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Fig. 3 | Microbial Cell Factories

Fig. 3

From: Aadh2p: an Arxula adeninivorans alcohol dehydrogenase involved in the first step of the 1-butanol degradation pathway

Fig. 3

Physical maps of the YRC102-(6H)-AADH2 and YIC102-(6H)-AADH2 used for transformation of A. adeninivorans G1212 (a) and time-course traces of ADH activity by transgenic A. adeninivorans strains G1212/YRC102 and G1212/YRC102-AADH2 grown in various media (be). a Both cassettes contain the selection marker module with ATRP1m gene fused to the ALEU2 promoter and the expression module with TEF1 promoter—(6H)-AADH2 gene—PHO5 terminator. In addition YRCs (AscI fragments) are flanked by 25S rDNA sequences for targeting, whereas YICs (SbfI fragments) contain only the selection marker and expression modules. Transformants G1212/YRC102 (b, d) and G1212/YRC102-AADH2 (c, e) were cultured in shake-flasks for 96 h in YMM-glucose-NaNO3 at 30 °C (b, c) and YEPD at 30 °C (d, e). At the indicated times, 2 mL aliquots of the culture were used to determine biomass (dcw in g/L—filled square, dashed line) and to assay the intracellular Aadhp activity (U/L culture—filled circle, solid line) using butyraldehyde as substrate, and to calculate the Aadhp output Y (P/X) (U/g dcw—filled triangle, dotted line). For the determination of the dcw, 2 mL yeast culture was centrifuged and the pellet was washed with 1 mL water. The pellet was lyophilized and the tube with dried cells was weighted. Measurements were done in triplicate

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