Project description:Sugarcane plantlets from a variety with high inputs of N obtained from BNF (genotype SP70-1143, CTC, Brazil) free of microorganisms were obtained by sterile meristem culture and micropropagation according to the method of Hendre et al. (1983). In vitro-grown SP70-1143 rooted sugarcane plantlets were inoculated as described by James et al. (1994) with 0.1 ml of 106–107 bacterial suspension. Controls were inoculated with medium only. Endophytic diazotrophic bacteria used were Gluconacetobacter diazotrophicus (PAL5 strain) or a mixture of Herbaspirillum seropedicae (HRC54 strain) and H. rubrisubalbicans (HCC103 strain). All plants were maintained at 30°C with an irradiance of 60 µmol photons m–2 s–1 for 12 h d–1. One day after the inoculation, plant tissues were examined for bacterial colonization by the Most Probable Number (MPN) estimation, according to the methods of Reis et al. (1994) and plantlets were collected and immediately frozen in liquid nitrogen. Five plantlets were polled for each treatment. Extraction of total RNA was performed separately on each sample pool. Keywords: comparison of associations with different endophytic bacterias
Project description:To explore the molecular mechanism of low-K tolerance in sugarcane, we have employed whole genome microarray expression profiling to identify sugarcane genes in response to low-K stress. seeldings were transplanted to low-K hydroponic (containing 0.1 mmol.L-1 K+) and the roots were collected at 0 (CK), 8, 24 and 72 h after exposure to low-K condition. The expressions of genes in sugarcane roots were detected by microarray analysis. Totally 1545 genes at 8 h, 1053 genes at 24 h and 3155 at 72 h differentially expressed under low-K stress, when the 2-fold change was adopted as the threshold for determining differentially expressed genes. Among these genes, a certain amount of transcription factors, transporters, kinases, oxidative stress-related genes and genes in Ca+ and ethylene signaling pathway were detected to differentially express. Seeldings were treated with low-K hydroponic (containing 0.1 mmol.L-1 K+) and after 0 (CK), 8, 24 and 72 h exposure to low -K stress, the roots of sugarcane were collected. Four independent experiments were performed using roots collected at different time points
Project description:In order to increase our understanding on the epigenetic regulation in response to abiotic stresses in plants, sRNA regulation in sugarcane plants submitted to drought stress was analyzed. Deep sequencing analysis was carried out to identify the sRNA regulated in leaves and roots of sugarcane cultivars with different drought sensitivities. An enrichment of 22-nt sRNA species was observed in leaf libraries. The pool of sRNA selected allowed the analysis of different sRNA classes (miRNA and siRNA). Twenty eight and 36 families of conserved miRNA were identified in leaf and root libraries, respectively. Dynamic regulation of miRNA was observed and the expression profile of eight miRNA was verified in leaf samples by stem-loop qRT-PCR assay. Altered miRNA regulation was correlated with changes in mRNA levels of specific targets. 22-nt miRNA triggered siRNA-candidates production by cleavage of their targets in response to drought stress. Some genes of sRNA biogenesis were down-regulated in tolerant genotypes and up-regulated in sensitive in response to drought stress. Our analysis contributes to increase the knowledge on the roles of sRNA in epigenetic-regulatory pathways in sugarcane submitted to drought stress. Screenning of sRNA transcriptome of sugarcane plants under drougth stress
Project description:In order to increase our understanding on the epigenetic regulation in response to abiotic stresses in plants, sRNA regulation in sugarcane plants submitted to drought stress was analyzed. Deep sequencing analysis was carried out to identify the sRNA regulated in leaves and roots of sugarcane cultivars with different drought sensitivities. An enrichment of 22-nt sRNA species was observed in leaf libraries. The pool of sRNA selected allowed the analysis of different sRNA classes (miRNA and siRNA). Twenty eight and 36 families of conserved miRNA were identified in leaf and root libraries, respectively. Dynamic regulation of miRNA was observed and the expression profile of eight miRNA was verified in leaf samples by stem-loop qRT-PCR assay. Altered miRNA regulation was correlated with changes in mRNA levels of specific targets. 22-nt miRNA triggered siRNA-candidates production by cleavage of their targets in response to drought stress. Some genes of sRNA biogenesis were down-regulated in tolerant genotypes and up-regulated in sensitive in response to drought stress. Our analysis contributes to increase the knowledge on the roles of sRNA in epigenetic-regulatory pathways in sugarcane submitted to drought stress.
