The Application Effect of Doctor-Nurse Collaborative and Hierarchical Management Combined with Nursing Risk Management in Nursing Management of Patients with Postpartum Hemorrhage.

Background: To study the application effect of doctor-nurse collaborative and hierarchical management combined with nursing risk management in nursing management of patients with postpartum hemorrhage. Methods: Overall 200 patients with postpartum hemorrhage treated in Guangdong Second Provincial General Hospital (Guangzhou, China) from Mar 2018 to Mar 2019 were divided into the experimental group and the control group. The experimental group received the nursing model of doctor-nurse collaborative and hierarchical management combined with nursing risk management while the control group received nursing risk management to compare the satisfaction, medical record quality before and after nursing, incidence of nursing risk events and treatment effect between the two groups. Results: After nursing management, the bleeding volume in the experimental group was significantly less than that in the control group (P<0.05). The quality score of nursing records in the experimental group was higher than that in the control group (P<0.01). The occurrence of nursing risk events in the experimental group was significantly less than that in the control group (P<0.05). The overall nursing satisfaction of the experimental group was higher than that of the control group (P<0.01). The treatment effect of the experimental group was significantly better than that of the control group (P<0.01). Conclusion: Doctor-nurse collaborative and hierarchical management combined with nursing risk management had a significant effect in the nursing management of patients with postpartum hemorrhage, which is worthy of promotion and application.

The Impact of a Knowledge Discovery-Based Psychoanalytic Intervention in the Treatment of Tuberculosis in University Students with Different Doses of Isoniazid.

Tuberculosis (TB) is an infectious disease that poses a serious threat to the health of the population in China, and TB outbreaks in universities have aroused great concern in society. Psychological emotions have a large impact on the academic lives of university students, and nowadays it is not only labour-intensive but also slow to monitor and analyse and deal with the psychology of university students' daily lives in a uniform manner. If psychological problems are not detected and given feedback in a timely manner, they can have a series of negative effects on the individual university student. In this paper, we apply the Bi-LSTM model and the CNN model neural network algorithm to learn the text data, and finally have 95.55% and 90.03% accuracy in the sentiment analysis experiment, respectively, which provides a feasible solution to solve the batch rapid analysis of the psychological changes reflected in the daily text of university students. Risk communication for TB emergencies should emphasize public participation, timely release of information about the epidemic, and good monitoring of public opinion.

Coibamide A kills cancer cells through inhibiting autophagy.

Natural products are useful tools for biological mechanism research and drug discovery. Due to the excellent tumor cell growth inhibitory profile and sub-nanomolar potency, Coibamide A (CA), an N-methyl-stabilized depsipeptide isolated from marine cyanobacterium, has been considered as a promising lead compound for cancer treatment. However, the molecular anti-cancer mechanism of the action of CA remains unclear. Here, we showed that CA treatment induced caspase-independent cell death in breast cancer cells. CA treatment also led to severe lysosome defects, which was ascribed to the impaired glycosylation of lysosome membrane protein LAMP1 and LAMP2. As a consequence, the autophagosome-lysosome fusion was blocked upon CA treatment. In addition, we presented evidence that this autophagy defect partially contributed to the CA treatment-induced tumor cell death. Together, our work uncovers a novel mechanism underlying the anti-cancer action of CA, which will promote its further application for cancer therapy.

TALEN-based editing of TFIIAy5 changes rice response to Xanthomonas oryzae pv. Oryzae.

The xa5 gene encodes a basal transcription factor (TFIIAγ) protein with wide spectrum resistance to bacterial blight caused by Xanthomonas oryzae pv. Oryzae (Xoo) in rice. It was only found in a few rice ecotypes, and the recessive characteristics limited its application in breeding. Here, we employed a TALEN-based technique to edit its dominant allelic TFIIAγ5 and obtained many mutant TFIIAγ5 genes. Most of them reduced rice susceptibility to varying degrees when the plants were challenged with the Xoo. In particular, the knocked-out TFIIAγ5 can reduce the rice susceptibility significantly, although it cannot reach the xa5-mediated resistance level, indicating TFIIAγ5 is a major component involved in disease susceptibility. In addition, the mutant encoding the protein with deletion of the 32nd amino acid or amino acid insertion between 32nd and 33rd site confers rice with the similar resistance to that of the knocked-out TFIIAγ5. Thus, the amino acids around 32nd site are also the important action sites of TFIIAγ5 besides the 39th amino acid previously reported. Moreover, the integration of xa5 into TFIIAγ5-knockout plants conferred them with a similar resistance as IRBB5, the rice variety containing the homozygous xa5 gene. Thus, TFIIAγ5 was not simply regarded as a resistant or a susceptible locus, as the substitution of amino acids might shift its functions.

