Additional file 3.
Expression of FLAG-tagged secretion candidates by C. burnetii transformants to confirm secretion.C. burnetii transformed with plasmids encoding FLAG-tagged secretion candidates were cultured for 48 h, then expression of tagged protein induced by addition of aTc for 24 h. Supernatants were harvested, TCA precipitated and analyzed by immunoblotting using antibody directed against the FLAG-tag. Supernatants of samples that were positive for secretion were then probed using antibody directed against EF-Ts to rule out cell lysis as a source of protein present in supernatants. Whole cell lysate of C. burnetii expressing FLAG-tagged CBU1764a was used as a positive control (+ve). To confirm that proteins not present in supernatants were expressed by C. burnetii transformants, lysates of bacterial pellets were probed with antibody directed against the FLAG-tag.
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Stead et al. BMC Microbiology 2013 13:222 doi:10.1186/1471-2180-13-222