Project description:C57BL/6 mice were maintained in specific pathogen-free conditions in accordance with the Institutional Animal Care and Use Committee. Listeria monocytogenes (clinical isolate 10403s) was from V. Boyartchuk (NTNU, Trondheim, Norway) and infected as previously described (PMID 24477914) for 24 hours before harvesting the spleen. PBS was used as control. . Single cell suspensions from spleens were used to make total RNA. 4 μg of total RNA was used to generate libraries for RNA-sequencing (Illumina unstranded mRNA kits). Samples were sized, quantified and validated on a Bioanalyzer. Libraries were sequenced on a High-Seq System (Illumina 2000, San Diego, CA) as paired-end 100 reads. Sequence reads were aligned to the mouse genome (assembly NCBI m37/mm9) using TopHat.Gene level read counts based on the resulting alignments were calculated using HTSeq. The DESeq R package was used to normalize gene counts, calculate fold change in gene expression, estimate p-values and adjusted p-values for change in gene expression values, and to perform a variance stabilized transformation on read counts.
Project description:Several Toll-like receptors are activated by Listeria monocytogenes infection, resulting in the activation of MyD88 dependent signaling pathway. However, the negative role of MyD88 in gene expresson is unclear. To address this, we performed microarray analysis of mRNAs from WT or MyD88-/- peritoneal macrophages infected with Listeria monocytogenes.
Project description:Listeria monocytogenes cells (strain LI0521) were digested with trypsin for the identification of surface proteins. The supernatant was filter-sterilized and subjected to identification by LC-MS/MS. Concurrently secreted or shed proteins were identified by isolating filter-sterilized supernatants following incubation of L. monocytogenes cells in buffer without trypsin. This was followed by trypsin digest of the sterilized supernatant and identification by LC-MS/MS.
Project description:Time course study of the mouse infection by comparing the genomic transcriptional patterns of Listeria monocytogenes EGDe grown under laboratory conditions (exponential growth phase) with that of in vivo-grown bacteria (in mouse spleens) over three days of infection. Time course study of the mouse infection by comparing the genomic transcriptional patterns of Listeria monocytogenes EGDe grown under laboratory conditions (exponential growth phase) with that of in vivo-grown bacteria (in mouse spleens) over three days of infection.
Project description:Time course study of the mouse infection by comparing the genomic transcriptional patterns of Listeria monocytogenes EGDe grown under laboratory conditions (exponential growth phase) with that of in vivo-grown bacteria (in mouse spleens) over three days of infection.
Project description:These studies were designed to examine the transcription of Listeria monocytogenes strains 10403S and LO28 during intracellular replication in mammalian macrophages. Duplicate WT Listeria monocytogenes (strains 10403S and LO28) were used to infect mouse bone marrow-derived macrophages (BMMs). Bacterial RNA was harvested at 4 hours post-infection.