Our team had the benefit of being able to work in a BSL-1 lab for the course of our project.
In our lab, we ensure that whenever we are handling bacterial cultures we will wear our personal protective equipment. This includes our gloves, labcoat and safety glasses when appropriate. The protocols that we are performing are routine synthetic biology protocols and do not pose any heightened risks. Our laboratory also utilizes CITI safety training; the following modules cover our work: Basic Introduction to Biosafety, Conflicts of Interest, Hazard Communication, Initial Biosafety Training, Lab Chemical Safety, and OSHA Bloodborne Pathogens. This extensive training is designed to provide team members with safety training relevant to their project and additional training relevant to general lab research more broadly. All training was completed before students worked in the lab.
We used a standard E.coli strain, MG1655, requiring BSL1 standard precautions. Our sensor was tested with manganese chloride solutions in the low mM range, and posed minimal risk. We engineered a sensor plasmid which incorporates a manganese-responsive promoter, riboswitch and sfGFP into a pSB3K3 backbone to sense Manganese (II) in water samples. The sensor plasmid was transformed into E.coli strain MG1655 for initial testing. A second plasmid with IPTG-inducible manganese responsive transcription factor mntR will also be prepared to see if it improves the sensitivity of the sensor plasmid.We tested the function and specificity of our sensor plasmid (i.e. pSB3K3 with manganese-responsive promoter, riboswitch and sfGFP) by performing manganese chloride dose-response curves and specificity tests. Further, we co-expressed the sensor plasmid with an IPTG-inducible mntR plasmid to see if increased mntR (a manganese-responsive transcription factor) increases the sensitivity of the sensor.
Although chronic exposure to mM levels of manganese has been linked to health effects, basic safety precautions including gloves, lab coats, and eye protection will be used to minimize risk. The potential risks of accidental exposure are minimal but chronic consumption of manganese has been linked to cognitive defects. These effects did not effect the students working in the labroatory.