Transcriptomics

Dataset Information

20

Interaction with Human Bronchial Epithelial Cells in vitro Affects Gene Expression in Moraxella catarrhalis


ABSTRACT: In this study, DNA microarray technology was used to measure global gene expression in M. catarrhalis cells that had attached to human bronchial epithelial cells in culture. Overall design: M. catarrhalis ATCC 43617 was allowed to attach (MOI~30) to a monolayer of 16HBE14 human bronchial epithelial cells for 2 hr in OmniTray cell culture plates. Additional portions of these bacteria were incubated in tissue culture medium in 75-cm2 cell culture flasks in the absence of 16HBE14 cells for 2 hr. After this incubation period, the medium in the flask with the 16HBE14 monolayer was decanted and replaced with tissue culture medium containing 25 mM sodium azide (to protect RNA). The 16HBE14 monolayer and attached bacteria were released from the plastic by scraping. After low-speed centrifugation to collect these bronchial cells and attached bacteria, the cell pellet was re-suspended in TE containing 1% saponin (which lyses the human cells). After a 10-min incubation at 37°C, the bacterial cells were collected by centrifugation. The bacteria growing in the flask without human cells were treated identically and then harvested by centrifugation. Total RNA was extracted from both sets of bacterial cells by using the Qiagen RNeasy Midi kit. DNA microarray analysis, slide scanning, and statistical analysis of the data were performed. RNA was isolated four times and used in five DNA microarray hybridizations; “dye swap” experiments were performed twice. Two additional independent experiments were performed to isolate two sets of RNA samples for qRT-PCR analysis to validate DNA microarray data.

INSTRUMENT(S): EJH Mcat Oligo-DNA Microarray 3

ORGANISM(S): Moraxella catarrhalis  

SUBMITTER: Eric J Hansen  

PROVIDER: GSE12990 | GEO |

SECONDARY ACCESSION(S): PRJNA112599

REPOSITORIES: GEO

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