Complete genome sequence of a novel capillovirus infecting Hevea brasiliensis in China.

Tapping panel dryness (TPD) is a complex disorder that causes partial or complete cessation of latex drainage upon tapping of rubber trees (Hevea brasiliensis). In this work, we determined the complete genome sequences of a novel virus identified in a rubber tree with TPD syndrome in China. The genome of the virus consists of 6811 nt and possesses two overlapping open reading frames (ORF1 and ORF2), encoding a polyprotein and a movement protein, respectively. The polyprotein shares 37% amino acid sequence identity with cherry virus A (CVA, ARQ83874.1) over 99% coverage. The genome architecture is similar to that of members of the genus Capillovirus (family Betaflexiviridae). Phylogenetic analysis of the replicase proteins showed that the virus clustered together with members of the genus Capillovirus. The new virus is tentatively called "rubber tree virus 1" (RTV1). RTV1 is the first virus reported to infect rubber trees. This work lays a foundation for research into finding the potential causal agent of TPD in Hevea brasiliensis.

Characterization, expression, and antimicrobial activity of histones from Japanese flounder Paralichthys olivaceus.

Histone proteins are not only structurally important for chromosomal DNA packaging but also involved in the regulation of gene expression and the immune response of host against pathogens. Japanese flounder (Paralichthys olivaceus) as one of the most important marine flatfish, suffered from widespread outbreaks of diseases, and its immunological functioning remained to be elucidated. In the present study, we reported the expression patterns of four histones (H1, H2A, H3, and H3.3) and functional characterization of the histone H3.3 from flounder. Quantitative real time RT-PCR (RT-qPCR) analysis showed that expression of the four histones occurred in multiple tissues, but their levels of expression were relatively high in immune organs, and inducible in response to pathogens infection. Infection with extracellular and intracellular bacterial pathogens and viral pathogen regulated the expression of histones in a manner that depended on tissue type, pathogen, and infection stage. Specifically, H1 expression was highly induced by intracellular viral pathogens; H2AX and H3 expressions were highly induced by intracellular bacterial pathogen; dissimilarly, H3.3 expression was slightly induced by extracellular bacterial pathogen, but was inhibited by intracellular bacterial and viral pathogens. To further investigate H3.3 function, recombinant H3.3 (rH3.3) was obtained, and in vitro experiments showed rH3.3 possessed the capability of binding to both Gram-negative and Gram-positive bacteria and inhibiting the growth of some target bacteria. Consistently, In vivo results showed that overexpression of H3.3 promoted the host defense against invading pathogenic microorganism and regulated the expressions of several cytokines. These results suggested that flounder histones exhibit different expression patterns in response to the infection of different microbial pathogens, and H3.3 serves as an immune-related protein and plays an important role in antimicrobial immunity of Japanese flounder. Taken together, this study is the first report about the expression profile of different histones upon different kind of pathogens and anti-infectious immunity of H3.3 in teleost, which offered new insights into the immunological function of histones in teleost.

Major histocompatibility complex class I (MHC Iα) of Japanese flounder (Paralichthys olivaceus) plays a critical role in defense against intracellular pathogen infection.

