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Paraquat is one of the most widely used nonselective herbicides in agriculture. Due to its wide use, paraquat resistant weeds have emerged and is becoming a potential threat to agriculture. The molecular mechanisms of paraquat resistance in weeds remain largely unknown. Physiological studies indicated that the impaired translocation of paraquat and enhanced antioxidation could improve paraquat resistance in plants. However, the detoxification of paraquat via active metabolism by plants has not been reported to date. Here we report that an activated expression of At1g01600 encoding the P450 protein CYP86A4 confers paraquat resistance as revealed by the gain-of-function mutant paraquat tolerance 11D (pqt11D), in which a T-DNA with four 35S enhancers was inserted at 1646 bp upstream the ATG of At1g01600. The paraquat resistance can be recapitulated in Arabidopsis wild type by overexpressing AtPQT11 (At1g01600), while its knockout mutant is hypersensitive to paraquat. Moreover, AtPQT11 also confers paraquat resistance in E. coli when overexpressed. We further demonstrate that AtPQT11 has P450 enzyme activity that converts paraquat to N-demethyl paraquat nontoxic to Arabidopsis, therefore detoxifying paraquat in plants. Taken together, our results unequivocally demonstrate that AtPQT11/ CYP86A4 detoxifies paraquat via active metabolism, thus revealing a novel molecular mechanism of paraquat resistance in plants and providing a means potentially enabling crops to resist paraquat.

Plants are talented biochemists that produce a broad diversity of small molecules. These so-called specialized metabolites (SMs) play critical roles in the adaptive evolution of plants to defend against biotic and abiotic stresses, attract pollinators, and modulate soil microbiota for their own benefits. Many plant SMs have been used as nutrition and flavor compounds in our daily food, as well as drugs for treatment of human diseases. Current multi-omics tools have significantly accelerated the process of biosynthetic pathway elucidation in plants through correlation analyses, genetic mapping, and de novo biosynthetic gene cluster predictions. Understanding the biosynthesis of plant SMs has enabled reconstitution of naturally occurring specialized metabolic pathways in microbial hosts, providing a sustainable supply of these high-value molecules. In this review, we illustrate the general functions of several typical plant SMs in natural ecosystems and for human societies. We then provide an overview of current methods elucidating the biosynthetic pathways of plant SMs, and synthetic biology strategies that optimize the efficiency of heterologous biosynthetic pathways in microbial hosts. Moving forward, dissection of the functions and application of plant SMs by using current multidiscipline approaches would be greatly benefit to the scientific community and human societies.

Plants supply both food and medicinal compounds, which are ascribed to diverse metabolites produced by plants. However, studies on domestication-driven changes in the metabolome and genetic basis of bioactive molecules in perennial fruit trees are generally lacking. Here, we conducted multidimensional analyses revealing a singular domestication event involving the genomic and metabolomic selection of jujube trees (Ziziphus jujuba Mill.). The genomic selection for domesticated genes was highly enriched in metabolic pathways, including carbohydrates and specialized metabolism. Domesticated metabolome profiling indicated that 187 metabolites exhibited significant divergence as a result of directional selection. Malic acid was directly selected during domestication, and the simultaneous selection of specialized metabolites, including triterpenes, consequently lead to edible properties. Cyclopeptide alkaloids (CPAs) were specifically targeted for the divergence between dry and fresh cultivars. We identified 1080 significantly associated loci for 986 metabolites. Among them, 15 triterpenes were directly selected at six major loci, allowing the identification of a homologous cluster containing seven 2,3-oxidosqualene cyclases (OSCs). An OSC gene was found to contribute to the reduction in the content of triterpenes during domestication. The complete pathway for synthesizing ursolic acid was dissected by integration of the metabolome and transcriptome. Additionally, an N-methyltransferase involved in the biosynthesis of CPA and responsible for inter-cultivar content variation was identified. The present study promotes our understanding of the selection process of the global metabolome subsequent to fruit tree domestication and facilitates the genetic manipulation of specialized metabolites to enhance their edible traits.

