Fig. 3

Construction and growth studies of the chassis strain S. coelicolor A3(2)-2023. (A) The overall strategy for the construction of the chassis strain S. coelicolor A3(2)-2023 using a double-crossover approach and Cre-mediated site-specific recombination. The targeted endogenous red BGC was replaced with vox-lox71-apra-lox66-vox2261 by a double crossover. Sites lox71 and lox66 were recombined by Cre to eliminate the apramycin resistance gene. S. coelicolor A3(2)-2023 was constructed with five RMCE sites (vox-vox2261, rox-rox2232, lox5171-lox2272, FRT-F3, and attB-attB15) sequentially inserted into the chromosome of S. coelicolor A3(2). (B) Construction of the knockout plasmid pKC1139-red-vox-apra-vox2261. The resulting plasmid was used for red BGC knockout and vox-vox2261 site insertion on the S. coelicolor A3(2) chromosome. (C) Growth curves of S. coelicolor A3(2) and S. coelicolor A3(2)-2023 in TSB liquid medium. (D) Growth and sporulation of S. coelicolor A3(2) and S. coelicolor A3(2)-2023 on MS agar medium. The color of the MS agar medium with S. coelicolor A3(2) was initially red due to the production of prodiginines (red complex) and then turned to blue upon the production of actinorhodin (blue product)