Fig. 2.
Optimization of taxadiene production through regulating the expression of the upstream and downstream modular pathways. (A) Response in taxadiene accumulation to changes in upstream pathway strengths for constant values of the downstream pathway. (B) Dependence of taxadiene on the downstream pathway for constant levels of upstream pathway strength. (C) Taxadiene response from strains (17 to 24) engineered with high upstream pathway overexpressions (6 to 100 a.u.) at two different downstream expressions (31 a.u. and 61 a.u.). (D) Modulation of a chromosomally integrated upstream pathway by using increasing promoter strength at two different downstream expressions (31 a.u. and 61 a.u.). (E) Genotypes of the 32 strain constructs whose taxadiene phenotype is shown in Fig. 2, A to D. E, E. coli K12MG1655 ΔrecAΔendA; EDE3, E. coli K12MG1655 ΔrecAΔendA with DE3 T7 RNA polymerase gene in the chromosome; MEP, dxs-idi-ispDF operon; GT, GPPS-TS operon; TG, TS-GPPS operon; Ch1, 1 copy in chromosome; Trc, Trc promoter; T5, T5 promoter; T7, T7 promoter; p5, pSC101 plasmid; p10, p15A plasmid; and p20, pBR322 plasmid.