Implementing shallow screening strategies to accelerate microbial strain improvement

The ability to move rapidly through the Design-Generate-Test-Learn (DGTL) cycle is essential for effective forward engineering of microbial phenotypes. The Onyx^® platform is an automated CRISPR-based genome editing technology that enables rapid diversity generation. To optimize the DGTL cycle, effective diversity generation must be paired with an appropriate screening strategy for isolated genetic variants. Here we discuss the benefits of creating a higher edit diversity in cell populations while using a shallow screening approach with high-throughput and automated phenotyping systems such as Rapid Fire Mass Spec and plate-based fluorometric assays. We demonstrate the effectiveness of this approach using both metabolic and strain engineering examples, including a 14,000-fold improvement in lysine biosynthesis in E. coli and 4‑fold increase in heterologous protein production in S. cerevisiae.