Characterization of mycobacterial oligopeptide permease system

ABSTRACT: M. tuberculosis strain 1254 wild type (WT) and its isogenic delta Rv3662c-Rv3665c KO (opp KO) and complemented opp::opp were cultured in vitro, in liquid 7H9 media supplemented with OADC and 0.05% Tween 80, 0.5% glycerol, in a 150 mL volume, contained in roller bottles, incubated at 37oC. Growth was monitored as OD600 nm and recorded on a daily basis. In all experiments, the starting OD600 nm was 0.04-0.05. Aliquots were removed at each OD600 nm to make serial dilutions, plate onto 7H10 OADC agar plates for CFU counts. When cells reached OD600 nm of 0.3, 1.0, and 1.5, a 10-30 mL aliquot was taken, centrifuged 3 min at 3000 g, at room temperature, and the supernatant was discarded, with cell pellets being immediately frozen by inclusion into dry ice. Frozen cell pellets were stored at -80oC for later RNA isolation. In all experiments, RNA from WT was labeled with Cy5, and RNA from the opp KO was labeled with Cy3. In all experiments, the microarray comparsion used RNA from cells of the same growth stage, i.e., WT OD 0.3 vs opp KO OD 0.3, WT OD 1.0 vs opp KO OD 1.0, and WT OD 1.5 vs opp KO OD 1.5. When comparing the Complemented opp::opp vs the mutant opp KO, RNA from the later strain was Cy3-labeled. A cell type comparison design experiment design type compares cells of different type for example different cell lines.

ORGANISM(S): Mycobacterium tuberculosis

PROVIDER: GSE18426 | GEO | 2009/10/07

SECONDARY ACCESSION(S): PRJNA118223

REPOSITORIES: GEO