Fig. 2: Engineering violacein production into Panamanian golden frog bacterial isolates.
a Skin isolates were tested for resistance to carbenicilin (A, 100 μg/mL), chloramphenicol (C, 35 μg/mL), kanamycin (50 μg/mL), spectinomycin (S, 100 μg/mL), and tetracycline (T, 10 μg/mL). Strains were categorized as either resistant (red) or susceptible (pink) to the antibiotic (“Materials and methods”). Combinations that were not tested are colored black. b Fluorescence of Diaphorobacter 63F strains with (+) or without (−) plasmid pJAB582 (BHR lacIQ PA1lacO-1-sfGFP oriT aphA3). IPTG (1 mM) was added to induce expression of gfp from pJAB582. The bar graph shows the average of three replicates, dots represent individual data points, and error bars are the s.d. c Violacein biosynthetic pathway. d Plasmid containing violacein pathway library. Ribosome binding sites were randomized using degenerate primers (“Materials and methods”). e Image of 63F strains grown on R2A media for 72 h at 23 °C. f HPLC-UV profiles of a violacein standard (top), 63F strain carrying plasmid pJAB631 (BHR PA1lacO-1-vioABDCE oriT aphA3) (middle), and engineered Diaphorobacter 63F:vio strain containing PA1lacO-1-vioDABCE integrated into the genome (bottom). The identity of the peaks was determined by LC-MS. g Placement of the violacein pathway in the Diaphorobacter 63F chromosome. h Doubling times of wild type (gray) and engineered (purple) 63F strains. The bar graph shows the average of three replicates, dots represent individual data points, and error bars are the s.d.
