A novel genetic switch with an optimal ON/OFF ratio to preserve growth performance prior to Escherichia coli autolysis for enhanced plasmid release

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When the biotechnology workhorse organism, Escherichia coli (E. coli) is used to propagate commercially valuable plasmids, such as those demanded by the globally expanding cell and gene therapy sector, efficient recovery of plasmids from the E. coli cell is key. However, this recovery of the desired plasmid product can prove challenging, with requirements for cell disruption and plasmid isolation from a complex process stream at scale.The aim of this study is to apply a novel genetic switch, with an optimal ON/OFF ratio to control E.coli autolysis such that normal growth performance is maintained whilst allowing simpler less shear induced separation. Several technologies have been reported in which E. coli autolysis is triggered to improve liberation of recombinant protein or plasmid DNA products. However, to date these E. coli strains consistently exhibit compromised growth, and therefore yield, performance due to 'leakiness' of the genetic trigger mechanism that controls initiation of autolytic events. This technology if successful will provide a key approach in reducing the number of steps in DNA manufacture, making the process more cost effective and sustainable.