Team:HK GTC/2019.igem.org/Team:HK GTC/Safety

Safety

Safe project design

Our goal of the project is to design the mutation sites and to generate purified proteins of PETase mutants. Their efficiencies in degradation are tested via enzyme activities for pNP-aliphatic esters. Our products will be used in a closed system to prevent leakage of enzyme into the environment. Our approach also serves as a safe alternative to prevent the release of engineered bacteria into the wild.

Safe organisms

We choose safe strains of E.coli called DH5a and BL21(DE3) derivative C41(DE3) as a chassis. The strain C41(DE3) has at least one mutation that allow the expression of toxic proteins, as PETase is toxic to E. coli, using it is favourable. Both E.coli strains have no harm on the natural environment and humans.

Safety lab

To protect from exposure to health and safety hazard, we wear personal protective equipment in the lab all the times. This includes safety goggles, lab coat, disposable gloves and covered shoes. We are working in a university lab and we are accompanied and supervised at all times by our guiding teacher, lab technicians and lab professor.

We chemically transformed plasmid containing gene encoding mutated forms of PETase into DH5a for gene verification and then C41(DE3) for protein expression.

We use Midori Green stain directly to molten agarose during the gel pouring step to visualize DNA fragments as a safe alternative to ethidium bromide.

We utilized antiseptic techniques and used Bunsen burner to maintain sterility of the workplace and experimental materials.