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Geraniol derived from essential oils of various plant species is widely used in the cosmetic and perfume industries. It is also an essential trait of the pleasant smell of rose flowers. In contrast to other monoterpenes which are produced in plastids via the methyl erythritol phosphate pathway, geraniol biosynthesis in roses relies on cytosolic NUDX1 hydrolase which dephosphorylates geranyl diphosphate (GPP). However, the metabolic origin of cytosolic GPP remains unknown. By feeding Rosa chinensis “Old Blush” flowers with pathway-specific precursors and inhibitors, combined with metabolic profiling and functional characterization of enzymes in vitro and in planta, we show that geraniol is synthesized through the cytosolic mevalonate (MVA) pathway by a bifunctional geranyl/farnesyl diphosphate synthase, RcG/FPPS1, producing both GPP and farnesyl diphosphate (FPP). The downregulation and overexpression of RcG/FPPS1 in rose petals affected not only geraniol and germacrene D emissions but also dihydro-β-ionol, the latter due to metabolic cross talk of RcG/FPPS1-dependent isoprenoid intermediates trafficking from the cytosol to plastids. Phylogenetic analysis together with functional characterization of G/FPPS orthologs revealed that the G/FPPS activity is conserved among Rosaceae species. Site-directed mutagenesis and molecular dynamic simulations enabled to identify two conserved amino acids that evolved from ancestral FPPSs and contribute to GPP/FPP product specificity. Overall, this study elucidates the origin of the cytosolic GPP for NUDX1-dependent geraniol production, provides insights into the emergence of the RcG/FPPS1 GPPS activity from the ancestral FPPSs, and shows that RcG/FPPS1 plays a key role in the biosynthesis of volatile terpenoid compounds in rose flowers.

For volatile organic compounds (VOCs) to be released from the plant cell into the atmosphere, they have to cross the plasma membrane, the cell wall, and the cuticle. However, how these hydrophobic compounds cross the hydrophilic cell wall is largely unknown. Using biochemical and reverse-genetic approaches combined with mathematical simulation, we show that cell-wall localized non-specific lipid transfer proteins (nsLTPs) facilitate VOC emission. Out of three highly expressed nsLTPs in petunia petals, which emit high levels of phenylpropanoid/benzenoid compounds, only PhnsLTP3 contributes to the VOC export across the cell wall to the cuticle. A decrease in PhnsLTP3 expression reduces volatile emission and leads to VOC redistribution with less VOCs reaching the cuticle without affecting their total pools. This intracellular build-up of VOCs lowers their biosynthesis by feedback downregulation of phenylalanine precursor supply to prevent self-intoxication. Overall, these results demonstrate that nsLTPs are intrinsic members of the VOC emission network, which facilitate VOC diffusion across the cell wall.

Biosynthesis of secondary metabolites relies on primary metabolic pathways to provide precursors, energy, and cofactors, thus requiring coordinated regulation of primary and secondary metabolic networks. However, to date it remains largely unknown how this coordination is achieved. Using Petunia hybrida flowers, which emit high levels of phenylpropanoid/benzenoid volatile organic compounds (VOCs), we uncovered genome-wide dynamic deposition of histone H3 lysine 9 acetylation (H3K9ac) during anthesis as an underlying mechanism to coordinate primary and secondary metabolic networks. The observed epigenome reprogramming is accompanied by transcriptional activation, at gene loci involved in primary metabolic pathways that provide precursor phenylalanine, as well as secondary metabolic pathways to produce volatile compounds. We also observed transcriptional repression among genes involved in alternative phenylpropanoid branches that compete for metabolic precursors. We show that GNAT family histone acetyltransferase(s) (HATs) are required for the expression of genes involved in VOC biosynthesis and emission, by using chemical inhibitors of HATs, and by knocking down a specific HAT, ELP3, through transient RNAi. Together, our study supports that chromatin level regulatory mechanisms may play an essential role in activating primary and secondary metabolic pathways to regulate VOC synthesis in petunia flowers.HIGHLIGHTOur study shows that posttranslational modification of histones is essential for regulating the biosynthesis and emission of floral scent compounds, thus providing insights into chromatin level regulation of secondary metabolism.