Tanshinones are the bioactive nor-diterpenoid constituents of the Chinese medicinal herb Danshen (Salvia miltiorrhiza). These groups of chemicals have the characteristic furan D-ring, which differentiates them from the phenolic abietane-type diterpenoids frequently found in the Lamiaceae family. However, how the 14,16-epoxy is formed has not been elucidated. Here, we report an improved genome assembly of Danshen using a highly homozygous genotype. We identify a cytochrome P450 (CYP71D) tandem gene array through gene expansion analysis. We show that CYP71D373 and CYP71D375 catalyze hydroxylation at carbon-16 (C16) and 14,16-ether (hetero)cyclization to form the D-ring, whereas CYP71D411 catalyzes upstream hydroxylation at C20. In addition, we discover a large biosynthetic gene cluster associated with tanshinone production. Collinearity analysis indicates a more specific origin of tanshinones in Salvia genus. It illustrates the evolutionary origin of abietane-type diterpenoids and those with a furan D-ring in Lamiaceae., Salvia miltiorrhiza is a medicinal plant that can produce the bioactive tanshinones. Here, the authors report the improved genome assembly and reveal the possible roles of three CYP71Ds in catalyzing the reactions leading to the formation of the characteristic furan D-ring of transhinones.

ZnS ceramic has been extensively used in various optical applications, e.g., full-wave band infrared window. The attractiveness of this material is explained by its wider transmission radiation range and higher infrared transmittance. An infrared window has often to be bonded to the protective metal frame due to high risk of involving demanding operational conditions, e.g., a vibration. So, an innovative joining technique must be necessarily uncovered. The objective of this work was designed to use a low-temperature glass powder to directly solder/braze ZnS ceramics during the temperature range between 400 and 450 °C. Our hypothesis was that the mechanical strength between ZnS glass and PbO–B2O3–ZnO powder would be significantly enhanced by doping the solder/braze by PbTiO3 particles. The formed interfacial reaction layer consists of PbS and the uniformly dispersed PbTiO3 particles. A heuristic explanation of the bonding mechanism is offered, and it is found that the use of sintering remelting can reduce defects within the glass joints. Owning to the second-phase diffusion strengthening of PbTiO3 particles and an interfacial chemical bonding in the joint, the mechanical properties are enhanced.

Diosgenin saponins isolated from

Crop breeding heavily relies on natural genetic variation. However, additional new variations are desired to meet the increasing human demand. Inflorescence architecture determines grain number per spike, a major determinant of bread wheat (Triticum aestivum L.) yield. Here, using Brachypodium distachyon as a wheat proxy, we identified DUO-B1, encoding an APETALA2/ethylene response factor (AP2/ERF) transcription factor, regulating spike inflorescence architecture in bread wheat. Mutations of DUO-B1 lead to mild supernumerary spikelets, increased grain number per spike and, importantly, increased yield under field conditions without affecting other major agronomic traits. DUO-B1 suppresses cell division and promotes the expression of BH

Diosgenin saponins isolated from Dioscorea species such as D. zingiberensis exhibit a broad spectrum of pharmacological activities. Diosgenin, the aglycone of diosgenin saponins, is an important starting material for the production of steroidal drugs. However, how plants produce diosgenin saponins and the origin and evolution of the diosgenin saponin biosynthetic pathway remain a mystery. Here we report a high-quality, 629-Mb genome of D. zingiberensis anchored on 10 chromosomes with 30 322 protein-coding genes. We reveal that diosgenin is synthesized in leaves (‘source’), then converted into diosgenin saponins, and finally transported to rhizomes (‘sink’) for storage in plants. By evaluating the distribution and evolutionary patterns of diosgenin saponins in Dioscorea species, we find that diosgenin saponin-containing may be an ancestral trait in Dioscorea and is selectively retained. The results of comparative genomic analysis indicate that tandem duplication coupled with a whole-genome duplication event provided key evolutionary resources for the diosgenin saponin biosynthetic pathway in the D. zingiberensis genome. Furthermore, comparative transcriptome and metabolite analysis among 13 Dioscorea species suggests that specific gene expression patterns of pathway genes promote the differential evolution of the diosgenin saponin biosynthetic pathway in Dioscorea species. Our study provides important insights and valuable resources for further understanding the biosynthesis, evolution, and utilization of plant specialized metabolites such as diosgenin saponins.

