Team:DTU-Denmark/Safety considerations

Most E.coli strains are harmless and normally part of human gut flora. However, environmental strains such as serotype O157:H7 can cause serious food poisoning in humans. The genetic set up of the E.coli laboratory strain DH5α has been engineered in such a way, that it can be considered as a safe laboratory strain, if standard safety protocols are followed. Moreover E.coli is one of the most used laboratory strains and requires only a biosafety level 1 laboratory. This fact alone eliminates most safety issues with regard to laboratory applications like in our case.

The genetic characteristics of the DH5α strains are:

F- endA1 glnV44 thi-1 recA1 relA1 gyrA96 deoR nupG Φ80dlacZΔM15 Δ(lacZYA-argF)U169, hsdR17(rK- mK+), λ–

Amongst the most important genetic modifications is F-, which denotes the strain´s inability to develop F-pili. This prevents transfer of genetic information by horizontal gene transfer. Another important genetic modification is recA1, making the DH5α strain recombination deficient, preventing recombination of the plasmid into the E.coli genome so that the plasmid inserts are more stable. This leaves DH5α also very sensitive to UV-light as a result of the strains inability to undergo recombination, thereby impairing its DNA repair mechanism.

With regard to its prospective use as a biosensor within industrial, environmental, medical or food applications safety aspects have to be reconsidered and implemented in product development strategies. For example if our E.coli biosensor would be deployed as an oral drug it will be necessary to prevent the bacteria spreading to other parts of the human body or giving rise to an immune response. A possible solution to these problems could be to encapsulate the bacteria in some way or/and to limit the number of bacteria reproduction cycles.

Local biosafety group

We have a local biosafety group at our institution, and they have advised us to follow standard safety protocols for genetic engineering and molecular biology that are the standard practice at our institution. None of the BioBricks submitted in this project comprise any elevated danger of researcher safety, public safety, or environmental safety, if the standard practice is followed.

At the institute all equipment used for bacteria are cleaned with cetyl and autoclaved. This treatment kill the bacteria but the DNA might still be intact. In order for the DNA to be inactivated more autoclaves would be needed. Another way to inactivate DNA is by using chlorine. However, this raises an issue with regard to nature.

When working in a class II laboratory it is important to handle all waste carefully. This means that everything should be autoclaved.

The Department for systems biology have developed an safety protocol that we have all read, signed and followed during the duration of our project. This applies to the given laws and regulation.

• Woodcock, D.M. et al. (1989) Nucl. Acids Res., 17, 3469–3478.
• Raleigh, E.A., Lech, K. and Brent, R. (1989). In F.M. Ausebel et al. (Eds.), Current Protocols in Molecular Biology, (p. 1.4). New York: Publishing Associates and Wiley Interscience.
• http://en.wikipedia.org/wiki/Escherichia_coli