Machine Learning Guided Batched Design of a Bacterial Ribosome Binding Site

[Image: see text] Optimization of gene expression levels is an essential part of the organism design process. Fine control of this process can be achieved by engineering transcription and translation control elements, including the ribosome binding site (RBS). Unfortunately, the design of specific genetic parts remains challenging because of the lack of reliable design methods. To address this problem, we have created a machine learning guided Design–Build–Test–Learn (DBTL) cycle for the experimental design of bacterial RBSs to demonstrate how small genetic parts can be reliably designed using relatively small, high-quality data sets. We used Gaussian Process Regression for the Learn phase of the cycle and the Upper Confidence Bound multiarmed bandit algorithm for the Design of genetic variants to be tested in vivo. We have integrated these machine learning algorithms with laboratory automation and high-throughput processes for reliable data generation. Notably, by Testing a total of 450 RBS variants in four DBTL cycles, we have experimentally validated RBSs with high translation initiation rates equaling or exceeding our benchmark RBS by up to 34%. Overall, our results show that machine learning is a powerful tool for designing RBSs, and they pave the way toward more complicated genetic devices.