|Application:||Gene expression microarray analysis|
|Number of samples:||6|
|Release date:||Sep 25 2013|
|Last update date:||Sep 25 2013|
|Dataset link||Differential expression of Dictyostelium discoideum AX2 upon infection with wt L. pneumophila JR32 vs. uninfected 48h p.i. (timecourse experiment)|
The bacterial strain used in this study was L. Pneumophila Philadelphia I JR32. The strain was grown on buffered charcoal yeast extract agar (BCYE) at 37°C with 5% CO2 atmosphere for 3 days. The D. discoideum wild-type strain AX2 was grown at 23°C in 75 cm2 cell-culture flasks with 10 ml HL5 medium. For infection, Dictyostelium cells were harvested, resuspended in a 1:1 solution of HL5 medium and Soerensen buffer. Fifteen millilitres of a 1×10e6 cells/ml suspension were seeded into a 75 cme2 cell culture flask and the amoebae were inoculated with 10e7 bacteria/ml. After different time intervals of incubation (1, 3, 6, 24, and 48 h) the RNA was isolated from 1.5×10e7 Dictyostelium cells. Usually two or three parallel cultures for the experiment and the control were inoculated per infection. The percentage of infected cells was determined by in situ hybridization with Legionella-specific 16S rRNA probes. The average from three independent determinations was 34, 42, 42, 57 and 82% of infected Dictyostelium cells after 1, 3, 6, 24 and 48 h, respectively.