This project aims to develop the full versatility that paper-based sensors can have. This system uses the idea of a three-hybrid system, where one part binds the promoter, another part is a subunit of E. coli RNA Polymerase, and the analyte of choice provides the bridge that recruits the RNAP to the DNA. For this system, we chose two analytes: VEGF-A, which is a small dimeric protein involved in many cancers, and anti-MUC1 antibodies, which are present in many cancer patients. Both of these sensors use the same DNA binding domains and RNA Polymerase domain, inspired by a bacterial two-hybrid system developed in 2000. (Joung 2000) In fact, the only difference between the sensors are the proteins fused to these domains. In the VEGF-A sensor, we used a single chain variable fragment antibody, which is extremely specific to its target. (Chen 1999) In the anti-MUC1 antibody sensor, the bait is a portion of the MUC1 protein, as shown in the image below.
Project State
Currently, we are cloning the constructs using PCR tricks. Once we have the plasmids, it will be a simple matter to apply the 3-day process to create sensor extracts. Since we have worked out most of the kinks in the sensor extract protocol, we expect to get results within a month, which will be posted on this page.