In plants, phenylalanine biosynthesis occurs via two compartmentally separated pathways. Overexpression of petunia chorismate mutase 2 (PhCM2), which catalyzes the committed step of the cytosolic pathway, increased flux in cytosolic phenylalanine biosynthesis, but paradoxically decreased the overall levels of phenylalanine and phenylalanine-derived volatiles. Concomitantly, the levels of auxins, including indole-3-acetic acid and its precursor indole-3-pyruvic acid, were elevated. Biochemical and genetic analyses revealed the existence of metabolic crosstalk between the cytosolic phenylalanine biosynthesis and tryptophan-dependent auxin biosynthesis mediated by an aminotransferase that uses a cytosolic phenylalanine biosynthetic pathway intermediate, phenylpyruvate, as an amino acceptor for auxin formation. In plants, the cytosolic phenylalanine biosynthetic intermediate phenylpyruvate can serve as an amino acceptor in tryptophan-dependent auxin biosynthesis, thus facilitating crosstalk between these two distinct primary metabolic pathways.

Chloroplasts are semi-autonomous organelles governed by the precise coordination between the genomes of their own and the nucleus for functioning correctly in response to developmental and environmental cues. Under stressed conditions, various plastid-to-nucleus retrograde signals are generated to regulate the expression of a large number of nuclear genes for acclimation. Among these retrograde signaling pathways, the chloroplast protein GENOMES UNCOUPLED 1 (GUN1) is the first component identified. However, in addition to integrating aberrant physiological signals when chloroplasts are challenged by stresses such as photooxidative damage or the inhibition of plastid gene expression, GUN1 was also found to regulate other developmental processes such as flowering. Several partner proteins have been found to interact with GUN1 and facilitate its different regulatory functions. In this study, we report 15 possible interacting proteins identified through yeast two-hybrid (Y2H) screening, among which 11 showed positive interactions by pair-wise Y2H assay. Through the bimolecular fluorescence complementation assay in Arabidopsis protoplasts, two candidate proteins with chloroplast localization, DJC31 and HCF145, were confirmed to interact with GUN1 in planta. Genes for these GUN1-interacting proteins showed different fluctuations in the WT and gun1 mutant under norflurazon and lincomycin treatments. Our results provide novel clues for a better understanding of molecular mechanisms underlying GUN1-mediated regulations.

Out of the three aromatic amino acids, the highest flux in plants is directed towards phenylalanine, which is utilized to synthesize proteins and thousands of phenolic metabolites contributing to plant fitness. Phenylalanine is produced predominantly in plastids via the shikimate pathway and subsequent arogenate pathway, both of which are subject to complex transcriptional and post-transcriptional regulation. Previously, it was shown that allosteric feedback inhibition of arogenate dehydratase (ADT), which catalyzes the final step of the arogenate pathway, restricts flux through phenylalanine biosynthesis. Here, we show that in petunia (Petunia hybrida) flowers, which typically produce high phenylalanine levels, ADT regulation is relaxed, but not eliminated. Moderate expression of a feedback-insensitive ADT increased flux towards phenylalanine, while high overexpression paradoxically reduced phenylalanine formation. This reduction could be partially, but not fully, recovered by bypassing other known metabolic flux control points in the aromatic amino acid network. Using comparative transcriptomics, reverse genetics, and metabolic flux analysis, we discovered that transcriptional regulation of the d-ribulose-5-phosphate 3-epimerase gene in the pentose phosphate pathway controls flux into the shikimate pathway. Taken together, our findings reveal that regulation within and upstream of the shikimate pathway shares control over phenylalanine biosynthesis in the plant cell.

Galling insects gain food and shelter by inducing specialized anatomical structures in their plant hosts. Such galls often accumulate plant defensive metabolites protecting the inhabiting insects from predation. We previously found that, despite a marked natural chemopolymorphism in natural populations of Pistacia palaestina, the monoterpene content in Baizongia pistaciae-induced galls is substantially higher than in leaves of their hosts. Here we show a general up-regulation of key structural genes in both the plastidial and cytosolic terpene biosynthetic pathways in galls as compared with non-colonized leaves. Novel prenyltransferases and terpene synthases were functionally expressed in Escherichia coli to reveal their biochemical function. Individual Pistacia trees exhibiting chemopolymorphism in terpene compositions displayed differential up-regulation of selected terpene synthase genes, and the metabolites generated by their gene products in vitro corresponded to the monoterpenes accumulated by each tree. Our results delineate molecular mechanisms responsible for the formation of enhanced monoterpene in galls and the observed intraspecific monoterpene chemodiversity displayed in P. palaestina. We demonstrate that gall-inhabiting aphids transcriptionally reprogram their host terpene pathways by up-regulating tree-specific genes, boosting the accumulation of plant defensive compounds for the protection of colonizing insects.

