Fig 1.
Schematic of the biosynthesis pathway of AP-3 and a brief workflow.
Table 1.
Primers used in this study.
Fig 2.
HPLC analysis of AP-3 production.
(A) AP-3 standard curve. (B) Products of the fermentation sample. (C) AP-3 standard. Red arrows indicate the AP-3 position in the graphs.
Fig 3.
Generation of the asm25-inactivated strain.
(A) Schematic diagram of plasmid construction. Gel electrophoresis of pWHU2653 harboring asm25 sgRNA (both lanes). Plasmids were purified from E. coli. (C) Gel electrophoresis of homologous arms of asm25. Lanes 1 and 2 indicate arm R, Lanes 3 and 4 indicate arm L, and Lane 5 indicates arm LR fusion. (D) Gel electrophoresis of selected double-crossover exconjugantes. The primers asm25-seq-F and asm25-seq-R were used for PCR.
Fig 4.
Determination of AP-3 yield in asm25 knockout and WT strains by HPLC.
Student’s t test was used for statistical analysis. *: p<0.05. The mean ± SD from three independent experiments (n = 3) are presented. Blue line: WT strain; Red line: asm25 knockout strain.
Fig 5.
Determination of AP-3 yield in the presence of isobutanol.
(A) AP-3 yield curve in the WT strain. (B) AP-3 yield curve in the asm25 knockout strain. Lines in color represent various concentrations of isobutanol in fermentation. The mean ± SD from three independent experiments (n = 3) are presented. The comparison and statistical analysis for AP-3 yield at Day 9 among groups were performed by the ANOVA. **: p<0.01; ***: p<0.001.