| 초록 |
In this work, an integration helper plasmid was designed for rapid gene manipulation in E. coli. This plasmid (pCW611) expresses two recombinases (Red and Cre) under separate inducible promoters (PlacUV5 and PBAD). Thus, iterative plasmid transformation and curing is avoided, resulting in significant time savings compared to the traditional, two-plasmid approach. This enabled target gene deletion in 3 days. The system was verified by deleting adhE, sfcA, frdABCD, and ackA individualy and pairwise (adhE-aspA and sfcA-aspA). Finally, a fumaric acid producing E. coil strain was developed by deleting four genes (fumB, iclR, fumA, and fumC) in 10 days as a proof-of-concept. [This work was supported by the Technology Development Program to Solve Climate Changes on Systems Metabolic Engineering for Biorefineries from the Ministry of Science, ICT and Future Planning (MSIP) through the National Research Foundation (NRF) of Korea (NRF-2012-C1AAA001-2012M1A2A2026556). MG was additionally supported by the Swedish research council Formas]. |
