Deployment of Sensors with Cell-Free Systems

Developments in synthetic biology have demonstrated an ever-increasing capability to manipulate and repurpose cellular function, thus removing the living cell from the equation. Synthetic gene circuits can harness cell-free transcription and translation machinery to respond to any stimuli nature or adversaries can create. Cell-free protein synthesis reactions can maintain functionality after being dried, stored, and rehydrated. Given this, we aim to develop and provide Warfighters a detection technology that is wide-ranging, simple, cheap, and that will not release genetically modified organisms. Here we describe developments in this area by evaluating a range of material matrices for our cell-free reactions. We tested a commercial cell-free transcription/translation product called PURExpress (NEB), as well as a cell lysate we produced at the CCDC Chemical Biological Center. We have successfully utilized colorimetric, fluorescent, and luciferase-based reporters for easy visualization purposes. We ultimately intend to integrate this technology into a small handheld reader that images cell-free detection tickets and processes results in real time. A variety of materials were tested to serve as reaction matrices, including glass microfibers, cellulose-based papers, and synthetic polymer materials. Testing was conducted by placing disks of material in 384-well plates, acoustically dispersing cell-free reaction components onto the disks, and lyophilizing. Reactions were then re-hydrated, placed in a scanner, and time- course images were collected. Image analysis software developed at CCDC CBC was utilized to determine the speed of reaction .