Expanding the Diversity of Reporters for Use in Multiplexed Cell-Free Protein Synthesis Systems for Paper-Based Sensing Platforms

Cell-free protein synthesis (CFPS) systems are emerging as an attractive new avenue to bring developments in synthetic biology closer to real-world application. In particular, the preservation of cell-free systems until reactivation by hydration at the point of need offers new capabilities, especially in remote sensing applications. One interesting advantage of this approach is the potential to multiplex the sensing of several analytes by including multiple DNA molecules that each produce a unique visual signal. At this stage, however, work has primarily utilized Green Fluorescent Protein (GFP) or β-galactosidase (LacZ) as readouts. Here we describe efforts to expand the diversity of available reporters for use in multiplexing paper-based cell-free systems, including a panel of colorimetric enzyme reporters to enable a palette of readouts, discernible by the naked eye.