Triterpenoids are one of the largest groups of secondary metabolites and exhibit diverse structures, which are derived from C30 skeletons that are biosynthesized via the isoprenoid pathway by cyclization of 2,3-oxidosqualene. Triterpenoids have a wide range of biological activities, and are used in functional foods, drugs, and as industrial materials. Due to the low content levels in their native plants and limited feasibility and efficiency of chemical synthesis, heterologous biosynthesis of triterpenoids is the most promising strategy. Herein, we classified 121 triterpene alcohols/ketones according to their conformation and ring numbers, among which 51 skeletons have been experimentally characterized as the products of oxidosqualene cyclases (OSCs). Interestingly, 24 skeletons that have not been reported from nature source were generated by OSCs in heterologous expression. Comprehensive evolutionary analysis of the identified 152 OSCs from 75 species in 25 plant orders show that several pentacyclic triterpene synthases repeatedly originated in multiple plant lineages. Comparative analysis of OSC catalytic reaction revealed that stabilization of intermediate cations, steric hindrance, and conformation of active center amino acid residues are primary factors affecting triterpene formation. Optimization of OSC could be achieved by changing of side-chain orientations of key residues. Recently, methods, such as rationally design of pathways, regulation of metabolic flow, compartmentalization engineering, etc., were introduced in improving chassis for the biosynthesis of triterpenoids. We expect that extensive study of natural variation of large number of OSCs and catalytical mechanism will provide basis for production of high level of triterpenoids by application of synthetic biology strategies.

Odd-numbered primary alcohols are components of plant cuticular wax, but their biosynthesis remains unknown. We isolated a rice wax crystal-sparse leaf 5 (WSL5) gene using a map-based cloning strategy. The function of WSL5 was illustrated by overexpression and knockout in rice, heterologous expression in Arabidopsis and transient expression in tobacco leaves. WSL5 is predicted to encode a cytochrome P450 family member CYP96B5. The wsl5 mutant lacked crystalloid platelets on the surface of cuticle membrane, and its cuticle membrane was thicker than that of the wild-type. The wsl5 mutant is more tolerant to drought stress. The load of C23 -C33 alkanes increased, whereas the C29 primary alcohol reduced significantly in wsl5 mutant and WSL5 knockout transgenic plants. Overexpression of WSL5 increased the C29 primary alcohol and decreased alkanes in rice leaves. Heterologous expression of WSL5 increased the C29 primary alcohol and decreased alkanes, secondary alcohol, and ketone in Arabidopsis stem wax. Transient expression of WSL5 in tobacco leaves also increased the production C29 primary alcohol. WSL5 catalyzes the terminal hydroxylation of alkanes, yielding odd-numbered primary alcohols, and is involved in the formation of epidermal wax crystals on rice leaf, affecting drought sensitivity.

ZnS has been widely used as a material for windows in optical systems. This is because it has a good transmittance in both mid-infrared and far-infrared bands, a stable chemical performance, a moderate linear thermal expansion coefficient and an adequate mechanical strength. ZnS windows need the so-called splice technology implementation to satisfy the integrity of large size applications or special surface structures. Traditional ZnS light windows are mechanically assembled with metal frames. However, the strength of the large-size ZnS formed by mechanical assembly is low. Consequently, an adequate joining technology is needed to overcome the low splice strength and improve the mechanical strength of the assembly. In this paper, we proposed a pre-metallization method by using an electroless plating of Ni on ZnS surface. Subsequently, a low temperature joining by using a Sn63Pb37 solder was completed. By a study of the process and the corresponding mechanism of the electroless nickel plating, a Ni–P layer with a controllable P content and thickness has been formed, and a smooth surface and a good combination with ZnS has been obtained. The microstructure of the Sn63Pb37 and Ni–P/ZnS was analyzed, and the phase composition and element distribution within the joint were determined. The influences of process parameters on the microstructures of the joints have been explored, including the brazing temperature, the time of thermal energy preservation and the thickness of the coating. Finally, the shear strengths of joints under different imposed parameters were optimized leading to the peak soldering temperature of 250 °C and the dwell at the peak of 1 min. The fractures were along Ni–P/ZnS ceramic interface, therefore, a successfully low temperature joining process on ZnS ceramics using Ni–P plating and Sn63Pb37 solder had been performed.