Project description:A collection of 237,000 expressed sequence tags generated by the Sugarcane EST sequencing project (SUCEST) was analyzed in search of signal transduction components. The SUCAST (Sugarcane Signal Transduction) Catalogue contains over 3500 components, with around 2900 involved in several aspects of cell signaling and transcription. Sequence comparisons and conserved protein domain analysis revealed 477 receptors, 510 protein kinases, 107 protein phosphatases, a large number of small GTPases, G-proteins, members of the calcium and inositol metabolism, and other signal transduction-related proteins. Over 600 transcription factors were also indexed. Moreover, 437 genes with no matches in the public databases and 111 genes of unknown function were catalogued. Several of the SUCEST cDNA libraries were derived from plants submitted to abiotic stresses or infected with endophytic nitrogen fixing bacteria and stress and pathogen response-related genes were also annotated. The abundance of transcripts among six different sugarcane tissues (flowers, roots, leaves, lateral buds, 1st and 4th internodes) was evaluated using microarrays and expression profile clustering. We identified 216 genes that are significantly more abundant in one of the tissues analyzed. A subset of the data was validated by real-time PCR. Additionally, genes with similar expression levels among different tissues were identified. The characterization of these elements and their promoters can aid in the development of tools for the genetic manipulation of this plant species and other economically important grasses. Keywords: other
Project description:The soldier fly is an endemic pest of sugarcane in Australia. Small numbers of larvae can cause significant damage to roots and reduce the crop yields. Little is known about the composition and function of the soldier fly salivary gland, its secretions, and their roles in insect-plant interactions. In this study, we performed transcriptome analysis of the salivary glands of starved and sugarcane root-fed soldier fly larvae. A total of 31,119 highly expressed assembled contigs were identified in the salivary glands and almost 50% of them showed high levels of similarity to known proteins in Nr databases. Of all the obtained contigs, only 9,727 sequences contain an open reading frame of over 100 amino acids. Around 31% of contigs were predicted to encode secretory proteins, including some digestive and detoxifying enzymes and potential effectors. Some known salivary secreted peptides such as serine protease, cysteine proteinase inhibitors, antimicrobial peptides and venom proteins were among the top 100 highly expressed genes. Differential gene expression analysis revealed significant modulation of 850 transcripts in salivary glands upon exposure to plant roots or starvation stress. Here, we identified some venom proteins which were significantly upregulated in the salivary glands of soldier fly larvae exposed to sugarcane roots. In other insects and nematodes some of these proteins have been used to manipulate host plant defense systems and facilitate the invasion of the host plant. These findings provide a further insight into the identification of potential effector proteins involved in soldier fly- sugarcane interactions.
Project description:Endophytic bacteria influence plant growth and development and therefore are an attractive resource for applications in agriculture. However, little is known about the impact of these microorganisms on secondary metabolite (SM) production by medicinal plants. Here we assessed, for the first time, the effects of root endophytic bacteria on the modulation of SMs in the medicinal plant Lithospermum officinale (Boraginaceae family), with a focus on the naphthoquinones alkannin/shikonin (A/S). The study was conducted using a newly developed in vitro system as well as in the greenhouse. Targeted and non-targeted metabolomics approaches were used and supported by expression analysis of the gene PGT, encoding a key enzyme in the A/S biosynthesis pathway. Three bacterial strains, Chitinophaga sp. R-73072, Xanthomonas sp. R-73098 and Pseudomonas sp. R-71838 induced a significant increase of diverse SMs, including A/S, in L. officinale in both systems, demonstrating the strength of our approach for screening A/S derivative-inducing bacteria. Our results highlight the impact of root-endophytic bacteria on secondary metabolism in plants and indicate that production of A/S derivatives in planta likely involves cross-modulation of different metabolic pathways that can be manipulated by bacterial endophytes.