The major histocompatibility complex (MHC) is a highly polymorphic region of the vertebrate genome that plays a critical role in initiating immune responses towards invading pathogens. It is well known that MHC I molecules play a central role in the immune response to viruses. However, rare literatures were reported the role of MHC I in the resistance to intracellular bacteria. Sequences of MHC Iα were identified in multiple teleost species, including Japanese flounder (Paralichthys olivaceus), however, the immunological function of MHC Iα remain largely unknown. In this study, we examined the expression profile and biological activity of an MHC Iα homologue, PoMHC Iα, from P. olivaceus. Structural analysis showed that PoMHC Iα possesses conserved structural characteristics of MHC Iα proteins, including MHC_I domain, IGc1 domain, transmembrane region. Expression of PoMHC Iα was upregulated in a time-dependent manner by extracellular and intracellular bacterial pathogens and viral pathogen infection. Different expression patterns were exhibited in response to the infection of different types of microbial pathogens in different immune tissues. Recombinant PoMHC Iα increased the capability of host cells to defense against intracellular pathogen Edwardsiella tarda infection and enhanced the expression of immune related genes. The knockdown of PoMHC Iα attenuated the ability of cells to eliminate E. tarda, which was sustained by the in vivo results that overexpression of PoMHC Iα promoted the host defense against invading E. tarda. Antigen uptake assay indicated PoMHC Iα participated in cells antigen presentation. Collectively, this study is the first report that MHC Iα plays an important role in immune defense against intracellular bacterial pathogen in teleost. Taken together, these findings add new insights into the biological function of teleost MHC Iα and emphasize the importance of MHC I gene products for the control of E. tarda infection.

Transcriptomic analyses reveal molecular mechanisms underlying growth heterosis and weakness of rubber tree seedlings.

BACKGROUND: Breeding rubber tree seedling with growth heterosis is vital for natural rubber production. It is the prerequisites for effectively utilizing growth heterosis to elucidate its molecular mechanisms, but the molecular mechanisms remain poorly understood in rubber tree. To elucidate seedling growth heterosis, we conducted comparative transcriptomic analyses between the two hybrids and their parents. RESULTS: By identifying and comparing differently expressed genes (DEGs), we found that the hybrids (BT 3410 and WC 11) show significantly differential expression profiles from their parents (PR 107 and RRIM 600). In BT 3410-parent triad, 1092 (49.95%) and 1094 (50.05%) DEGs indicated clear underdominance or overdominance, respectively. Whereas in WC 11-parent triad, most DEGs (78.2%, 721) showed low- or high-parent dominance; 160 (17.35%) exhibited expression patterns that are not statistically distinguishable from additivity, and 8 (0.87%) and 33 (3.58%) DEGs exhibited underdominance and overdominance, respectively. Furthermore, some biological processes are differentially regulated between two hybrids. Interestingly, the pathway in response to stimulus is significantly downregulated and metabolic pathways are more highly regulated in BT 3410. CONCLUSIONS: Taken together, the genotypes, transcriptomes and biological pathways (especially, carbohydrate metabolism) are highly divergent between two hybrids, which may be associated with growth heterosis and weakness. Analyzing gene action models in hybrid-parent triads, we propose that overdominance may play important roles on growth heterosis, whereas dominance on hybrid weakness in rubber tree seedlings. These findings bring new insights into our understanding of growth heterosis of rubber tree seedling.

Molecular cloning and characterization of S-adenosylmethionine decarboxylase gene in rubber tree (Hevea brasiliensis).

S-Adenosylmethionine decarboxylase (SAMDC) is a key rate-limiting enzyme involved in polyamines biosynthesis, and it plays important roles in plant growth, development and stresses response. However, no SAMDC gene was reported in rubber tree. Here we report characteristics of an SAMDC gene (HbSAMDC1) in rubber tree. HbSAMDC1 contains a 1080 bp open reading frame (ORF) encoding 359 amino acids. Quantitative real-time PCR analyses revealed that HbSAMDC1 exhibited distinct expression patterns in different tissues and was regulated by various stresses, including drought, cold, salt, wounding, and H2O2 treatments. HbSAMDC1 5' untranslated region (UTR) contains a highly conserved overlapping tiny and small upstream ORFs (uORFs), encoding 2 and 52 amino acid residues, respectively. No introns were located in the main ORF of HbSAMDC1, whereas two introns were found in the 5' UTR. In transgenic tobaccos, the highly conserved small uORF of HbSAMDC1 is found to be responsible for translational repression of downstream ß-glucuronidase reporter. To our knowledge, this is the first report on molecular cloning, expression profiles, and 5' UTR characteristics of HbSAMDC1. These results lay solid foundation for further elucidating HbSAMDC1 function in rubber tree.

