We collaborated with the Boston University Hardware team. We followed the protocols they provided us and assembled three types of chips: PCR, digestion and a toehold chip they designed specifically for our project. To test out functionality of the chips we used color dye to imitate the process of biological fluid flowing into the chip. The toehold chip had a relatively serious leakage problem, and air bubbles in the valves blocked the way for color dye to get through. We hypothesize several reasons that explained why the leakage happened and redid some steps to improve the assembly. After several rounds of tests and trials with all the chips, we gave BU Hardware team some feedback about our thoughts of the chips and how it could potentially be improved. We also pointed out steps on the protocols that appeared unclear to us as users and suggested them to explain purposes of certain steps better to make sure users understand why they are important.
MIT Collaboration: Adding an MS2 Domain to Triggers
We collaborated with MIT to test the potential for their RNA binding protein (MS2) to sterically protect our RNA trigger from exonuclease degradation. For the MS2 protein to be able to recognize our trigger DNA, a hairpin loop structure was added at one end. After DNA is transcribed into RNA, the MS2 protein binds to the hairpin loop on the RNA sequence. The below is a flowchart showing the procedures of collaboration. MIT designed the hairpin loop that was added to the trigger sequences we provided them. We eventually put MS2 protein, trigger and toehold sequence into cell-free. However, due to potential contamination of trigger RNA, we were not able to get valuable cell-free data.