Generally, Escherichia coli TOP10 cells are used for cloning. They are derivatives of the non-pathogenic K-12 laboratory strains. In our lab, DH5alpha is used. The detailed information of the E. coli mutations can be found in E.coli wiki. The likelihood of a human getting ill from working with these bacteria is therefore low. The most probable route of transmission would occur by accidental aerosol formation or ingestion. The Environmental Protection Agency states that the K-12 strain (which both DH5-Alpha and JM109 are derivatives of) is poorly retained in the human gut, so the chance of it becoming pathogenic by mutations is low[1]. Mainly for these reasons, this organism is classified as Bio-Safety Level 1.

Aspergillus niger N593, and 872.11 are all derivatives of N400, which is the sequenced CBS strain. They are non-pathogenic industrial workhorses. Additionally, the N593 and 872.11 have the pyrA auxotrophy marker. (see Fungi-related safety).

Public safety

In case of the accidental release of bacteria containing our BioBrick system, the physical conditions outside the lab are harsh to the E. coli strain chassis that are used, so the bacteria would be unable to grow and the DNA of the BioBrick system will not be widely. By means of natural genetic transformation the DNA of the BioBrick system could be taken up by other bacteria.

Additionally, as expression of the plasmid would incur a significant metabolic burden on a (probably non-pathogenic) soil bacterium, and do little to increase its fitness, it is considered unlikely that the plasmid would be propagated. Unfortunately it is not possible to know in advance what the actual effect of a natural transformation would be, but the odds seem to be in favour of it to cause little impact on the public and environment.

Environmental safety

There are several scenarios in which unintentional release of genetically modified material could take place. Labelling of the lab equipment and glassware used is necessary to prevent loss and improper waste disposal by a fellow researcher. The following problems are not that easy to prevent:

The air filtering system will have a hard time in keeping aerosols in the lab when a window gets broken. If this happens through a thunderstorm, an electricity break-down is not unlikely and would increase the chance of release furthermore. Under Good Microbial Practice, though, the formation of aerosols is prevented as much as possible. Thus in total, the probability is relatively low.

All waste containing genetically modified organisms is required to be sterilized by autoclaving before disposal. Direct actions should be taken if it is discovered that an autoclave has been malfunctioning after the waste is discarded outside of the lab. To reduce the risk on environmental contamination it is necessary to check upon the autoclave’s functionality (by monitoring its operational temperature). If the aforementioned hazards do occur, they should be reported to the Minister of ‘Housing, Spatial Planning and the Environment’ and involved institutions to make the hazards undone as soon as possible.

The usage of antibiotic resistance markers increases the chance of spreading antibiotic resistance to pathogens. By conjugation, transduction or natural genetic transformation, DNA can be transferred between bacteria. There is a chance the antibiotic resistance genes end up in a pathogenic bacterium which is not intrinsically resistant to antibiotics. The chance that these genes persist is low, since there is no direct evolutionary benefit for micro-organisms living around the lab to take up the extraneous genes. The plasmids used in our lab can bear resistance markers for Ampicillin, Chloramphenicol and Kanamycin. These are all resistance markers that are used in BSL-1 laboratories on a standard basis.

Risk and benefits

Generally, under Good Microbiological Practices the risk of working with the BioBrick system is rather small to the researchers. Despite the involved BioBrick system, the cell chassis in our project is considered to be safe to researchers, publics and environment. Even if the system was to be released into the surroundings, there is a low risk that it would enter a pathogenic host organism. However, there might be some risks involved if a person or organization has malicious intents by using this system as producers for toxins. In this case they would be able to use it to produce some secondary metabolites which have bad influences.

We believe, though, that every new tool that is made could be used for malicious purposes, but by no means does this mean they shouldn’t be made. In our case, we think that the possible benefits of our system and the likelihood of it being used for those purposes outweigh the risk of it being used for evil purposes. Based on these considerations, we conclude that the potential benefits in successfully executing our project outweighs the estimated risks.