Expression Profiles, Characterization and Function of HbTCTP in Rubber Tree (Hevea brasiliensis).

As a highly conserved protein, the translationally controlled tumor protein (TCTP) carries out vital roles in various life processes. In rubber tree, two TCTP genes, HbTCTP and HbTCTP1, were cloned, but only HbTCTP1 was studied in details. In this study, cis-acting regulatory elements, expression patterns, subcellular localization, interacting proteins, and antioxidant activity of HbTCTP were systematically analyzed. Besides the common cis-acting regulatory elements, HbTCTP promoter also harbored various known cis-elements that respond to hormone/stresses. Being consistent with the aforementioned results, HbTCTP was regulated by drought, low temperature, high salt, ethylene (ET), wounding, H2O2, and methyl jasmonate (MeJA) treatments. HbTCTP was expressed throughout different tissues and developmental stages of leaves. In addition, HbTCTP was associated with tapping panel dryness (TPD). HbTCTP was localized in the membrane, cytoplasm and the nucleus, and interacted with four proteins rubber elongation factor (REF), 17.5 kDa heat shock family protein, annexin, and REF-like stress related protein 1. Being similar to HbTCTP1, HbTCTP also indicated antioxidant activity in metal-catalyzed oxidation (MCO) system. Our results are useful for further understanding the molecular characterization and expression profiles of HbTCTP, but also lay a solid foundation for elucidating the function of HbTCTP in rubber tree.

[Integration and expression of Xa21 in transgenic rice CX8621].

Agrobacterium tumefaciens-mediated transformation system has been widely applied. However, the function of target gene is affected by multiple factors. With this system, we obtained a transgenic rice line CX8621 carrying the bacterial blight resistance gene Xa21. In previous work, we have confirmed that it was selectable maker-free and vector backbone-free. And after 16 generations of breeding, it still maintained perfect resistance to bacterial blight disease. On this basis, we analyzed the integration and expression of Xa21 in CX8621 at the present study. First, based on the border sequences of plasmid pBXa21 and Xa21, we designed nested primers and assured the integrity of Xa21 in CX8621. Second, we cloned the flanking sequences and located Xa21 on chromosome 2 using improved Tail-PCR. Then we analyzed the expression pattern of Xa21 in several tissues and at different developmental stages by RT-PCR. The results show that Xa21 can be stably expressed in CX8621, agreeing well with the disease resistance response as reported previously. In addition, we detected the protein levels of XA21 in CX8621 with antibody of natural XA21 protein. Surprisingly, no XA21 protein was detected in the seeds of CX8621. Thus, the integration and expression analysis of Xa21 in CX8621 provided a part of scientific evidences for the safety assessment of genetically modified rice.

Rice Xa21 primed genes and pathways that are critical for combating bacterial blight infection.

Rice bacterial blight (BB) is a devastating rice disease. The Xa21 gene confers a broad and persistent resistance against BB. We introduced Xa21 into Oryza sativa L ssp indica (rice 9311), through multi-generation backcrossing, and generated a nearly isogenic, blight-resistant 9311/Xa21 rice. Using next-generation sequencing, we profiled the transcriptomes of both varieties before and within four days after infection of bacterium Xanthomonas oryzae pv. oryzae. The identified differentially expressed (DE) genes and signaling pathways revealed insights into the functions of Xa21. Surprisingly, before infection 1,889 genes on 135 of the 316 signaling pathways were DE between the 9311/Xa21 and 9311 plants. These Xa21-mediated basal pathways included mainly those related to the basic material and energy metabolisms and many related to phytohormones such as cytokinin, suggesting that Xa21 triggered redistribution of energy, phytohormones and resources among essential cellular activities before invasion. Counter-intuitively, after infection, the DE genes between the two plants were only one third of that before the infection; other than a few stress-related pathways, the affected pathways after infection constituted a small subset of the Xa21-mediated basal pathways. These results suggested that Xa21 primed critically important genes and signaling pathways, enhancing its resistance against bacterial infection.