Metabolic engineering is embraced as a method to sustainably enhance production of valuable phytochemicals with beneficial properties. However, successful production of these compounds in plants is not always predictable even when the pathways are fully known, frequently due to the lack of comprehensive understanding of plant metabolism as a whole, and interconnections between different primary, secondary, and hormone metabolic networks. Here, we highlight critical hidden constraints, including substrate availability, silent metabolism, and metabolic crosstalk, that impair engineering strategies. We explore how these constraints have historically been manifested in engineering attempts and propose how modern advancements will enable future strategies to overcome these impediments.

Chloroplasts are semi-autonomous organelles, with more than 95% of their proteins encoded by the nuclear genome. The chloroplast-to-nucleus retrograde signals are critical for the nucleus to coordinate its gene expression for optimizing or repairing chloroplast functions in response to changing environments. In chloroplasts, the pentatricopeptide-repeat protein GENOMES UNCOUPLED 1 (GUN1) is a master switch that senses aberrant physiological states, such as the photooxidative stress induced by norflurazon (NF) treatment, and represses the expression of photosynthesis-associated nuclear genes (PhANGs). However, it is largely unknown how the retrograde signal is transmitted beyond GUN1. In this study, a protein GUN1-INTERACTING PROTEIN 1 (GIP1), encoded by At3g53630, was identified to interact with GUN1 by different approaches. We demonstrated that GIP1 has both cytosol and chloroplast localizations, and its abundance in chloroplasts is enhanced by NF treatment with the presence of GUN1. Our results suggest that GIP1 and GUN1 may function antagonistically in the retrograde signaling pathway.

Experimental research was conducted using concrete produced by adding the polycarboxylic-type water-reducing agent. The cement, sand, aggregates, fillers and water-reducing agent were used as the important raw materials for the concrete. Then the following properties of the concrete were analyzed: compressive strength, elastic modulus and frost resistance. The compressive strength was 84 MPa at 56 days and the modulus of elasticity was 45.3 GPa at 28 days. Therefore, the water-reducing agent played an important role for the concrete. All the properties met the technical requirements of the CRTS III track plate.

Compared with the traditional water reducer, polycarboxylicwater-reducing agent exhibits the advantages of high water-reducing rate, cement paste fluidity and low slump loss, etc. The structure of polycarboxylates water reducing agent molecular is comb type. Water reducing agent can be used in the molecular design because it has high water reducing rate, low dosage, good slump stability, and have great potential in increase strength. In recent years, it has attracted many researchers' attention. Water reducing agent can block or destroy cement granular flocculation structure, through the surface function, complexation, electrostatic repulsion force and stereo repulsive force. Research on water reducing agent based on the application of poly carboxylic acid can realize functional design of water reducing agent, so as to promote the development of high-performance concrete.

The conversion of etioplasts into chloroplasts in germinating cotyledons is a crucial transition for higher plants, enabling photoautotrophic growth upon illumination. Tight coordination of chlorophyll biosynthesis and photosynthetic complex assembly is critical for this process. ORANGE (OR), a DnaJ-like zinc finger domain-containing protein, was reported to trigger the biogenesis of carotenoid-accumulating plastids by promoting carotenoid biosynthesis and sequestration. Both nuclear and plastidic localizations of OR have been observed. Here, we show that Arabidopsis (Arabidopsis thaliana) OR physically interacts with the transcription factor TCP14 in the nucleus and represses its transactivation activity. Through this interaction, the nucleus-localized OR negatively regulates expression of EARLY LIGHT-INDUCIBLE PROTEINS (ELIPs), reduces chlorophyll biosynthesis, and delays development of thylakoid membranes in the plastids of germinating cotyledons. Nuclear abundance of OR decreased upon illumination. Together with an accumulation of TCP14 in the nucleus, this derepresses chloroplast biogenesis during de-etiolation. TCP14 is epistatic to OR and expression of ELIPs is directly regulated by the binding of TCP14 to Up1 elements in the ELIP promoter regions. Our results demonstrate that the interaction between OR and TCP14 in the nucleus leads to repression of chloroplast biogenesis in etiolated seedlings and provide new insights into the regulation of early chloroplast development.

