Transcriptomic response to Nod Factor treatments on Medicago
ABSTRACT: NGS2013-04: Transcriptomic response to Nod Factor treatments on Medicago Role of the root hair in water and nutrient uptake and the establishment of the nitrogen-fixing symbiotic interaction with rhizobia. The RNA was extracted from root hairs of Medicago: control vs treated by nod factors (2 biological replicates)
Project description:affy_ralstonia_medicago - Ralstonia solanacearum is the causal agent of the devastating bacterial wilt disease. Its infection process was studied with an in vitro inoculation procedure on intact roots of Medicago truncatula. The pathosystem involved susceptible A17 and resistant F83005.5 M truncatula lines infected with the pathogenic strain GMI1000. The mutant A17 line, Sickle, which showed a resistant phenotype was also part of the experiment. To identify host signaling pathway triggered by R. solanacearum infection with a focus on the involvment of ethylene, we used the Medicago Affymetrix array to monitore the expression profiles and the molecular process associated with initial symptoms development (12hpi) and colonization (72hpi). In order to maximize chances to observe differential gene expression, RNA samples were extracted from the root infection zone (root tips) -Three Medicago truncatula lines, A17, F83005.5 and sickle were inoculated with GMI1000 Ralstonai solanacearum strain (107 cfu/ml). RNA were extracted from root extremities (1 cm above the root tip) at time 0, 12h and 72h post inoculation. Three biological repeats were conducted normal vs disease comparison, time course, 27 arrays - Medicago
Project description:Plants show a remarkable plasticity to adapt their root architecture to biotic and abiotic constraints of the soil environment. Although some of these modifications are fine-tuned by miRNAs, there are still shadow zones in these regulations. In the model legume Medicago truncatula, we analyzed the small RNA (smRNA) transcriptome of roots submitted to symbiotic and pathogenic interactions. Mapping on the genome and prediction of pre-miRNA hairpins allowed the identification of 416 candidates. Out of them, we found known and novel variants of 77 miRNA families, already reported in miRBase. In addition, thanks stringent criteria of miRNA prediction, 53 mtr-miRNAs were discovered, including 27 putative miRtrons. Exploring polymorphism in 26 M. truncatula ecotypes, higher polymorphism was observed in conserved rather than legume-specific miRNA genes. An average of 19 targets, mainly involved in environmental responses and signaling, was predicted per novel miRNA. In addition, taking advantage of our large number of smRNA libraries, we identified sets of miRNAs responsive to root pathogens or to symbiotic interactions and the related Nod and myc-LCO signals. 23 libraries of small RNA (smRNA) of roots submitted to symbiotic and pathogenic interactions.
Project description:affy_ralstonia_peeters_medicago - We have identified two essential virulence determinants (GALA7, a type III secretion effector and HpaP, a chaperone-like protein) of Ralstonia solanacearum for the infection and colonisation of the host plant Medicago truncatula. The scope of this project is to identify the GALA7 and HpaP-specific transcriptome alteration. For this purpose wild type and mutant infected root material (13h and 72h postinfection) will be analysed on M. truncatula affymetrix chips. Medicago truncatula (A17 line) are grown in vitro on Farheus medium (with Nitrogen source) plantlets are inoculated with water R. solanacearum wt, gala7 and hpap mutants, and root tips are collected at 13h and 72h postinoculation. Experiment was performed 3 times independently. 4 bacteria conditions x 2 harvest times x 3 biological repeats = 24 samples Keywords: gene knock out,normal vs disease comparison,time course,treated vs untreated comparison 24 arrays - Medicago
Project description:A comparative assessment between both technologies, RNASeq and microarrays to detect differential expression in Arabidopsis transcriptome. The sequencing approach use High-throughput sequencing on different Solexa technologies (GAII,HiSeq2000 multiplex or not) Wild type samples were analyzed from 2 tissus (flower buds and leaves) which have a very contrasted transcriptomic profile (i.e very high number of genes differentially expressed). The RNA was extracted from 2 tissus Flower Buds and Leaves from Arabidopsis. The associated GEO series with array part is GSE45345
Project description:The involvement of ROS in the legume – Rhizobium symbiotic interaction has been highlighted (Santos et al., 2001; Rubio et al., 2004). This interaction is characterized by the formation of a new organ on the root, the nodule and by the penetration, in parallel, of the bacteria into the root tissue via an infection thread (IT) (Parniske and Downie, 2003; Gage, 2004). H2O2 production has been shown in ITs during the Medicago – Sinorhizobium meliloti interaction (Santos et al., 2001). Moreover, S. meliloti mutants impaired in H2O2 detoxification mechanism possess in planta symbiotic phenotypes. As H2O2 production is known to orchestrate plant gene expression, our goal is focused on identifying the H2O2 regulated genes in the symbiotic process. We focus our analysis in the M. truncatula transcriptome as we are characterising the S. meliloti H2O2 transcriptome. -M. truncatula seedlings were grown in axenic condition on modified Fahraeus medium during seven days. Then they were transferred on new plates supplemented either with dimethyl sulfoxide (DMSO, mock treatement) or diphenylene iodonium (DPI, 10 µM). Twenty four hours later, they were inoculated either with water (mock inoculation; DMSO-H2O and DPI-H2O) or wild type S. meliloti 2011 strain (DMSO-INOC and DPI-INOC). Roots without apexes were harvested 48 hours after inoculation. After RNA extraction (Trizol), quality of the treatment was verified by RT-PCR (ENOD11: inoculation efficiency; GSHS1: DNA contamination; MTC27: constitutive) Keywords: treated vs untreated comparison 8 arrays - Medicago
Project description:This study compares the response of wild type (A17) and ethylene insensitive mutant (sickle) lines of the model legume Medicago truncatula to infection by the root-infecting necrotrophic fungal pathogen, Rhizoctonia solani AG8 (WAC10335). Two time points were taken, 2 and 7 days after inoculation along with corresponding mock-treated controls.