Electrically conductive adhesives (ECAs) are mainly used in IC packaging, LED packaging and other electronic industry to replace the traditional soldering and lead-free solders, therefore, there is a constant need to improve the electrical conductive and mechanical property of ECAs. In this study, the electrical resistivity and shear strength of the modified nano-Ag coated Cu particles in ECA was investigated and compromised, in terms of filler contents, curing temperature and curing time. The conductive mechanism of the modified nano-Ag coated Cu particles in ECA was discussed as the filler content increased. Finally, the filler content was optimized by 23.5 vol% and 170 °C 100 min curing condition was selected to obtain the compromised property (5.159 × 10−6 Ω cm 6.121 MPa), which is better than the previous one (4.99 × 10−6 Ω cm, 4.406 MPa) at 200 °C. The tested data, as well as the failure mode and fracture interfaces, were carried out in this study.

Toosendanin, bearing a furan ring, is a limonoid belonging to the group of tetranortriterpenoids. Toosendanin is a phytochemical found in the medicinal plant Melia toosendan Sieb. et Zucc. of the Meliaceae family. Toosendanin and its derivatives demonstrate high insecticidal activity and are important pesticides derived from plants. Despite intensive investigation of limonoids over several decades, the biosynthetic pathway of these triterpenoids is less understood. To identify the key enzymes involved in the toosendanin biosynthetic pathway, we analyzed the contents of toosendanin in various plant tissues and parts and found that the highest level of toosendanin was found in the developing fruit and gradually decreased as the fruit matured. More than 346 116 transcripts were assembled based on 394 million paired-end Illumina reads and 6 million PacBio reads from the pooled RNA samples of fruits, leaves and young barks. A total of 186 263 genes were predicted. Six 2,3-oxidosqualene cyclase (OSC) genes were identified by analyzing the association between gene expression and metabolite profiles. Functional analyses using the Nicotiana benthamiana transient expression assay showed that MtOSC1 catalyzed 2,3-oxidosqualene to produce a tetracyclic triterpene skeleton, tirucalla-7,24-dien-3β-ol, which is predicted as the precursor for toosendanin biosynthesis. We identified another OSC, MtOSC6, which is a lupeol synthase. Using synthetic biology methods, these identified enzymes could be used to model a biosynthetic pathway to produce large quantities of toosendanin.

Global food security depends on cereal crops with durable disease resistance. Most cereals are colonized by rust fungi, which are pathogens of major significance for global agriculture1. Cereal rusts display a high degree of host specificity and one rust species or forma specialis generally colonizes only one cereal host2. Exploiting the non-host status and transferring non-host resistance genes between cereal crop species has been proposed as a strategy for durable rust resistance breeding. The molecular determinants that define the host status to rusts, however, are largely unknown. Here, we show that orthologous genes at the Rphq2 locus for quantitative leaf rust resistance from cultivated barley3 and Rph22 from wild bulbous barley4 affect the host status to leaf rusts. Both genes encode lectin receptor-like kinases. We transformed Rphq2 and Rph22 into an experimental barley line that has been bred for susceptibility to non-adapted leaf rusts, which allowed us to quantify resistance responses against various leaf rust species. Rphq2 conferred a much stronger resistance to the leaf rust of wild bulbous barley than to the leaf rust adapted to cultivated barley, while for Rph22 the reverse was observed. We hypothesize that adapted leaf rust species mitigate perception by cognate host receptors by lowering ligand recognition. Our results provide an example of orthologous genes that connect the quantitative host with non-host resistance to cereal rusts. Such genes provide a basis to exploit non-host resistance in molecular breeding.

Flavonols and proanthocyanidins are the major components that determine the black colouration of cucumber fruit spines and their synthesis is regulated by CsMYB60., Spine colour is an important fruit quality trait that influences the commercial value of cucumber (Cucumis sativus). However, little is known about the metabolites and the regulatory mechanisms of their biosynthesis in black spine varieties. In this study, we determined that the pigments of black spines are flavonoids, including flavonols and proanthocyanidins (PAs). We identified CsMYB60 as the best candidate for the previously identified B (Black spine) locus. Expression levels of CsMYB60 and the key genes involved in flavonoid biosynthesis were higher in black-spine inbred lines than that in white-spine lines at different developmental stages. The insertion of a Mutator-like element (CsMULE) in the second intron of CsMYB60 decreased its expression in a white-spine line. Transient overexpression assays indicated that CsMYB60 is a key regulatory gene and Cs4CL is a key structural gene in the pigmentation of black spines. In addition, the DNA methylation level in the CsMYB60 promoter was much lower in the black-spine line compared with white-spine line. The CsMULE insert may decrease the expression level of CsMYB60, causing hindered synthesis of flavonols and PAs in cucumber fruit spines.

