My long-term research goal is to develop a general method to rapidly and easily monitor small molecules relevant to the study of biology and the environment. A research objective of my research lab is to engineer riboswitch-based biosensors to monitor intracellular concentrations of small organic molecules. Riboswitches are naturally occurring non-coding RNA elements that act as direct sensors of diverse small molecules and can signal molecular recognition through altered protein expression. The rationale for this project is that the demonstrated plasticity of riboswitches coupled with the utility of genetic selections can form the basis of a powerful method for engineering whole-cell biosensors for small molecules. Indeed, through this project, a naturally occurring riboswitch for an important bacterial signaling molecule, cyclic diguanylate (c-di-GMP), was used to create a c-di-GMP sensor. When applied to the water-borne pathogen, Vibrio cholerae, the sensor was able to identify specific extracellular signals that affect intracellular c-di-GMP levels and likely contribute to the persistence of the bacteria in aquatic reservoirs. To demonstrate the full range and potential of riboswitch-based sensors, a novel riboswitch was engineered through in vivo directed evolution (Figure 2). This de novo development of a riboswitch is transformative as it provides a general platform from which many more synthetic riboswitches may be discovered, addressing a major obstacle to broadening the scope of riboswitch-based technology.