In plants, phenylalanine biosynthesis occurs via two compartmentally separated pathways. Overexpression of petunia chorismate mutase 2 (PhCM2), which catalyzes the committed step of the cytosolic pathway, increased flux in cytosolic phenylalanine biosynthesis, but paradoxically decreased the overall levels of phenylalanine and phenylalanine-derived volatiles. Concomitantly, the levels of auxins, including indole-3-acetic acid and its precursor indole-3-pyruvic acid, were elevated. Biochemical and genetic analyses revealed the existence of metabolic crosstalk between the cytosolic phenylalanine biosynthesis and tryptophan-dependent auxin biosynthesis mediated by an aminotransferase that uses a cytosolic phenylalanine biosynthetic pathway intermediate, phenylpyruvate, as an amino acceptor for auxin formation.

Pepper ( Capsicum annuum) fruits are a rich source of carotenoids. Geranylgeranyl diphosphate (GGPP) is the precursor for carotenoid biosynthesis and is produced by GGPP synthase (GGPPS), which belongs to the prenyl transferase (PTS) family. In this study, we identified from the pepper genome a total of eight PTS homologues. Our subcellular localization, enzymatic activity, and expression level analyses proved that among these homologues Capana04g000412 is the only functional GGPPS (CaGGPPS1) for carotenoid biosynthesis in pepper fruits. We demonstrated that CaGGPPS1 interacts with a catalytically inactive small subunit homologue protein CaSSUII, and such an interaction promotes CaGGPPS1 enzymatic activity. We also revealed a protein-protein interaction between CaSSUII and a putative phytoene synthase and the repression of carotenoid accumulation by silencing CaSSUII in pepper fruits. Taken together, our results suggest an essential contribution of the CaGGPPS1/CaSSUII interaction to carotenoid biosynthesis in ripening pepper fruits.

Carotenoid metabolism in red algae is not well understood. Geranylgeranyl diphosphate (GGPP), synthesized by GGPP synthase (GGPS), is a precursor for the biosynthesis of many biologically important metabolites, including carotenoids and chlorophylls. GGPSs have been functionally characterized in many organisms, but not in species of the primitive red algal order Bangiales. Here, we cloned and characterized the gene encoding GGPS (PuGGPS) in Pyropia umbilicalis (Bangiales). PuGGPS encodes a protein of 345 amino acids with an N-terminal transit peptide. The catalytic activity of PuGGPS for the production of GGPP was verified by a color complementation assay in Escherichia coli and subsequent high-performance liquid chromatography analysis. Homology modeling of PuGGPS showed that its tertiary structure resembles that of other known GGPSs and that this structure allows for the precise docking of the enzymatic product of PuGGPS, GGPP. When leafy thalli of P. umbilicalis were treated with norflurazon, an inhibitor of the key carotenoid metabolism enzyme phytoene desaturase, the expression of PuGGPS increased by twofold compared with that of the control in the first 2 h, suggesting a prompt response to metabolic perturbation. Prolonged norflurazon treatment failed to increase PuGGPS expression. Sequence analysis showed that PuGGPS shares seven conserved motifs with other previously identified GGPSs from different organisms, including two aspartate-rich GGPS signature motifs. Phylogenetic analysis also indicated that PuGGPS is a member of the type II GGPSs found in eubacteria and plants.

