Biosafety

For people working with genetic modified organisms, having biosafety as an important focus point is a must. This is because when working with organisms so small, that even the naked eye can’t see them it can still have a big impact on the researchers health, and in an even worse scenario, it can potentially affect the environment and the public safety in a negative way.
Because of this we have taken the biosafety questions very serious, and made a point of identifying all the dangerous situations that could occur in the laboratory.
This involves taking necessary precautions, like only working with nonpathogenic bacteria[i].
We have been working in a class 1 laboratory and have followed all the safety rules needed to work in such an environment. In doing this we have also abided the local Danish laws[ii] on biosafety and read the specific guidelines for our institute.
Another thing we found important concerning biosafety, is if our biobrick could give the bacteria pathogenic abilities. We have tried our best to prevent it from giving the bacteria any advantages, compared to naturally occurring bacteria, which could pose an environmental problem.
We also had two extra safety measures in the planning phase:
The first safety measure we planned was a L-ramnose Kill-switch. This would be a safety measure in the completed product. The consumer would get a small bag of L-ramnose, (a sugar that the body can’t digest) with the product, that they could consume some time after drinking our yogurt with inulin producing bacteria.
This would kill any of our constructed bacteria left in the gut and leave the naturally occurring bacteria unharmed, while not being toxic to humans.
Our other safety is a response to the way our construct will work, we need to have our genes in two plasmids, which will let the bacteria synthesize the inulin. Because of the two different plasmids we are able to make a paired double toxin/anti-toxin kill-switch (see the figure below). This will prevent some of the mutation that can happen in the gene, and at the same time prevent horizontal gene transfer.
The way the kill-switch is going to work is by inserting a toxin and a non corresponding anti-toxin in both of our genes, so that the toxin and its corresponding anti-toxin are on different plasmids.
This will hopefully prevent horizontal gene transfer by killing the new host and also prevent frameshift mutations from happening in the new host.
This will of course require that the anti-toxin is downstream from the toxin so that it won’t be able to lose the toxin without losing the anti-toxin.

Would any of your project ideas raise safety issues in terms of researcher safety, public safety, or environmental safety?

We are working in a class 1 laboratory following all the lab rules. We are using nonpathogenic bacteria[i] and we are planning on adding different kinds of kill-switches.
Another thing we see as safety preclusion is that the biobrick we are adding is a disadvantage for the bacteria, the biobrick makes the bacteria produce inulin out of its food source which slows down the grow rate of the bacteria giving it a disadvantage competing with other bacteria in the wild.

Do any of the new BioBrick parts (or devices) that you made this year raise any safety issues?

We don’t see any safety issues in any of our biobricks. In our planning phase though, we have come up with an idea on how to prevent frameshift mutations from occurring in the plasmids and how to prevent horizontal gene transfer.
This will be solved through a paired, double toxin/anti-toxin kill-switch.

What does your local biosafety group think about your project? And is there any local laws?

Our local biosafety group is called “Arbejdsmiljøgruppen”, they are responsible for safety in the laboratory, both biological and work related. We talked with a representative from the group about our project, and she did not find any problems with our project, as long as we followed the safety rules of a class 1 laboratory.
We also looked into the local laws and found that the local law we had to follow is “Bekendtgørelsen om Genteknologi og Arbejdsmiljø[ii]” (eng. The Order on Gene-technology and Working Environment) of 2008, which follows the rules laid down by the European Union in 1990 in the ”Directive on the Contained Use of Genetically Modified Micro-organisms”.

Do you have any other ideas how to deal with safety issues that could be useful for future iGEM competitions?
How could parts, devices and systems be made even safer through biosafety engineering?

One of the safety measurements we are using it the paired, double toxin/anti-toxin kill-switch which we think should be a part of coming iGEM teams safety precautions.

Apendix

Bacteria: E. coli Strains: TOP10 and XL gold
[i] link til apendix
[ii]link til https://www.retsinformation.dk/Forms/R0710.aspx?id=12325)