Brachypodium distachyon (Brachypodium) has been developed as a model system for the temperate grasses. Hence, establishing a large insertion mutant population and identifying T-DNA insertion sites are imperative for functional genomics studies. Currently, Brachypodium has an established T-DNA collection resource, but it is far less than needed. In addition, the conventional methods for the isolation of flanking sequence tags (FSTs) characterizing the T-DNA inserts, such as TAIL-PCR, adapter ligation-mediated PCR, and inverse PCR, are time-consuming, laborious, and expensive. In this study, Agrobacterium-mediated transformation system was optimized to generate T-DNA mutants. Approximately 7000 T-DNA insertion lines were obtained. Furthermore, a simple and highly efficient method was developed to isolate T-DNA flanking sequence tags (FSTs). The procedures simplified the multi-step reaction that is required for the conventional inverse PCR and several reactions were conducted within a microscale reaction system in one tube. It is flexible to isolate FSTs for either individual or large numbers of T-DNA lines. To rapidly process the large-scale sequence data, a serial of Perl scripts were developed using the Perl Programming Language ( http://www.perl.org ). Using these methods, a total of 794 flanking sequences were isolated and analyzed in detail. Although the methods were developed in the process of isolating and analyzing T-DNA flanking sequences of Brachypodium, it can be applied to other plants.

Gas chromatography-mass spectrometry (GC-MS) is a robust analytical platform for analysis of small molecules. Recently, it is widely used for large-scale metabolomics studies, in which hundreds or even thousands of samples are analyzed simultaneously, producing a very large and complex GC-MS datasets. A number of software are currently available for processing GC-MS data, but it is too compute-intensive for them to efficiently and accurately align chromatographic peaks from thousands of samples. Here, we report a newly developed software, QPMASS, for analysis of large-scale GC-MS data. The parallel computing with an advanced dynamic programming approach is implemented in QPMASS to align peaks from multiple samples based on retention time and mass spectra, enabling fast processing large-scale datasets. Furthermore, the missing value filtering and backfilling are introduced into the program, greatly reducing false positive and false negative errors to be less than 5%. We demonstrated that it took only 8 h to align and quantify a GC-TOF-MS dataset from 300 rice leaves samples, and 17 h to process a GC-qMS dataset from 1000 rice seed samples by using a personal computer (3.70 GHz CPU, 16 GB of memory and > 100 GB hard disk). QPMASS is written in C++ programming language, and is able to run under Windows operation system with a user-friendly interface.

Plants produce a variety of metabolites that are essential for plant growth and human health. To fully understand the diversity of metabolites in certain plants, lots of methods have been developed for metabolites detection and data processing. In the data-processing procedure, how to effectively reduce false-positive peaks, analyze large-scale metabolic data, and annotate plant metabolites remains challenging. In this review, we introduce and discuss some prominent methods that could be exploited to solve these problems, including a five-step filtering method for reducing false-positive signals in LC-MS analysis, QPMASS for analyzing ultra-large GC-MS data, and MetDNA for annotating metabolites. The main applications of plant metabolomics in species discrimination, metabolic pathway dissection, population genetic studies, and some other aspects are also highlighted. To further promote the development of plant metabolomics, more effective and integrated methods/platforms for metabolite detection and comprehensive databases for metabolite identification are highly needed. With the improvement of these technologies and the development of genomics and transcriptomics, plant metabolomics will be widely used in many fields., Mass spectrometry-based metabolomics approaches are often used in analysis of complex metabolites in plants. This review discusses the methods on how to effectively reduce false-positive peaks, analyze large-scale datasets, and annotate plant metabolites. LC-MS- and GC-MS-based metabolome analyses have been used for species discrimination, metabolic pathway dissection, and population genetic studies, and plant metabolomics will be widely exploited in many fields of plant sciences.

Metabolomics is a rapidly emerging field of post-genomic research that aims to comprehensively analyze all metabolites in biological samples. Potential biomarkers that distinguish prostate cancer samples were successfully identified through metabolomics analysis (Sreekumar et al., 2009). Metabolome quantitative trait loci (mQTL) and genome-wide association studies coupled with metabolomics analysis (mGWAS) also became efficient tools to decipher the genetic basis of complex metabolic traits in large populations (Gong et al., 2013; Chen et al., 2014).

