ABSTRACT: The differential response of the chicken trachea to chronic infection with virulent Mycoplasma gallisepticum strain Ap3AS and Vaxsafe MG (strain ts-304) – A transcriptional profile
Project description:To identify differential gene expression profiles of chicken tracheal epithelial cells (TECs) upon exposure to Mycoplasma gallisepticum virulent strain Rlow and avirulent strain Rhigh and corresponding lipid associated membrane proteins(LAMP) at 1.5 hours in vitro. Goal of this experiment was to identify relative comtribution of LAMPs in up-regulation of inflammatory gene compared to the live strains. Several genes were identified to be differentially regulated in all exposures, but the virulent strain up-regulated more number genes as well as at a higher extent. We identified 6 important inflammatory mediators and did confirmatory RT-qPCR analysis at 1.5, 6 and 24 hours in vitro as well as at 1.5 and 6 hours ex-vivo. RT-qPCR was also employed to identify expression of these 6 genes in presence of different signalling inhibitors and we were able to identify that Mycoplasma gallisepticum LAMPs up-regulate these inflammatory genes via TLR-2 in an NF-M-NM-:B dependent pathway. Primary chicken tracheal epithelial cells (TECs) were exposed to either 500 MOI of a virulent Mycoplasma gallisepticum strain Rlow or an avirulent strain Rhigh and the corresponding lipid associated membrane proteins (LAMPs) at 5M-BM-5g/mL for 1.5 hours. 4 biological replicates along with a dye swap technique totalling 8 replicates were utilized for all microarray experiments
Project description:To identify differential gene expression profiles of chicken tracheal epithelial cells (TECs) upon exposure to Mycoplasma gallisepticum virulent strain Rlow and avirulent strain Rhigh and corresponding lipid associated membrane proteins(LAMP) at 1.5 hours in vitro. Goal of this experiment was to identify relative comtribution of LAMPs in up-regulation of inflammatory gene compared to the live strains. Several genes were identified to be differentially regulated in all exposures, but the virulent strain up-regulated more number genes as well as at a higher extent. We identified 6 important inflammatory mediators and did confirmatory RT-qPCR analysis at 1.5, 6 and 24 hours in vitro as well as at 1.5 and 6 hours ex-vivo. RT-qPCR was also employed to identify expression of these 6 genes in presence of different signalling inhibitors and we were able to identify that Mycoplasma gallisepticum LAMPs up-regulate these inflammatory genes via TLR-2 in an NF-κB dependent pathway.
2014-09-19 | GSE61520 | GEO
Project description:Transcriptomic analysis of long-term protective immunity induced by vaccination with Mycoplasma gallisepticum strain ts-304
Project description:Genomic content of Vaccine strains were probed against the known sequence of the virulent strain Rlow of M. gallisepticum to identify divergent or absent genes in the attenuated strains. Genomic DNA was extracted from in vitro grown cultures of M. gallisepticum strains Rlow, F, ts-11, and 6/85 and three samples from each were selected for hybirdization to oligonucleotide microarrays using standard methods from the Bioprime CGH kit. All samples were labeled with Cy3 dye and scanned at a PMT gain of 700 using a GenePix 4000B scanner. Features are duplicated on the slides and data were averaged between duplicate features on each slide. Median signal intensities were averaged between samples of the same strain and each feature from each vaccine strain was compared to Rlow. Features exhibiting four-fold or less hybridization in the vaccine strains were considered divergent or absent.
Project description:Genomic content of Vaccine strains were probed against the known sequence of the virulent strain Rlow of M. gallisepticum to identify divergent or absent genes in the attenuated strains.
Project description:Mycoplasma gallisepticum transcriptome comparison between in vitro grown cultures of strains Rlow and F utilizing oligo DNA microarrays.
Project description:Mycoplasma gallisepticum transcriptome comparison between in vitro grown cultures of strains Rlow and F utilizing oligo DNA microarrays. Two-condition experiment, Rlow vs. F strain cells. Biological replicates: 3. 1 technical replicate per biological replicate which includes a dye swap.
Project description:Mycoplasma gallisepticum is a convenient model object for studying the regulation of transcription because it has a reduced genome, lack of cell wall and many metabolic pathways, and also easy to culture and non-pathogenic to humans. For rapid investigation of gene expression we developed microarray design including 3 366 probes for 678 genes. They included 665 protein coding sequences and 13 antisense RNAs from 816 genes and 17 ncRNAs present in Mycoplasma gallisepticum. This work was carried out transcriptomic profiling for different types of effects on the expression of genes of Mycoplasma gallisepticum: 1) genetic knock-out mutants; 2) cell culture exposed to sublethal concentrations of antibiotics; and 3) well-characterized heat stress effect. The study was performed on Agilent one-color microarray with custom design and random-T7 polymerase primer for cDNA synthesis. Using set of different probes for each gene or ncRNA allows to increase accuracy of gene expression quality.