Molecular cloning, expression profiles, and characterization of a novel polyphenol oxidase (PPO) gene in Hevea brasiliensis.

The polyphenol oxidase (PPO) is involved in undesirable browning in many plant foods. Although the PPOs have been studied by several researchers, the isolation and expression profiles of PPO gene were not reported in rubber tree. In this study, a new PPO gene, HbPPO, was isolated from Hevea brasiliensis. The sequence alignment showed that HbPPO indicated high identities to plant PPOs and belonged to dicot branch. The cis-acting regulatory elements related to stress/hormone responses were predicted in the promoter region of HbPPO. Real-time RT-PCR analyses showed that HbPPO expression varied widely depending on different tissues and developmental stages of leaves. Besides being associated with tapping panel dryness, the HbPPO transcripts were regulated by ethrel, wounding, H2O2, and methyl jasmonate treatments. Moreover, the correlation between latex coagulation rate and PPO activity was further confirmed in this study. Our results lay the foundation for further analyzing the function of HbPPO in rubber tree.

Functional characterization of a wheat NHX antiporter gene TaNHX2 that encodes a K(+)/H(+) exchanger.

The subcellular localization of a wheat NHX antiporter, TaNHX2, was studied in Arabidopsis protoplasts, and its function was evaluated using Saccharomyces cerevisiae as a heterologous expression system. Fluorescence patterns of TaNHX2-GFP fusion protein in Arabidopsis cells indicated that TaNHX2 localized at endomembranes. TaNHX2 has significant sequence homology to NHX sodium exchangers from Arabidopsis, is abundant in roots and leaves and is induced by salt or dehydration treatments. Western blot analysis showed that TaNHX2 could be expressed in transgenic yeast cells. Expressed TaNHX2 protein suppressed the salt sensitivity of a yeast mutant strain by increasing its K(+) content when exposed to salt stress. TaNHX2 also increased the tolerance of the strain to potassium stress. However, the expression of TaNHX2 did not affect the sodium concentration in transgenic cells. Western blot analysis for tonoplast proteins indicated that the TaNHX2 protein localized at the tonoplast of transgenic yeast cells. The tonoplast vesicles from transgenic yeast cells displayed enhanced K(+)/H(+) exchange activity but very little Na(+/)H(+) exchange compared with controls transformed with the empty vector; Na(+)/H(+) exchange was not detected with concentrations of less than 37.5 mM Na(+) in the reaction medium. Our data suggest that TaNHX2 is a endomembrane-bound protein and may primarily function as a K(+)/H(+) antiporter, which is involved in cellular pH regulation and potassium nutrition under normal conditions. Under saline conditions, the protein mediates resistance to salt stress through the intracellular compartmentalization of potassium to regulate cellular pH and K(+) homeostasis.

Do transgenesis and marker-assisted backcross breeding produce substantially equivalent plants? A comparative study of transgenic and backcross rice carrying bacterial blight resistant gene Xa21.

BACKGROUND: The potential impact of genetically modified (GM) plants on human health has attracted much attention worldwide, and the issue remains controversial. This is in sharp contrast to the broad acceptance of plants produced by breeding through Marker Assisted Backcrossing (MAB). RESULTS: Focusing on transcriptome variation and perturbation to signaling pathways, we assessed the molecular and biological aspects of substantial equivalence, a general principle for food safety endorsed by the Food and Agricultural Organization and the World Health Organization, between a transgenic crop and a plant from MAB breeding. We compared a transgenic rice line (DXT) and a MAB rice line (DXB), both of which contain the gene Xa21 providing resistance to bacterial leaf blight. By using Next-Generation sequencing data of DXT, DXB and their parental line (D62B), we compared the transcriptome variation of DXT and DXB. Remarkably, DXT had 43% fewer differentially expressed genes (DEGs) than DXB. The genes exclusively expressed in DXT and in DXB have pathogen and stress defense functions. Functional categories of DEGs in DXT were comparable to that in DXB, and seven of the eleven pathways significantly affected by transgenesis were also perturbed by MAB breeding. CONCLUSIONS: These results indicated that the transgenic rice and rice from MAB breeding are substantial equivalent at the transcriptome level, and paved a way for further study of transgenic rice, e.g., understanding the chemical and nutritional properties of the DEGs identified in the current study.