Global food security depends on cereal crops with durable disease resistance. Most cereals are colonized by rust fungi, which are pathogens of major significance for global agriculture

Transposon tagging is a powerful tool that has been widely applied in several species for insertional mutagenesis in plants. Several efforts have aimed to create transfer-DNA (T-DNA) insertional mutant populations in Brachypodium distachyon, a monocot plant used as a model system to study temperate cereals, but there has been a lack of research aimed at using transposon strategies. Here, we describe the application of a maize (Zea mays) Dissociation (Ds) transposon tagging system in B. distachyon. The 35S::AcTPase cassette and Ds element were constructed within the same T-DNA and transformed into B. distachyon plants. The Ds element was readily transposed to other chromosomes or to the same chromosome under the function of Activator (Ac) transposase. Through homologous chromosome synapsis, recombination, and segregation, the Ds element separated from the Ac element. We selected stable Ds-only plants using G418 and GFP assays and analyzed 241 T(0) lines, some of which were highly efficient at producing Ds-only progeny. Through thermal asymmetric interlaced PCR, we isolated 710 independent Ds flanking sequences from Ds-only plants. Furthermore, we identified a large collection of mutants with visible developmental phenotypes via this transposon tagging system. The system is relatively simple and rapid in comparison to traditional T-DNA insertion strategies, because once efficiency lines are obtained they can be reused to generate more lines from nontransposed plants without the use of time-consuming tissue culture steps.

Triterpene synthases, also known as 2,3-oxidosqualene cyclases (OSCs), synthesize diverse triterpene skeletons that form the basis of an array of functionally divergent steroids and triterpenoids. Tetracyclic and pentacyclic triterpene skeletons are synthesized via protosteryl and dammarenyl cations, respectively. The mechanism of conversion between two scaffolds is not well understood. Here, we report a promiscuous OSC from rice (Oryza sativa) (OsOS) that synthesizes a novel pentacyclic triterpene orysatinol as its main product. The OsOS gene is widely distributed in indica subspecies of cultivated rice and in wild rice accessions. Previously, we have characterized a different OSC, OsPS, a tetracyclic parkeol synthase found in japonica subspecies. Phylogenetic and protein structural analyses identified three key amino acid residues (#732, #365, #124) amongst 46 polymorphic sites that determine functional conversion between OsPS and OsOS, specifically, the chair–semi(chair)–chair and chair–boat–chair interconversions. The different orientation of a fourth amino acid residue Y257 was shown to be important for functional conversion. The discovery of orysatinol unlocks a new path to triterpene diversity in nature. Our findings also reveal mechanistic insights into the cyclization of oxidosqualene into tetra- and pentacyclic skeletons, and provide a new strategy to identify key residues determining OSC specificity.

Quantitative resistance (QR) refers to a resistance that is phenotypically incomplete and is based on the joined effect of several genes, each contributing quantitatively to the level of plant defense. Often, QR remains durably effective, which is the primary driver behind the interest in it. The various terms that are used to refer to QR, such as field resistance, adult plant resistance, and basal resistance, reflect the many properties attributed to it. In this article, we discuss aspects connected to those attributions, in particular the hypothesis that much of the QR to biotrophic filamentous pathogens is basal resistance, i.e., poor suppression of PAMP-triggered defense by effectors. We discuss what role effectors play in suppressing defense or improving access to nutrients. Based on the functions of the few plant proteins identified as involved in QR, vesicle trafficking and protein/metabolite transportation are likely to be common physiological processes relevant to QR.

In flowering plants, the pollen coat protects the released male germ cells from desiccation and damage during pollination. However, we know little about the mechanism by which the chemical composition of the pollen coat prevents dehydration of pollen grains. Here we report that deficiency of a grass conserved triterpene synthase, OsOSC12/OsPTS1, in rice leads to failure of pollen coat formation. The mutant plants are male sterile at low relative humidity (RH 80%). The lack of three major fatty acids in the pollen coat results in rapid dehydration of pollen grains. We show that applying mixtures of linolenic acid and palmitic acid or stearic acid are able to prevent over-dehydration of mutant pollen grains. We propose that humidity-sensitive genic male sterility (HGMS) could be a desirable trait for hybrid breeding in rice, wheat, maize, and other crops., In flowering plants, the pollen coat surrounds the male germ cells and protects against dehydration, damage and pathogen attack. Here, the authors show that a deficiency in terpenoid synthesis results in rice pollen over-dehydration and leads to a humidity-sensitive conditional male sterile phenotype.

