There is risk associated with almost any laboratory experiment, especially when working with live biological agents. However, our team has gone to great lengths to minimize the risk posed to the researcher, the public, and the environment. Overall, our project can be considered low risk since the finished product is not intended to come in contact with humans in any form. Our E. coli and Rhodobacter sphaeroides cultures should only be grown and tested in a bioreactor with proper laboratory safety technique. Even in the event that either strain was ingested, its is very likely that no harm would occur since the bacteria are not able to survive the environment of the human digestive system. The two major safety concerns associated with our project arise during DNA purification using gel electrophoresis and extraction of PCB from spirulina powder.
Gel electrophoresis is necessary to purify DNA for almost all steps of our project. To make DNA visible as it moves through the agarose gel, Ethidium Bromide (EtBr) is added to the gel to act as a fluorescent tag. Ethidium Bromide itself is a potent mutagen and a known carcinogen that may be absorbed through the skin. For this reason, a separate lab bench has been set aside for all items coming into contact with the toxin including pipets, tips, gel rigs, glassware, and DC power sources (for electrophoresis). In addition there is a vessel designated as contaminated where all disposables (i.e. pipet tips) maybe deposited and properly disposed of at a later date. It is important that all items coming in contact with EtBr remain on the EtBr lab bench and all noncontaminated lab materials are not brought to this bench. Nitrile gloves must be worn when handling EtBr and replaced frequently, especially when going back and forth from this bench to another one.
Phycocyanobilin (PCB) is necessary for the final testing and characterization of our strain of Rhodobacter sphaeroides and is most easily obtained by purifying it from spirulina, a cyanobacteria. There are two potentially harmful chemicals used in this process: methanol and Mercury (II) Chloride (HgCl2). Methanol is highly flammable and poisonous if ingested. Methanol can cause blindness if swallowed in even small quantities. HgCl2 is a highly toxic form of Mercury because it is in a soluble form and can accumulate in fatty tissues. All steps in PCB extraction must be performed on a designated lab bench in much the same way as gel electrophoresis (although separate from EtBr contamination). A fume hood should also be used with nitrile gloves to prevent human contact. Special decontamination vessels are needed that are large enough to hold all waste products given that there are several washings which produce a large volume of toxins. Mercury requires special disposal procedures that may vary depending on lab protocol.
The general public is at little to no risk from our experimentation since it is not intended for use outside of a bioreactor or for human trials in any way. Our labspace was shared with several classes so it was important that we clean up our lab when finished and that we properly label all chemicals and contamination areas. Environmental procedures were also followed with the disposal of all chemicals, glassware, and sharps. None of our live cultures were released into the environment and therefore would not affect natural populations of bacteria.
All research conducted at Washington University (medical school and main campus) is under the supervision of the Office of the Vice Chancellor for Research (OVCR). This office is responsible for all concerns pertaining to research and publications produced by the university. Among other things, our lab must comply with the Environmental Health and Safety Policy and Procedures laid out by the OVCR. To view the full list of Environmental Health and Safety Policies and Prcedures, please click [http://ehs.wustl.edu/new/safetycommittee.htm here]. Furthermore, every team member has completed an Annual Regulatory and Safety Review for Laboratory Personnel offered by the Environmental Health and Safety committee at Washington University. This review is offered annually and all of our team members have at least one year of lab experience working with live cultures
Our new BioBricks consist of composites of existing BioBricks and new ones derived from the Rhodobacter sphaeroides genome. None of these BioBricks are hazardous or require any special storage procedure beyond that of ordinary DNA fragments.
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