Team:Toronto/Safety

Would the materials used in your project and your final prduct pose safety risks?

Our project this year consist of engineering DH5a E.coli to produce magnetite crystals in the periplasm and using these crystals to engineer a novel gene expression system. Our team worked exclusively on E.Coli in a BL1 lab. Both groups adhered to the safety protocols outlined by University of Toronto.

When studying the formation of magnetite under synthetic conditions great care was taken to minimize accidents. A new purification protocol was designed to minimize the use of highly concentrated acids and all purification protocols involving highly corrosive acids were limited to the fume hood and disposed of according to protocols outlined by the University of Toronto.

In terms of biological hazards the team has been trained in Biosafety protocols by the University and extra care was taken to minimize possible release of our bacterium to the environment by immediately changing PPE when contaminated. PPE such as lab coats and gloves are worn at all times when handling and disposing of bacterial cultures and colonies.

Also due to the fact our bacterium is designed to be used in the lab we have determined that our constructs with the exception of the repressor proteins would not be active due to the lack of stimuli from IPTG or aTc.

We believe that our final project will have a high impact on future iGEM's in providing a novel way of sensing magnetic fields and allowing bacterium to react to them. While the bacterium is safe and innocuous it is a very versatile due to its foundational nature as a novel way of activating genes. This can be misused by a malicious individual. The bacterium's function as a mechanism to form nano-particles of magnetite however is difficult to use maliciously as currently it requires stringent conditions.

Do the new BioBrick parts that you made this year raise safety issues?

The only part which may raise a possible safety concern is our use of a fusion protein containing part of the ToxR protein. The ToxR protein regulates the production of Cholera toxin in Vibrio Cholerae. In our project we have designed a fusion protein which contains the the transmembrane and cytoplasmic domain of the ToxR protein fused to MMS6 which binds to magnetite. In order to ensure that the ToxR protein was safe to use we blasted the genome of E.coli for 3 repeats of GATTTTT which is the minimal sequence to which the ToxR cytoplasmic domain would bind. In our searches we were unable to determine known genes which contained 3 upstream GATTTTT repeats. Therefore there should be no expression of a native E.coli gene. In addition we blasted the NCBI database to see if another other organisms would contain the CTX promoter in case our protein was horizontally transferred to another organism. Many of the hits received were from cellular organisms which are far removed from E.coli or where from organisms associated with Cholerae such as a Cholerae phage.

Given that the cytoplasmic domains of ToxR need to dimerize inorder to function and the fact that magnetite is not formed in the periplasm of Vibrio Cholerae, we feel that there is minimal chance of a horizontal transfer of genetic material causing a threat to the environment by creating a virulent strain of Vibrio Cholerae. This will be documented on our Parts once we submit this part to the registry.

What biosafety provisions do you operate under?

The iGEM Toronto team operates under the biosafety rules of the University of Toronto. Which adhere to the 3rd edition of the Laboratory Biosafety Guidelines set out by the Public Health Agency of Canada. These provisions can be found at the following links.

http://www.ehs.utoronto.ca/services/biosafety.htm or http://www.phac-aspc.gc.ca/publicat/lbg-ldmbl-04/index-eng.php

In addition, our team have all undertaken a laboratory safety workshop offered by the Faculty Medicine which trains all new staff and students on lab safety.

Does your institution have a Biosafety committee or equivalent group?

Yes The biosafety division of the Office of the Environmental Health and Safety is responsible for biosafety at the University of Toronto. We spoke with a senior biosafety officer, who was enthusiastic about the applications of the project. He explained that the main hazards of the project were proper handling and disposal of iron oxide nanoparticles, and he also directed us to a research group in the Department of Civil Engineering that is undertaking a similar project and needs to dispose of similar materials. From these leads, we have learned the appropriate measures to facilitate safe lab protocols, avoiding accidental harm.

Does your country have a national biosafety regulations or guidelines

Yes, Canada does have national biosafety regulations and guidelines. They can be found at the following link

http://www.phac-aspc.gc.ca/publicat/lbg-ldmbl-04/index-eng.php