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. 2021 Aug 9;203(17):e00210-21. doi: 10.1128/JB.00210-21

FIG 5.

FIG 5

Analysis of abortive transcripts from the carD promoter. (A) carD promoter sequence from −36 to +50 from the 122-nt linear fragment (−72 to +50) used in the abortive transcription reactions. The −35, −10, and extended −10 elements (with positions matching E. coli consensus sequence shown in bold), position T18, and the transcription start site (+1) are all indicated. The runoff (full-length [FL]) transcript is 50 nt. (B) Image of 20% acrylamide-urea gel showing 5′ radiolabeled abortive and productive transcripts produced from the carD promoter fragment with RNAP (120 nM), with or without CarD (4,800 nM) in transcription buffer with 50 mM NaCl. The RNA size markers (lane 5), the positions of the abortive RNAs, and the full-length transcript (FL) are indicated. (C) Amount of full-length (50-nt) transcript without CarD relative to with CarD is plotted. (D) Amount of 6-mer abortive RNA from the carD promoter without CarD relative to with CarD is plotted. (E) Productive yield (FL transcript as a percentage of total product, abortive plus FL) without CarD or with CarD is plotted. The percentage yields are average values, and the error bars represent standard deviations determined from 5 assays. (F) Abortive yield-to-productive yield ratio without CarD (cyan) or with CarD (gray). (G) Abortive probabilities at positions in the ITS of the carD promoter. The abortive probabilities for each position, calculated as described in Materials and Methods and in reference 55, are shown without CarD and with CarD. The 5-mer transcript was not detected in the absence of CarD, so no value is shown. The 5-mer value in the presence of CarD and the 6-mer value in the absence of CarD are the abortive yields for those positions since abortive probabilities cannot be calculated without values for a 4-mer and 5-mer, respectively. Error bars represent mean and standard deviations from 5 assays. P values for the difference in abortive probability in the presence versus the absence of CarD for each of the following ITS positions from R-7 through R-11: R-7 (residue 7), 0.011; R-8, 0.002; R-9, <0.001; R-10, <0.001; and R-11, 0.002.