Characterization of Ca(2+)/H(+) exchange in the plasma membrane of Saccharomyces cerevisiae.

The characteristics of the Ca(2+)/H(+) exchange were directly investigated in functionally inverted (inside-out) plasma membrane vesicles isolated from yeast using an aqueous two-phase partitioning method. Results showed that following the generation of an inside-acid pH gradient (fluorescence quenching), addition of Ca(2+) caused movement of H(+) out of the vesicles (fluorescence recovery). The Ca(2+)/H(+) exchange displayed saturation kinetics with respect to extravesicular Ca(2+) and ATP concentrations in the plasma membrane, and showed specificity for Ca(2+). The protonophore FCCP (carbonyl cyanide p-(trifluoromethoxy) phenylhydrazone), abolished the fluorescence quenching and consequently inhibited Ca(2+)/H(+) exchange in plasma membrane vesicles. Vanadate, which is known to inhibit the plasma membrane H(+)-ATPase, significantly decreased the Ca(2+)-dependent transport of H(+) out of vesicles. When the electrical potential across the plasma membrane was dissipated with valinomycin and potassium, the rate of Ca(2+)/H(+) exchange increased compared to that of the control without valinomycin, indicating that the stoichiometric ratio for this exchange is greater than 2H(+):Ca(2+). These data suggest that Ca(2+) is transported out of yeast cells through a Ca(2+)/H(+) exchange system that is driven by the proton-motive force generated by the plasma membrane H(+)-ATPase.

RNA-Seq analysis of Cocos nucifera: transcriptome sequencing and de novo assembly for subsequent functional genomics approaches.

BACKGROUND: Cocos nucifera (coconut), a member of the Arecaceae family, is an economically important woody palm grown in tropical regions. Despite its agronomic importance, previous germplasm assessment studies have relied solely on morphological and agronomical traits. Molecular biology techniques have been scarcely used in assessment of genetic resources and for improvement of important agronomic and quality traits in Cocos nucifera, mostly due to the absence of available sequence information. METHODOLOGY/PRINCIPAL FINDINGS: To provide basic information for molecular breeding and further molecular biological analysis in Cocos nucifera, we applied RNA-seq technology and de novo assembly to gain a global overview of the Cocos nucifera transcriptome from mixed tissue samples. Using Illumina sequencing, we obtained 54.9 million short reads and conducted de novo assembly to obtain 57,304 unigenes with an average length of 752 base pairs. Sequence comparison between assembled unigenes and released cDNA sequences of Cocos nucifera and Elaeis guineensis indicated that the assembled sequences were of high quality. Approximately 99.9% of unigenes were novel compared to the released coconut EST sequences. Using BLASTX, 68.2% of unigenes were successfully annotated based on the Genbank non-redundant (Nr) protein database. The annotated unigenes were then further classified using the Gene Ontology (GO), Clusters of Orthologous Groups (COG) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases. CONCLUSIONS/SIGNIFICANCE: Our study provides a large quantity of novel genetic information for Cocos nucifera. This information will act as a valuable resource for further molecular genetic studies and breeding in coconut, as well as for isolation and characterization of functional genes involved in different biochemical pathways in this important tropical crop species.

Development and characterization of intron-flanking EST-PCR markers in rubber tree (Hevea brasiliensis Muell. Arg.).

