Project description:Cultures were grown at 28 oC for 48 h on a shaker at 250 rpm. MG132 (60 ï¾µM, final concentration) in phenylmethylsulfonyl fluoride (PMSF) was added after 46 h or PMSF was added alone to the cultures without MG132. Following sample collection, mycelia were washed with 0.9 % (w/v) NaCl in RNAse-free (diethyl pyrocarbonite [DEPC]-treated) water, frozen in liquid nitrogen and stored at -80 oC.
Project description:Iron (Fe) deficiency is a yield-limiting factor for a variety of field crops across the world and generally results from the interaction of limited soil Fe bioavailability and susceptible genotype cultivation. Tomato, a Strategy I, model plant for Fe deficiency, is an important economical crop. Tomato responses in order to improve Fe uptake are based on acidification of rhizosphere, reduction of Fe3+ to Fe2+ and transport of Fe2+ into the cells. Transcriptional profile obtained by roots (27-d) of 21-d-old tomato plants starved of iron for an additional week was compared with the transcriptional profile obtained for roots (27-d) of 21-d-old tomato plants grown for an additional week at 100 M-NM-<M Fe. Tomato plants were hydroponically grown in both cases. Three different biological replicates were used for each sample repeating the experiment three times. All samples were obtained pooling roots of six plants (27-d-old).
Project description:In dairy ruminants transcriptome profiling has enabled the identification of genes, pathways and regulatory networks activated in mammary tissues during experimental infection by various pathogens, including E. coli, S. aureus and S. uberis. Information in goats are still low and many host-pathogen interaction mechanisms have to be explained. The objectives of the present study were (1) to identify the network of genes that becomes activated in caprine blood and milk somatic cells in early response towards a S. aureus challenge in order to better understand the local and sistemic response and (2) to search any difference in this immune response by using two animal groups belonging to a caprine reference family established based on founders with adverse SCC breeding values. Udders from ten healthy French Alpine goats were infected with S. aureus and samples of blood and milk cells were collected at 0, 24 and 30 hours after infection. Alterations in the transcriptome profile were investigated using a custom bovine DNA microarray containing 43.822 unique gene probes.
Project description:A wide range of environmental stresses lead to an elevated production of reactive oxygen species (ROS) in plant cells thus resulting in oxidative stress. The biological nitrogen fixation in the legume - Rhizobium symbiosis is at high risk of damage from oxidative stress. Common bean (Phaseolus vulgaris) active nodules exposed to the herbicide Paraquat (1,1 '-Dimethyl-4, 4'-bipyridinium dichloride hydrate) that generates ROS accumulation, showed a reduced nitrogenase activity and ureide content. We analyzed the global gene response of stressed nodules using the Bean CombiMatrix Custom Array 90K, that includes probes from some 30,000 expressed sequence tags (EST). A total of 4,280 ESTs were differentially expressed in oxidative stressed bean nodules; of these 2,218 were repressed. These genes were grouped in 44 different biological processes as defined by Gene Onthology. Analysis with the PathExpress bioinformatic tool, adapted for bean, identified five significantly repressed metabolic path This work presents the transcriptional profile of bean nodules, induced by strain Rhizobium tropici CIAT 899, under oxidative stress, generated experimentally by adding the herbicide Paraquat (1,1 '-Dimethyl-4, 4'-bipyridinium dichloride hydrate) for 48 hours. We analyzed the transcript profile, via microarray hybridization, using the Bean CombiMatrix Custom Array 90K, that includes probes from some 30,000 expressed sequence tags (EST). A total of 4,280 ESTs were differentially expressed in oxidative stressed bean nodules; of these 2,218 were repressed.
Project description:In this study, we investigated the transcriptome of Bifidobacterium bifidum PRL2010 during in vitro growth by micro array technology. When B. bifidum PRL2010 was grown in liquid broth, 425 of the 1644 PRL2010 genes represented on the array were expressed in at least one of the three investigated growth phases, i.e., lag-, exponential and stationary phase. These transcriptional analyses identified a core in vitro transcriptome encompassing 150 genes, which resulted expressed in all phases. A proportion of the latter genes were further investigated as potential reference genes by Quantitative Real Time PCR (qRT-PCR) assays. Their expression stability was evaluated under different growth conditions, encompassing cultivation on different carbon sources, exposure to environmental stresses (thermal, acidic and osmotic) and growth phases. Our analyses validated six reference genes suitable for normalizing mRNA expression levels in qRT-PCR experiments applied to bifidobacteria. Transcriptional profiling of B.bifidum PRL2010 at different growth phases (lag phase, early exponential phase, late exponential phase, early stationary phase).
