Dual UTR-A novel 5′ untranslated region design for synthetic biology applications

Bacterial 5′ untranslated regions of mRNA (UTR) involve in a complex regulation of gene expression; however, the exact sequence features contributing to gene regulation are not yet fully understood. In this study, we report the design of a novel 5′ UTR, dual UTR, utilizing the transcriptional and translational characteristics of 5′ UTRs in a single expression cassette. The dual UTR consists of two 5′ UTRs, each separately leading to either increase in transcription or translation of the reporter, that are separated by a spacer region, enabling de novo translation initiation. We rationally create dual UTRs with a wide range of expression profiles and demonstrate the functionality of the novel design concept in Escherichia coli and Pseudomonas putida using different promoter systems and coding sequences. Overall, we demonstrate the application potential of dual UTR design concept in various synthetic biology applications ranging from fine-tuning of gene expression to maximization of protein production.