Technological watch

AbstractEscherichia coli Nissle 1917 (EcN) is a probiotic microbe that has the potential to be developed as a promising chassis for synthetic biology applications. However, the molecular tools and techniques for utilizing EcN remain to be further explored. To address this opportunity, we systematically expanded the EcN?based toolbox, enabling EcN as a powerful platform for more applications. First, two EcN cryptic plasmids and other compatible plasmids were genetically engineered to enrich the manipulable plasmid toolbox for multiple gene coexpression. Next, we developed two EcN?based technologies, including the conjugation strategy for DNA transfer, and quantification of protein expression capability. Finally, we further expanded the EcN?based applications by developing EcN native integrase?mediated genetic engineering and establishing an in vitro cell?free protein synthesis (CFPS) system. Overall, this study expanded the toolbox for manipulating and making full use of EcN as a commonly used probiotic chassis, providing several simplified, dependable, and predictable strategies for researchers working in synthetic biology fields.This article is protected by copyright. All rights reserved