Carotene hydroxylases catalyze the hydroxylation of α- and β-carotene hydrocarbons into xanthophylls. In red algae, β-carotene is a ubiquitously distributed carotenoid, and hydroxylated carotenoids such as zeaxanthin and lutein are also found. However, no enzyme with carotene hydroxylase activity had been previously identified in red algae. Here, we report the isolation of a gene encoding a cytochrome P450-type carotene hydroxylase (PuCHY1) from Porphyra umbilicalis, a red alga with an ancient origin. Sequence comparisons found PuCHY1 belongs to the CYP97B subfamily, which has members from different photosynthetic organisms ranging from red algae to land plants. Functional complementation in Escherichia coli suggested that PuCHY1 catalyzed the conversion from β-carotene to zeaxanthin. When we overexpressed PuCHY1 in the Arabidopsis thaliana chy2 mutant, pigment analysis showed a significant accumulation of hydroxylated carotenoids, including neoxanthin, violaxanthin, and lutein in the leaves of transgenic plants. These results confirmed a β-hydroxylation activity of PuCHY1, and also suggested a possible ϵ-hydroxylation function. The pigment profile and gene expression analyses of the algal thallus under high-light stress suggested that P. umbilicalis is unlikely to operate a partial xanthophyll cycle for photoprotection.

The difference of gene expression between sclerotia-producing and non-sclerotia-producing single spore isolates from Morchella conica were preliminary analyzed by mRNA differential display reverse transcription-polymerase chain reaction (RT-PCR) technique and 67 differential gene fragments were obtained. Fifty-eight of their second PCR products were cloned and sequenced. Thirteen special differential gene fragments related to sclerotial formation were validated by semi-quantitative RT-PCR. Some gene fragments had certain homologies with lipoprotein, cyclin-dependent kinase C-3, glycerophosphoryl diester phosphodiesterase, Rho GDP-dissociation inhibitor, gamma-aminobutyrate permease, OmpA family protein, Transcript antisense to ribosomal RNA protein, sodium–calcium exchange protein and keratin-associated proteins 5, 6. In addition, the putative protein of some DNA fragments had higher similarity with hypothetical protein-coding gene in NCBI database, as well as some were only putative gene fragments. All these fragments were speculated to be the functional gene associated with sclerotial formation in morel.

The biosynthesis of chlorophylls and carotenoids and the assembly of thylakoid membranes are critical for the photoautotrophic growth of plants. Different factors are involved in these two processes. In recent years, members of the DnaJ-like zinc finger domain proteins have been found to take part in the biogenesis and/or the maintenance of plastids. One member of this family of proteins, PSA2, was recently found to localize to the thylakoid lumen and regulate the accumulation of photosystem I. In this study, we report that the silencing of PSA2 in Arabidopsis thaliana resulted in variegated leaves and retarded growth. Although both chlorophylls and total carotenoids decreased in the psa2 mutant, violaxanthin, and zeaxanthin accumulated in the mutant seedlings grown under growth condition. Lower levels of non-photochemical quenching and electron transport rate were also found in the psa2 mutant seedlings under growth condition compared with those of the wild-type plants, indicating an impaired capability to acclimate to normal light irradiance when PSA2 was silenced. Moreover, we also observed an abnormal assembly of grana thylakoids and poorly developed stroma thylakoids in psa2 chloroplasts. Taken together, our results demonstrate that PSA2 is a member of the DnaJ-like zinc finger domain protein family that affects light acclimation and chloroplast development.

Yunnan Province of China is one of the important centers for origin and evolution of cultivated rice worldwide. Wild rice is the ancestor of the cultivated rice. Many elite traits of wild rice have widened the genetic basis in cultivated rice. However, many populations of wild rice species have disappeared in the past few years. Therefore, the current status of wild rice resources should be updated and the genetic diversity of wild rice species should be examined for further germplasm preservation and utilization. Our investigations showed that the number of natural wild rice populations declined sharply in Yunnan Province during the past few years due to various reasons. Fortunately, one population of Oryza rufipogon, three of O. officinalis and ten of O. granulata have been newly found in different ecological sites, which were confirmed by inter-simple sequence repeat (ISSR) marker analysis in this study. ISSR analysis and investigation of some important traits of nutritional values indicated that the genetic diversity of the currently existing wild rice resources in Yunnan is still rich. The demonstration of genetic diversity of wild rice by a combined use of geographical distribution, morphological traits, nutrition contents and ISSR markers would be helpful for the conservation and exploration of these important wild rice resources.