The list of primary and secondary metabolic pathways in rice. (DOCX 13Â kb)

Seed vigor is an index of seed quality that is used to describe the rapid and uniform germination and the establishment of strong seedlings in any environmental conditions. Strong seed vigor in low-temperature germination conditions is particularly important in direct-sowing rice production systems. However, seed vigor has not been selected as an important breeding trait in traditional breeding programs due to its quantitative inherence. In this study, we identified and mapped eight quantitative trait loci (QTLs) for seed vigor by using a recombinant inbred population from a cross between rice (Oryza sativa L. ssp. indica) cultivars ZS97 and MH63. Conditional QTL analysis identified qSV-1, qSV-5b, qSV-6a, qSV-6b, and qSV-11 influenced seedling establishment and that qSV-5a, qSV-5c, and qSV-8 influenced only germination. Of these, qSV-1, qSV-5b, qSV-6a, qSV-6b, and qSV-8 were low-temperature-specific QTLs. Two major-effective QTLs, qSV-1, and qSV-5c were narrowed down to 1.13-Mbp and 400-kbp genomic regions, respectively. The results provide tightly linked DNA markers for the marker-assistant pyramiding of multiple positive alleles for increased seed vigor in both normal and low-temperature germination environments.

Summary Chemical mutagenesis is routinely used to create large numbers of rare mutations in plant and animal populations, which can be subsequently subjected to selection for beneficial traits and phenotypes that enable the characterization of gene functions. Several next-generation sequencing (NGS)-based target enrichment methods have been developed for the detection of mutations in target DNA regions. However, most of these methods aim to sequence a large number of target regions from a small number of individuals. Here, we demonstrate an effective and affordable strategy for the discovery of rare mutations in a large sodium azide-induced mutant rice population (F2). The integration of multiplex, semi-nested PCR combined with NGS library construction allowed for the amplification of multiple target DNA fragments for sequencing. The 8 9 8 9 8 tridimensional DNA sample pooling strategy enabled us to obtain DNA sequences of 512 individuals while only sequencing 24 samples. A stepwise filtering procedure was then elaborated to eliminate most of the false positives expected to arise through sequencing error, and the application of a simple Student’s t-test against position-prone error allowed for the discovery of 16 mutations from 36 enriched targeted DNA fragments of 1024 mutagenized rice plants, all without any false calls.

The cooking and eating qualities of rice are some of the problems facing by rice producers and consumers around the world. In this study we investigated the cooking and eating qualities of Basmati 385 and Hua Jing Xian 74. Both are rice cultivars with different phenotypic traits. The results show that Basmati 385 increased by an average of 102.14% after cooking while Hua Jing Xian 74 increased by an average of 69.91% after cooking. Basmati 385 had a pasting temperature of 78°C, while Hua Jing Xian 74 had a pasting temperature of 84°C. However, Hua Jing Xian 74 had a final viscosity of 4,686 cP while Basmati 385 had a final viscosity of 3,514 cP. The amylose contents of the two cultivars were also different; Hua Jing Xian 74 had an amylose content of 19.4%, while that of Basmati 385 was 18%. We also discovered that the starch granule morphology of the two cultivars were remarkably different. The lower pasting temperature of Basmati 385 positively correlated with good cooking qualities of rice. However, Hua Jing Xian 74 appears to have relatively better eating qualities due to its higher viscosity and higher amylose content. It would be pertinent to do more research about hybrid lines resulting from a cross of Hua Jing Xian 74 and Basmati 385, to see if they possess the good cooking qualities of Basmati 385 and the good eating qualities of Hua Jing Xian 74.

CRISPR/Cas9 is a powerful genome editing tool that has been extensively used in model plants and crops, such as Arabidopsis thaliana, rice, wheat, and soybean. Here, we report the use of CRISPR/Cas9 to precisely knock out the committed diterpene synthase gene (SmCPS1) involved in tanshinone biosynthesis in Salvia miltiorrhiza, a traditional Chinese medicinal herb with significant pharmacological activities, such as vasorelaxation, protection against ischemia-reperfusion injury, and antiarrhythmic effects. Three homozygous and eight chimeric mutants were obtained from 26 independent transgenic hairy root lines by Agrobacterium rhizogenes-mediated transformation. The metabolomic analysis based on LC-qTOF-MS and Q-TRAP-LC-MS/MS revealed that tanshinones, especially cryptotanshinone, tanshinone IIA and tanshinone I, are completely missing in homozygous mutants, without influencing other phenolic acid metabolites. By contrast, tanshinones are decreased but still detectable in chimeric mutants, which is similar to a previously-reported an RNAi study of SmCPS1. These results demonstrate that Agrobacterium rhizogenes- mediated transformation using CRISPR/Cas9 is a simple and efficient genome editing tool in S. miltiorrhiza, thus paving the way for large-scale genome editing in S. miltiorrhiza, which is important for pathway elucidation of secondary metabolites, quality improvement, and yield increases for this valuable traditional Chinese medicinal herb.

