Analysis of BBD18 repression of an <i>rpoS</i> promoter <i>lacZ_Bb</i> transcriptional fusion.

<p>(A) A schematic diagram of the transcriptional fusion of the <i>rpoS</i> promoter and 5’ untranslated region (UTR) fused directly to the <i>lacZ</i>_Bb open reading frame (ORF). The position of the <i>rpoS</i> transcriptional start site <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0093141#pone.0093141-Lybecker1" target="_blank">[58]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0093141#pone.0093141-Burtnick1" target="_blank">[63]</a> is indicated by a filled arrowhead. The Shine-Dalgarno sequence (RBS), and the translational start site of β-galactosidase (<i>lacZ_Bb</i>), indicated by the ATG, are also shown. Regions corresponding to the <i>rpoS</i> promoter and 5’ UTR (141bp) or the <i>lacZ</i>_Bb ORF are indicated with brackets. The "G" in the RBS marked with an asterisk is identified because the reporter construct harbored an A->G mutation relative to the published sequence at that position. Cell lysates from strains B31-S9 (wt), B31-S9/pBSV2G-<i>rpoS</i>p<i>₁₄₁</i>-<i>lacZ_Bb</i> (wt/<i>rpoS</i>p-<i>lacZ_Bb</i>), and B31-S9/pBSV2G- <i>rpoS</i>p<i>₁₄₁</i>-<i>lacZ_Bb</i>/pBSV28-<i>flaB</i>p-<i>bbd18</i> (wt/<i>rpoS</i>p-<i>lacZ_Bb</i>/<i>flaB</i>p-<i>bbd18</i>) were grown under <i>rpoS</i>-inducing conditions and analyzed with OspC antisera (B) or stained with Coomassie blue (C) to demonstrate equivalent protein loads in each lane. The positions of molecular mass standards are shown on the left in kiloDaltons (kDa). (D) β-galactosidase activity in cell lysates from strains B31-S9 (wt), B31-S9/pBSV2G-<i>rpoS</i>p<i>₁₄₁</i>-<i>lacZ_Bb</i> (wt/<i>rpoS</i>p-<i>lacZ_Bb</i>), and B31-S9/pBSV2G- <i>rpoS</i>p<i>₁₄₁</i>-<i>lacZ_Bb</i>/pBSV28-<i>flaB</i>p-<i>bbd18</i> (wt/<i>rpoS</i>p-<i>lacZ_Bb</i>/<i>flaB</i>p-<i>bbd18</i>) grown under <i>rpoS</i>-inducing conditions.