The complete nucleotide sequence of the sugar beet (Beta vulgaris ssp. vulgaris) chloroplast genome (cpDNA) was determined in this study. The cpDNA was 149,637 bp in length, containing a pair of 24,439 bp inverted repeat regions (IR), which were separated by small and large single copy regions (SSC and LSC) of 17,701 and 83,057 bp, respectively. 53.4% of the sugar beet cpDNA consisted of gene coding regions (protein coding and RNA genes). The gene content and relative positions of 113 individual genes (79 protein encoding genes, 30 tRNA genes, 4 rRNA genes) were almost identical to those of tobacco cpDNA. The overall AT contents of the sugar beet cpDNA were 63.6% and in the LSC, SSC and IR regions were 65.9%, 70.8% and 57.8%, respectively. Fifteen genes contained one intron, while three genes had two introns.

This study was conducted to evaluate the genetic diversity within and among the plants of four ancient tea gardens and two tableland tea gardens form Yunnan Province, China by AFLP technique. The percentage of polymorphic loci (P) of the plants from six tea gardens was 92.31%. The genetic diversity within the six gardens demonstrated by Nei cents genetic diversity (He) was estimated to be 0.1366, while Shannon indices (Ho) were 0.2323. The percentage of polymorphic loci of the four ancient tea populations was 45.55% on average, with a range of 36.44% (Mengsong) to 59.11% (Mengla). But the percentages of polymorphic loci of the plants from two tableland gardens were 13.77% (Yunkang 10) and 24.2% (Menghai Daye), respectively. There was a great genetic difference between ancient tea gardens and tableland tea gardens. The genetic diversity among the plants of the ancient tea garden was higher than those of the sexual tableland tea garden and the clone tableland tea garden based on P valve. The four ancient tea gardens and two tableland gardens could be differentiated with AFLP markers. The results show that AFLP marker is an effective tool in the discrimination of tea germplasm, as well as sundried green tea.

The cDNA library of Yuanjiang Oryza rifupongon leaf was constructed by using SMART technology. The titers of the non-amplified library and the amplified library were 1.1 x 106 pfu/mL and 3.98 x 107 pfu/mL, respectively. The recombination rate was more than 91%. The DNA sequence length of the most cDNAs in the library was between 500-2 000 bp. Some cDNAs chosen by random were sequenced. After BLAST analysis of some cDNAs, their possible function were predicted. It is found that these cDNAs show 98% similarity to Oryza sativa japonica in the NCBI database. These provided a base for further study on the structure and function of these cDNAs and evolutionary process of Yuanjiang Oryza rifupongon.

A 4,672 bp DNA sequence including the whole coding region and partial non-coding region of rice blast resistance gene Pi-ta+ has been cloned from Jinghong erect type of common wild rice (Oryza rufipogon Griff) in Yunnan by polymerase chain reaction method. The coding region shares 99.86% and 98.78% identity with the corresponding regions of the reported cultivated rice Yashiro-mochi and Yuanjiang type of common wild rice respectively. There are 4 nucleotides difference in the coding region and 6 in intron of the cloned Pi-ta+ gene,compared with Pi-ta from Yashiro-mochi. Pi-ta+ gene in Jinghong erect type of common wild rice has been proved to be a rare existing Pi-ta+ allele, because there was a alanine rather than a serine at the position 918 within the predicted amino acid sequence of PITA. Pi-ta+ allele can cause disease resistance response to rice blast pathogens in plant cells. Differences in DNA sequence, deduced amino acid sequence and antibacterial spectrum may make the Pi-ta+ allele new resistant characteristics. Finding and cloning of Pi-ta+ allele from Jinghong erect type of common wild rice in Yunnan provides a basement for further utilization of the wild rice resources.

Sometimes historic information about workflow execution is needed to analyze business processes. Process mining aims at extracting information from event logs for capturing a business process in execution. In this paper a process mining algorithm is proposed based on Synchro-Net which is a synchronization-based model of workflow logic and workflow semantics. With this mining algorithm based on the model, problems such as invisible tasks and short-loops can be dealt with at ease. A process mining example is presented to illustrate the algorithm, and the evaluation is also given.