With a long-term goal of constructing a linkage map enriched with gene-specific markers in rubber tree (Hevea brasiliensis Muell. Arg.), we utilized rubber tree ESTs associated with tapping panel dryness (TPD) to develop intron-flanking PCR markers. After downloading and assembling the rubber tree ESTs associated with TPD, we predicted the exon/exon junction sites (E/E) by aligning rubber tree transcripts with the genomic sequences of castor bean (Ricinus communis L.). Based on the predicted E/E, the primers flanking intron(s) and no intron were designed. Compared with the markers designed by conventional method, the PCR success rate of the markers designed with the predicted E/E increased 28-30%, whereas the polymorphism rate of intron-flanking EST-PCR markers was approximately 3.43-fold increase. Therefore, the intron-flanking marker was more polymorphism-generating efficient than the markers designed by conventional methods. In addition, analyzing the polymorphic information content (PIC) among Hevea germplasm showed that the polymorphism of wild rubber tree accessions was higher than one of cultivated rubber tree clones and Hevea species. This study enriches the categories and numbers of molecular markers in rubber tree, and the markers developed in this research will have a wide application in DNA fingerprinting, marker-assisted selection and genetic mapping in rubber tree. This research also indicates that it is possible to develop intron-flanking EST-PCR markers of rubber tree with castor bean genome as reference sequences, which provides new insights into developing intron-flanking EST-PCR markers for rubber tree or other plant species without genomic information.

RNA-Seq analysis and de novo transcriptome assembly of Hevea brasiliensis.

Hevea brasiliensis, being the only source of commercial natural rubber, is an extremely economically important crop. In an effort to facilitate biological, biochemical and molecular research in rubber biosynthesis, here we report the use of next-generation massively parallel sequencing technologies and de novo transcriptome assembly to gain a comprehensive overview of the H. brasiliensis transcriptome. The sequencing output generated more than 12 million reads with an average length of 90 nt. In total 48,768 unigenes (mean size = 436 bp, median size = 328 bp) were assembled through de novo transcriptome assembly. Out of 13,807 H. brasiliensis cDNA sequences deposited in Genbank of the National Center for Biotechnology Information (NCBI) (as of Feb 2011), 11,746 sequences (84.5%) could be matched with the assembled unigenes through nucleotide BLAST. The assembled sequences were annotated with gene descriptions, Gene Ontology (GO) and Clusters of Orthologous Group (COG) terms. In all, 37,432 unigenes were successfully annotated, of which 24,545 (65.5%) aligned to Ricinus communis proteins. Furthermore, the annotated uingenes were functionally classified according to the GO, COG and Kyoto Encyclopedia of Genes and Genomes databases. Our data provides the most comprehensive sequence resource available for the study of rubber trees as well as demonstrates effective use of Illumina sequencing and de novo transcriptome assembly in a species lacking genomic information.

Identification and characterization of genes associated with tapping panel dryness from Hevea brasiliensis latex using suppression subtractive hybridization.

BACKGROUND: Tapping panel dryness (TPD) is one of the most serious threats to natural rubber production. Although a great deal of effort has been made to study TPD in rubber tree, the molecular mechanisms underlying TPD remain poorly understood. Identification and systematical analyses of the genes associated with TPD are the prerequisites for elucidating the molecular mechanisms involved in TPD. The present study is undertaken to generate information about the genes related to TPD in rubber tree. RESULTS: To identify the genes related to TPD in rubber tree, forward and reverse cDNA libraries from the latex of healthy and TPD trees were constructed using suppression subtractive hybridization (SSH) method. Among the 1106 clones obtained from the two cDNA libraries, 822 clones showed differential expression in two libraries by reverse Northern blot analyses. Sequence analyses indicated that the 822 clones represented 237 unique genes; and most of them have not been reported to be associated with TPD in rubber tree. The expression patterns of 20 differentially expressed genes were further investigated to validate the SSH data by reverse transcription PCR (RT-PCR) and real-time PCR analysis. According to the Gene Ontology convention, 237 unique genes were classified into 10 functional groups, such as stress/defense response, protein metabolism, transcription and post-transcription, rubber biosynthesis, etc. Among the genes with known function, the genes preferentially expressed were associated with stress/defense response in the reverse library, whereas metabolism and energy in the forward one. CONCLUSIONS: The genes associated with TPD were identified by SSH method in this research. Systematic analyses of the genes related to TPD suggest that the production and scavenging of reactive oxygen species (ROS), ubiquitin proteasome pathway, programmed cell death and rubber biosynthesis might play important roles in TPD. Therefore, our results not only enrich information about the genes related to TPD, but also provide new insights into understanding the TPD process in rubber tree.