Wheat stripe rust, caused by Puccinia striiformis f. sp. tritici (PST), is one of the most destructive diseases and can cause severe yield losses in many regions of the world. Because of the large size and complexity of wheat genome, it is difficult to study the molecular mechanism of interaction between wheat and PST. Brachypodium distachyon has become a model system for temperate grasses’ functional genomics research. The phenotypic evaluation showed that the response of Brachypodium distachyon to PST was nonhost resistance (NHR), which allowed us to present this plant-pathogen system as a model to explore the immune response and the molecular mechanism underlying wheat and PST. Here we reported the generation of about 7,000 T-DNA insertion lines based on a highly efficient Agrobacterium-mediated transformation system. Hundreds of mutants either more susceptible or more resistant to PST than that of the wild type Bd21 were obtained. The three putative target genes, Bradi5g17540, BdMYB102 and Bradi5g11590, of three T-DNA insertion mutants could be involved in NHR of Brachypodium distachyon to wheat stripe rust. The systemic pathologic study of this T-DNA mutants would broaden our knowledge of NHR, and assist in breeding wheat cultivars with durable resistance.

Stripe (yellow) rust, caused by Puccinia striiformis Westend. f. sp. tritici Eriks (Pst), is one of the most important wheat (Triticum aestivum L.) diseases and causes significant yield losses. A recombinant inbred (RI) population derived from a cross between Yanzhan 1 and Xichang 76-9 cultivars was evaluated for resistance to wheat stripe rust strain CYR32 at both the seedling and adult plant stages. Four resistance quantitative trait loci (QTLs) were detected in this population, in which the major one, designated as Yrq1, was mapped on chromosome 2DS. The strategy of using the Brachypodium distachyon genome, wheat expressed sequence tags and a draft DNA sequences (scaffolds) of the D-genome (Aegilops tauschii Coss.) for the development of simple sequence repeat (SSR) markers was successfully used to identify 147 SSRs in hexaploid wheat. Of the 19 polymorphic SSRs in the RI population, 17 SSRs were mapped in the homeologous group 2 chromosomes near Yrq1 region and eight SSRs were genetically mapped in the 2.7 cM region of Yrq1, providing abundant DNA markers for fine-mapping of Yrq1 and marker-assisted selection in wheat breeding program. The effectiveness of Yrq1 was validated in an independent population, indicating that this resistance QTL can be successfully transferred into a susceptible cultivar for improvement of stripe rust resistance.

Integration of the genetic and metabolic fingerprinting can provide a new approach to differentiate similar Traditional Chinese Medical (TCM) materials. Two leguminous plants, Mojia Huangqi and Menggu Huangqi, are important medical herbs and share great similarities in morphology, chemical constituent, and genomic DNA sequence. The taxonomy of Mojia Huangqi and Menggu Huangqi has been debated for more than 50 years and discrimination of TCM materials directly affects the pharmacological and clinical effects. AFLP based genetic fingerprinting and GC-TOF/MS-based metabolic fingerprinting were used to successfully discriminate the two species. The results of AFLP supported the opinion that Menggu Huangqi was a variant of Mojia Huangqi. The metabolic fingerprinting showed growth locations have greater impacts on the metabolite composition and quantity than the genotypes (cultivated versus wild) in Menggu Huangqi. The difference of some soluble sugars, fatty acids, proline, and polyamine reflected plant adaptation to different growth environments. Using multivariate and univariate statistical analysis, three AFLP markers and eight metabolites were identified as candidate DNA and metabolic markers to distinguish the two herb materials. The correlation network between AFLP markers and metabolites revealed a complex correlation network, which indicated the special metabolic pathways and the regulation networks of Huangqi.