Synthesis of RpoS Is Dependent on a Putative Enhancer Binding Protein Rrp2 in Borrelia burgdorferi
Figure 3
Gene expression in B. burgdorferi strain OY173.
(A) Construction of an IPTG-inducible rrp2-FLAG expression shuttle plasmid. The plasmid pRrp2-FLAG pRrp2 was introduced into strain OY01 (rrp2[G239C]), yielding OY173. SDS-PAGE (B), immunoblot (C, D), and qRT-PCR analyses (E) were performed to analyze gene expression. In (B) and (C), spirochetes grown in BSK-II medium containing varying concentrations of IPTG were harvested when bacterial growth reached early stationary phase (∼108 cells per ml). In (D) and (E), spirochetes were grown in BSK-II medium. When bacterial growth reached mid-log phase (∼107 cells per ml), various amounts of IPTG were added into culture. Cells were collected at 9 h post-induction. In (B) and (D), concentrations of IPTG are indicated above the image. The arrow indicates OspC in (B). Specific antibodies, denoted as α- used in the immunoblot (C, D), are indicated on the left. In (E), the bars represent the mean measurements ± standard deviation. The mean values between induced groups (100-, 200-, or 500 µM IPTG) and the uninduced group (0 µM IPTG) were compared using the Student’s t test and are significantly different (p<0.05). For data normalization, the B. burgdorferi flaB gene was used as an internal control.