In addition to rice (Oryza sativa L.) cultivars, there are three wild rice species, namely O. rufipogon Griff, O. officinalis Wall and O. granulata Baill, in Yunnan Province, China. Each species has different subtypes and ecological distributions. Yunnan wild rice species are excellent genetic resources for developing new rice cultivars. The nutritional components of the husked seeds of wild rice have not been investigated thus far. Herein, we report on the contents of total protein, starch, amylose, 17 amino acids, and five macro and five trace mineral elements in husked seeds from three wild rice species and six O. sativa cultivars. The mean (±SD) protein content in the husked rice of O. rufipogon, O. officinalis, and O. granulata was (14.5 ± 0.6)%, (16.3 ± 1.1)%, and (15.3 ± 0.5)%, respectively. O. officinalis III originating from Gengma had the highest protein content (19.3%). In contrast, the average protein content of six O. sativa cultivars was only 9.15%. The total content of 17 amino acids of three wild rice species was 30%-50% higher than that of the six cultivars. Tyrosine, lysine, and valine content in the three wild rice species was 34%-209% higher than that of the cultivars. However, the difference in total starch content among different O. sativa varieties or types of wild rice species was very small. The average amylose content of O. rufipogon, O. officinalis, and O. granulata was 12.0%, 9.7%, and 11.3%, respectively, much lower than that of the indica and japonica varieties (14.37%-17.17%) but much higher than that of the glutinous rice cultivars (3.89%). The sulfur, phosphorus, magnesium, zinc, and ferrite content in the three wild rice species was 30%-158% higher than that of the six cultivars. The considerable difference in some nutritional components among wild rice species and O. sativa cultivars represents a wide biodiversity of Yunnan Oryza species. Based on the results of the present study, it is predicted that some good genetic traits, especially high protein and ideal amylose content, of Yunnan wild rice species may be useful in improving the nutritional value of rice. This is the first report regarding the amino acid, mineral element, protein and amylose content of husked seeds of some Yunnan wild rice species that have important genetic characteristics for rice quality and nutritional value. (Managing editor: Wei WANG)

In order to evaluate and preserve the endangered wild species of rice, Oryza granulata , we investigated the existing population distribution in Yunnan Province of Southwestern China. The genetic diversity among 14 populations (bulk samples) and within two populations (individual samples) was studied using random amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR) amplification markers. The field survey showed that 44% natural populations of O. granulata in Yunnan have become extinct during the last 30 years. Of the 37 remaining wild populations, 28 (76%) are on the verge of extinction and nine (24%) were unaffected. For the study among the 14 populations, the percentage of polymorphic bands (PPB) was 59% for RAPDs and 64% for ISSRs. However, within populations, the PPB generated by RAPDs and ISSRs was 26 and 26% in the first population (S1) and similarly within the second population (S2) (21 and 22%, respectively). The results showed that genetic variation is much higher among populations of O. granulata than within populations. Analyses of Nei’s gene diversity, unbiased genetic distance and Shannon’s index also agreed with these results. Therefore, we proposed an appropriate strategy for conserving the genetic resources of O. granulata in Yunnan; namely, rescuing and conserving the core populations for in situ conservation, and sampling and preserving more populations with fewer individuals from each population for ex situ conservation.

This study was conducted to evaluate the genetic diversity within and among the plants of four ancient tea gardens and two tableland tea gardens form Yunnan Province, China by AFLP technique. The percentage of polymorphic loci (P) of the plants from six tea gardens was 92.31%. The genetic diversity within the six gardens demonstrated by Nei cents genetic diversity (He) was estimated to be 0.1366, while Shannon indices (Ho) were 0.2323. The percentage of polymorphic loci of the four ancient tea populations was 45.55% on average, with a range of 36.44% (Mengsong) to 59.11% (Mengla). But the percentages of polymorphic loci of the plants from two tableland gardens were 13.77% (Yunkang 10) and 24.2% (Menghai Daye), respectively. There was a great genetic difference between ancient tea gardens and tableland tea gardens. The genetic diversity among the plants of the ancient tea garden was higher than those of the sexual tableland tea garden and the clone tableland tea garden based on P valve. The four ancient tea gardens and two tableland gardens could be differentiated with AFLP markers. The results show that AFLP marker is an effective tool in the discrimination of tea germplasm, as well as sundried green tea.