Project description:Nitrogen assimilation in plants is a tightly regulated process that integrates developmental and environmental signals. The legume-rhizobial symbiosis results in the formation of a specialized organ called root nodule, where the rhizobia fixes atmospheric nitrogen into ammonia. Ammonia is assimilated by the plant enzyme glutamine synthetase, which is specifically inhibited by PPT. The expression of key genes related to the regulation of root nodule metabolism will likely be affected by glutamine synthetase inhibition. We used microarrays to detail the global programme of gene expression in response to Glutamine synthetase inhibition in root nodules and identified genes differentially expressed over a time course. Medicago truncatula nodulated plants (20 days post inoculation) were treated with 0.25 mM of PPT. Root nodules were harvested at 4, 8 and 24 hours after PPT application. As a control, root nodules collected just before PPT application were used (PPT 0h). Three biological replicates consisting of pools of root nodules harvested from five distinct plants were used for RNA extraction and hybridization on Affymetrix GeneChips.
Project description:Despite their different origin and function, both pollen tubes and root hairs share the same sort of apical growth mechanism, i.e., the spatially focused cell expansion at the very apex. Ion fluxes, membrane trafficking, the actin cytoskeleton and their interconnection via signaling networks have been identified as fundamental processes underlying this kind of growth. Several molecules involved in apical growth have been identified, but the genetic basis is far from being fully characterized. We have used Affymetrix Arabidopsis ATH1 GeneChips to obtain the expression profiles of isolated Arabidopsis root hairs. A comparison with the expression profile of flow-sorted pollen grains reveals an overlap in the expression of 4989 genes, which corresponds to 42% of the root hair transcriptome and 76% of the pollen transcriptome, respectively. Our comparison with transcriptional profiles of vegetative tissues by principal component analysis and hierarchical clustering shows a clear separation of these samples comprised of cell types with diffuse growth from the two cell types with apical growth. 277 genes are enriched and 49 selectively expressed, respectively, in root hairs and pollen. From this set of genes emerges an apical growth signature containing novel candidate genes for apical growth determination. Root hairs were isolated from Arabidopsis seedlings and total RNA was isolated for expression profiling on Affymetrix ATH1 arrays. The study was performed with biological duplicates.
Project description:affy_aphanomyces_medicago - NFP is one of the putative Nod factor receptor and plays therefore a key role in the establishment of symbiosis. nfp allelic mutants are more susceptible to Aphanomyces euteiches (Ae), a root pathogen, than the A17 wild type (WT) line. In this study we want to compare the early plant responses, 1 day post inoculation (dpi) between WT and the nfp2 mutant, in order to identify NFP-dependent gene networks during infection.-Fifteen-day-old A17 or nfp-2 plants were grown in vitro on M medium. The entire roots were then inoculated (or not = controls) with a A. euteiches zoospores (105/ml) and harvested one day after inoculation. Three independent repeats were performed. Keywords: normal vs ems mutant comparison,near isogenic lines (hif) Overall design: 12 arrays - Medicago
Project description:affy_ralstonia_medicago - Ralstonia solanacearum is the causal agent of the devastating bacterial wilt disease. Its infection process was studied with an in vitro inoculation procedure on intact roots of Medicago truncatula. The pathosystem involved susceptible A17 and resistant F83005.5 M truncatula lines infected with the pathogenic strain GMI1000. The mutant A17 line, Sickle, which showed a resistant phenotype was also part of the experiment. To identify host signaling pathway triggered by R. solanacearum infection with a focus on the involvment of ethylene, we used the Medicago Affymetrix array to monitore the expression profiles and the molecular process associated with initial symptoms development (12hpi) and colonization (72hpi). In order to maximize chances to observe differential gene expression, RNA samples were extracted from the root infection zone (root tips) -Three Medicago truncatula lines, A17, F83005.5 and sickle were inoculated with GMI1000 Ralstonai solanacearum strain (107 cfu/ml). RNA were extracted from root extremities (1 cm above the root tip) at time 0, 12h and 72h post inoculation. Three biological repeats were conducted Overall design: normal vs disease comparison, time course, 27 arrays - Medicago