Bacterial-MERFISH measurements of E. coli in various media and B. thetaiotaomicron in the mouse colon

Single-cell decisions made in complex environments underlie many bacterial phenomena. Image-based, transcriptomics approaches offer an avenue to study such behaviors, yet these approaches have been hindered by the massive density of bacterial mRNA. To overcome this challenge, we combine 1000-fold volumetric expansion with multiplexed error robust fluorescence in situ hybridization (MERFISH) to create bacterial-MERFISH. This method enables high-throughput, spatially resolved profiling of thousands of operons within individual bacteria. Using bacterial-MERFISH, we dissect the response of E. coli to carbon starvation, systematically map subcellular RNA organization, and chart the adaptation of a gut commensal, B. thetaiotaomicron, to micron-scale niches in the mammalian colon. This deposition contains raw data and final analysis structures associated with the benchmarking of bacterial-MERFISH in E. coli in LB medium, profiling of E. coli through a glucose-xylose diauxic shift, mapping of the internal organization of the E. coli transcriptome in LB, and measurements of the spatial-niche-adaptation of B. theta to the mouse colon.