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Biosensor-Mediated Adaptive Laboratory Evolution of Escherichia coli for L-SerineOverproduction from Glycerol

In this study, adaptive laboratory evolution (ALE) combined with biosensor were employed to improving L-serine yield. First, serine-biosensor was constructed in E.coli based on the transcriptional regulator NCgl0581 of C. glutamicum, furthermore, the validity and sensitivity of the serinebiosensor was studied, and the results showed that serine-biosensor pDser from C. glutamicum was effective in E. coli and only cellular L-serine biosynthesized was monitored by serine-biosensor. Then E. coli 4W capable of producing 1.1 g/L L-serine from glycerol was used as a starting strain, and L-serine degradation pathway to glycine of 4W was deleted by CRISPR/ Cas9, resulting in strain 4WG, with L-serine titer of 2.01 g/L. 4WG was further evolved by using ALE combined with serine-biosensor, the evolved strain 4WGX was achieved and showed a yield of 0.41 g/g glycerol, and could produce 4.13 g/L L-serine, which was 105% and 275% higher than that of 4WG and 4W respectively, in addition, 4WGX showed better growth in the medium with 50 g/L L-serine addition, indicating its better L-serine tolerance. This work indicates that the serine-biosensor from C. glutamicum was useful in selecting serine over-producing E. coli, and this expanded the application of biosensor, and provided the more strategies for screening high performance strain.