Project description:We studied the global transcription profiling of mouse upon colonization with Bifidobacterium bifidum PRL2010 by using DNA microarrays. Two groups of mice consisting each of two animals were orally inoculated with either 109 CFU of PRL2010 cells (test strain) or water (control). Animals were 3 months old female BALB/c mice. Bacterial colonization was established by five consecutive daily administrations using a micropipette tip placed immediately behind the incisors.
Project description:In this work, 454 pyrosequencing was used to build up a 3M-bM-^@M-^Y cDNA fragment library from a normalized library constructed from pooled RNA samples extracted at different stages of A. quisqualis mycoparasitization process (recognition, early and late parasitization). An extensive catalogue of unique transcripts was compiled and used to develop a microarray for large-scale analysis of genes involved in this mycoparasitism. We examined the transcriptomic changes that occur during the first stage of mycoparasitism (conidial germination). Our results showed that 1,776 transcripts are regulated during germination in the presence of powdery mildew. A striking feature of the gene catalogue was the presence of a number of genes predicted to encode proteins involved in the production of, glucanases, chitinases and extracellular proteases. This suggests that A. quisqualis causes the degradation of powdery mildew macromolecular constituents to provide the carbon skeletons and energy for the synthesis of proteins and other components destined for the developing of the mycelium. Microarray oligo probes were designed based on 454 sequencing of 3'-ends of transcripts of a sample constituted by pooling RNAs extracted at different stages of A. quisqualis mycoparasitization process (recognition, early and late parasitization)
Project description:The aim of this study was to compare the tomato global transcriptional profiles in response to host attack by ToMV and Fol in order to identify genomic differences and similarities in incompatible interactions between a foliar and a vascular pathogen. In order to identify a set of genes of interest in tomato plants infected with F. oxysporum f. sp. lycopersici (Fol) and Tomato Mosaic Virus (ToMV) a transcriptional analysis was performed. Tomato genes differentially expressed upon inoculation with Fol and ToMV were identified at 2 days post-inoculation, using an un-inoculated sample as reference.
Project description:We studied the global transcription profiling of human cell lines upon colonization with Bifidobacterium bifidum PRL2010 by using DNA microarrays. We decided to use HT29 monolayer as an in vitro model to investigate the molecular impact of B. bifidum PRL2010 on human intestinal transcriptome. HT29 monolayers cultivated at 15 days of post confluence were placed in contact with PRL2010 cells for a range of time spanning from 0 h, 1 h (T1), 2 h (T2) to 4 h (T4).
Project description:Bifidobacteria represents one of the dominant group of microorganisms colonizing the intestine of infants. However, the genetic determinants supporting the establishment and the interaction with the human hosts are still largely unknown. Most commensal bacteria interacting with eukaryotic hosts express adhesive molecules on their surfaces that modulate interaction with host cell receptors or with soluble macromolecules. Whole genome transcription profiling of B. bifidum PRL2010, a strain isolated from infant stool, under in vitro as well as in vivo conditions revealed the expression of few common extracellular proteins among which type 1 pili encoding genes. To investigate the molecular mechanisms sustaining the interaction of PRL2010 strain with the human gut, we first explored the global genome transcription profiling of this strain in a in vitro human model such as in the presence of HT29 cell lines. The transcriptome was analyzed using a custom B. bifidum PRL2010 array representing the 90% of this organismM-bM-^@M-^Ys protein coding genes. To better evaluate the conserved responses by B. bifidum, the in vivo transcriptomes were quantified against a diverse set of transcriptome patterns identified for in vitro laboratory cultures of the strain, i.e., B. bifidum responses after growth on an cellM-bM-^@M-^Ys monolayers growth medium (DMEM); B.bifidum responses after growth on synthetic medium (MRS). Briefly, we analized five conditions, two of which are also used as references. Every experiment was performed in duplicate and in vivo condition was performed in quadruplicate.