Figure 1. SYBR-green based qPCR analysis shows that transcription limits heterologous production of oxytetracycline in E. coli.

(a) The 32 kb oxytetracycline biosynthetic gene cluster is shown. Five putative operons, oxyABCDE, oxyIHGF, oxyJKLMNO, oxyRQP, and oxyST are predicted for this gene cluster (b) qPCR analysis shows that over-expression of the alternative sigma factors σ⁵⁴, σ^S and FecI enable detectable levels of the oxyB transcript to be produced. Over-expression of no sigma factor, σ^E, σ^F and σ^H do not lead to detectable levels of the oxyB transcript. (c) qPCR analysis shows that over-expression of the alternative sigma factor σ⁵⁴ lead to detectable levels of transcripts for all five putative operons in the oxytetracycline biosynthetic pathway. In the absence of σ⁵⁴ over-expression, the oxyB transcript cannot be detected. See also Table S1. No oxyB, oxyF, oxyK, oxyP, oxyT transcripts were observed in the null strains which lacked the oxytetracycline gene cluster. (Figures S1, S2).