Targeted Transcriptional Activation Using a CRISPR-Associated Transposon System

Synthetic perturbation of gene expression is central to our ability to reliably uncover genotype–phenotype relationships in microbes. Here, we present a novel transcription activation strategy that uses theVibrio choleraeCRISPR-Associated Transposon (CAST) system to selectively insert promoter elements upstream of genes of interest. Through this strategy, we show robust activation of both recombinant and endogenous genes across theEscherichia colichromosome. We then demonstrate the precise tuning of expression levels by exchanging the promoter elements being inserted. Finally, we demonstrate that CAST activation can be used to synthetically induce ampicillin-resistant phenotypes inE. coli.