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: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: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:In this work, 454 pyrosequencing was used to build up a 3’ 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: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 organism’s 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 cell’s 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.
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:Petunia is an excellent model system, especially for genetic, physiological and molecular studies. Thus far, however, genome-wide expression analysis has been rarely applied because of the lack of sequence information. We applied next-generation sequencing to generate, through de novo read assembly, a large catalogue of transcripts for Petunia axillaris and Petunia inflata. On the basis of the transcriptome of each species, comprehensive microarray chips for gene expression analysis were established and used for the analysis of global- and organ-specific gene expression in both species. In addition, microarray analysis was applied to explore the molecular basis of the seed coat defects in Petunia hybrida mutants, homozygous for a null allele of the AN11 gene, encoding a WDR transcription regulator. Among the transcripts differentially expressed in an11 seeds compared to wild type, many expected targets of AN11 were found but also several interesting new candidates that might play a role in morphogenesis of the seed coat. Our results validate the combination of next-generation sequencing with microarray analyses strategies to identify the transcriptome of two petunia species without previous knowledge of their genome, and to develop comprehensive chips as useful tools for the analysis of gene expression in P. axillaris, P. inflata and P. hybrida. The manuscript describes the creation by next generation sequencing of a large catalogue of the transcriptome of the two Petunia species, that are considered to represent the natural material from which the breeders selected their varieties. This submission represents the transcriptome component of study. The high throughput sequencing data were submitted to SRA (accession numbers: SRA027293, SRP004866.1, SRX036999.2, SRX036998.2).
Project description:To obtain insights about the roles of VvMYB5a and VvMYB5b, here we perform complementation analyses using petunia regulatory mutants impaired in pigment accumulation in flower epidermis, proven to be a valid tool for gene functional studies. We created three transgenic petunia lines overexpressing VvMYB5a, VvMYB5b and VvMYBA1 and we compared petal transcriptomes of each overexpressors with the untransformed one.
Project description:Solanum torvum Sw is worldwide employed as rootstock for eggplant cultivation because of its vigour and resistance/tolerance to the most serious soil-borne diseasesas bacterial, fungal wilts and root-knot nematodes. A 30,0000 features custom combimatrix chip was designed and microarray hybridizations were conducted for both control and 14 dpi (day post inoculation) with Meloidogyne incognita-infected roots samples. We also tested the chip with samples from the phylogenetically-related nematode-susceptible eggplant species Solanum melongena.The genes identified from S. torvum catalogue, bearing high homology to knownnematode resistance genes, were further investigated in view of their potential role in the nematode resistance mechanism. total RNA was extracted from control and 14 days post-infection (infection with root-knot nematode Meloidogyne incognita) from roots of Solanum torvum and Solanum melongena. Three biological replicates were used for each condition and genotype for a total of 12 samples.
Project description:We have used cDNA-AFLP and microarray analyses to profile the response of the tomato meiotic anther transcriptome to moderate heat stress condition in Moneymaker or Falcorosso and HeatSet1 tomato genotype. Combimatrix microarray technology were applied to obtain a general overview of the gene expression in meiotic anthers of a tolerant and a sensitive tomato genotype. We analyzed three time points (0,2,and 6h) for each genotype. For each sample we performed three biological replicates.