Compared to Pi-ta(-) alleles, Pi-ta(+) alleles can cause blast resistance response. In this work, Pi-ta gene in multiple rice materials, including local rice cultivars, different types of O. rufipogon and O. longistaminata was detected by molecular cloning and sequence analysis. Results indicated that Pi-ta(+) alleles were rare alleles, because in all the tested materials, only the 'Erect' type of O. rufipogon (ETOR) from Jinghong county in Yunnan province contains a Pi-ta(+) allele. Another rice blast resistance gene, Pib, confers resistance to the Japanese strain of M. grisea, was also confirmed to be functional in this type of O. rufipogon. The results of pathogen inoculation test show that ETOR is more strongly resistant to the tested blast pathogen races than other types of O. rufipogon. The resistance of ETOR may at least partially depend upon the functioning of Pi-ta and Pib gene. As O. rufipogon has the same type of genome with the cultivated rice (O. sativa), Pi-ta(+) and Pib gene in Erect type of O. rufipogon can be used to improve the tolerance of cultivated rice to blast, either by traditional hybridization or by genetic engineering.

A 4,672 bp DNA sequence including the whole coding region and partial non-coding region of rice blast resistance gene Pi-ta+ has been cloned from Jinghong erect type of common wild rice (Oryza rufipogon Griff) in Yunnan by polymerase chain reaction method. The coding region shares 99.86% and 98.78% identity with the corresponding regions of the reported cultivated rice Yashiro-mochi and Yuanjiang type of common wild rice respectively. There are 4 nucleotides difference in the coding region and 6 in intron of the cloned Pi-ta+ gene,compared with Pi-ta from Yashiro-mochi. Pi-ta+ gene in Jinghong erect type of common wild rice has been proved to be a rare existing Pi-ta+ allele, because there was a alanine rather than a serine at the position 918 within the predicted amino acid sequence of PITA. Pi-ta+ allele can cause disease resistance response to rice blast pathogens in plant cells. Differences in DNA sequence, deduced amino acid sequence and antibacterial spectrum may make the Pi-ta+ allele new resistant characteristics. Finding and cloning of Pi-ta+ allele from Jinghong erect type of common wild rice in Yunnan provides a basement for further utilization of the wild rice resources.

Mapping a target gene is useful to isolate the gene or get molecular marker for molecular marker-assisted selec- tion. This paper summarized the recent studies of gene mapping in rice, including 194 genes. These genes are involved in disease resistance, cold tolerance, drought resistance, sterility, and fertility restoration of rice. The molecular markers that link with these genes were also summarized. Among these genes 14 have been cloned and sequenced. According to the results this paper also analyzed genes distribution rule in chromosomes.

The Xa21 gene previously cloned from the wild rice species Oryzae longistaminata confers broad-spectrum resistance to rice leaf blight caused by different strains of Xanthomonas oryzae pv. oryzae. Here we attempted to determine the existence of Xa21 homologs in other wild rice species and rice cultivars and the sequence differences between the homologs. We synthesized specific primers based on the reported Xa21 sequence to amplify homologs of the gene exon II from several rice cultivars and three wild rice species in Yunnan Province, China. The fragments cloned from various types of O. rufipogon Griff from Jinghong and Yuanjiang, Yunnan Province, were highly homologous to the reported Xa21 gene exon II. However, the fragment was not found in O. officinalis Wall. and O. meyeriana Baill. Sequence analysis suggested that differences in nucleotides were located randomly in the fragments we cloned.

A refractile body antigen Etp28 Gene of Eimeria tenella (Guangdong Strain) sporozoites was cloned by PCR from the synthesized first strand cDNA. It has a high homology with the previously reported Etp28 gene of Merck Strain LS18. The gene was expressed through standard procedures in the modified Autographa californica nuclear polyhedrosis virus (AcMNPV-OCC(-)) expression system and large amount of heterologous fusion protein (GST-6xHis-Etp28) was obtained. The expression product was about 21.3% of the total cellular protein of an infected cell and corresponding to approximately 0.42 mg of recombinant protein per 10(6) cells. And the immunoprotective trial showed that this candidate for recombinant vaccine conferred partial protection against chicken coccidiosis.