Figure 2

Gradual establishment of efficient D-xylose utilization during the mutagenesis and multiple evolutionary engineering steps. (a) CO₂ production as measured by weight loss in sequential semi-anaerobic batch fermentations in YP + 40 g/L xylose at 35°C. After EMS mutagenesis and one step of genome shuffling, the culture was submitted to 11 serial transfers, in which each time part of the culture after the batch fermentation was used to start a new fermentation. The CO₂ production profile of the first 9 serial batch fermentations is shown. The concentration of xylose was increased to 50 g/L and 60 g/L in the 8th and 9th batch fermentation, respectively. (GS1 stands for the first step of genome shuffling, and the next number indicates the step in the serial transfer.) (b) Volumetric CO₂ production in semi-anaerobic batch fermentation in YP + 40 g/L xylose at 35°C by single cell isolates obtained from the 2nd (GS1.2), 4th (GS1.4), 7th (GS1.7) and 11th (GS1.11) serial batch fermentation during the evolutionary adaptation process. The horizontal bar represents the mean with the standard deviation. The parent strain, HDY.GUF5, and a previously constructed industrial D-xylose utilizing stain, TMB3400, [37] are shown for comparison.