatment. Drought-tolerant plants reacted uniquely and rapidly by upregulating genes associated with jasmonic acid and salicylic acid metabolism, as well as genes known to cause endoplasmic reticulum stress and induction of programmed cell death. Conversely, active responses in drought-sensitive plants were delayed until 8 or 12 h after stress application. The results may help to identify biomarkers for selection of breeding materials with potentially improved drought tolerance.

The wheat and barley genes have been newly defined as key components of the light quality-dependent regulation of the freezing tolerance by the integration of phytochrome-mediated light and temperature signals. To further investigate the wavelength dependence of light-induced expression in cereals, we carried out a detailed study using monochromatic light treatments at an inductive and a non-inductive temperature. Transcript levels of gene in winter wheat Cheyenne, winter einkorn G3116 and winter barley Nure genotypes were monitored. We demonstrated that (1) is most effectively induced by blue light and (2) provide evidence that this induction does not arise from light-controlled gene expression. (3) We demonstrate that temperature shifts induce transcription independent of the light conditions and that (4) the effect of temperature and light treatments are additive. Based on these data, it can be assumed that temperature and light signals are relayed to the level of expression via separate signalling routes.

The primary aim of this study was to estimate genetic diversity among L. accessions using 22 previously published simple sequence repeat (SSR) markers. The plant material included 367 rye accessions comprising historical and contemporary cultivars, cultivated materials, landraces, and breeding strains from the Polish breeding company Danko. The studied accessions represented a wide geographical diversity. Several methods were employed to analyze genetic diversity among the L. accessions and to determine population structure: principal coordinate analysis (PCoA), neighbor-joining (NJ), and Bayesian clustering. We also defined a core collection of 25 rye accessions representing over 93 % of SSR alleles. The results of these analyses showed that accessions from the rye gene bank are clearly divergent in comparison with materials received directly from European breeding companies. Our findings suggest also that the genetic pool of current rye cultivars is becoming narrower during breeding processes. The selected panel of SSR markers performed well in detection of genetic diversity patterns and can be recommended for future germplasm characterization studies in rye.

Rye is a crop with relatively high resistance to biotic and abiotic stresses. However, the resistance to brown rust ( f. sp. ) and pre-harvest sprouting are still not satisfactory. High α-amylase activity is also among the main disadvantages of this species. Therefore, effective tools, e.g. molecular markers, allowing precise and environmentally independent selection of favourable alleles are desirable. In the present study, two kinds of association mapping-genome-wide association mapping (GWAM) based on sequences of DArTSeq markers and candidate gene association mapping (CGAM) based on sequences of genes-were chosen for development of molecular markers fulfilling these criteria. The analysed population consisted of 149 diverse inbred lines (DILs). Altogether, 67 and 11 single nucleotide polymorphisms (SNPs) identified in, respectively, GWAM and CGAM, were significantly associated with the investigated traits: 2 SNPs with resistance to brown rust, 71 SNPs with resistance to pre-harvest sprouting and 5 SNPs with α-amylase activity in the grain. Fifteen SNPs were stable across all environments. The highest number (13) of environmentally stable SNPs was associated with pre-harvest sprouting resistance. The test employing the Kompetitive Allele Specific PCR method proved the versatility of four markers identified in both GWAM and CGAM.

Peanut ( L.) is an important legume cultivated mostly in drought-prone areas where its productivity can be limited by water scarcity. The development of more drought-tolerant varieties is, therefore, a priority for peanut breeding programs worldwide. In contrast to cultivated peanut, wild relatives have a broader genetic diversity and constitute a rich source of resistance/tolerance alleles to biotic and abiotic stresses. The present study takes advantage of this diversity to identify drought-responsive genes by analyzing the expression profile of two wild species, and (AA and BB genomes, respectively), in response to progressive water deficit in soil. Data analysis from leaves and roots of (454 sequencing) and (suppression subtractive hybridization (SSH)) stressed and control complementary DNA (cDNA) libraries revealed several differentially expressed genes in silico, and 44 of them were selected for further validation by quantitative RT-PCR (qRT-PCR). This allowed the identification of drought-responsive candidate genes, such as Expansin, Nitrilase, , and transcription factors, displaying significant levels of differential expression during stress imposition in both species. This is the first report on identification of differentially expressed genes under drought stress and recovery in wild species. The generated transcriptome data, besides being a valuable resource for gene discovery, will allow the characterization of new alleles and development of molecular markers associated with drought responses in peanut. These together constitute important tools for the peanut breeding program and also contribute to a better comprehension of gene modulation in response to water deficit and rehydration.

The use of molecular markers allows for precise estimates of genetic diversity, which is an important parameter that enables breeders to select parental lines and designing breeding systems. We assessed the level of genetic diversity and population structure in a panel of 151 tropical maize inbred lines using 10,940 SNP (single nucleotide polymorphism) markers generated through the DArTseq genotyping platform. The average gene diversity was 0.39 with expected heterozygosity ranging from 0.00 to 0.84, and a mean of 0.02. Analysis of molecular variance showed that 97% of allelic diversity was attributed to individual inbred lines within the populations while only 3% was distributed among the populations. Both neighbor-joining clustering and STRUCTURE analysis classified the inbred lines into four major groups. The crosses that involve inbred lines from most divergent subgroups are expected to generate maximum heterosis and produce wide variation. The results will be beneficial for breeders to better understand and exploit the genetic diversity available in the set of maize inbred lines we studied.