Safe Project Design
The project is particularly suited for easy containment and safe handling.
- Thermococcus kodakarensis is nonpathogenic.
- Thermococcus kodakarensis is an obligate anaerobe, so it is easily contained under standard conditions
- Limonene is a nontoxic chemical, and is used in food products
- Thermococcus kodakarensis can only grow above 80°C, so it cannot spread under standard conditions.
Safe Lab Work
The lab that we grew Thermococcus kodakarensis in has several anaerobic chambers, which we cleaned with ethanol every time we worked in it. We also decontaminated all equipment that was brought into the chambers, and most commonly used equipment used inside the chamber stay inside of it at all times.
Our organism, Thermococcus kodakarensis, is an obligate anaerobe that will die when taken out of the anaerobic chambers. Due to its required growth conditions, working with this archaeon has very low risks to environmental and personal safety. There are no known ethical risks. PPE was worn as needed in the BSL-1 laboratory. All of the reagents and nutrients used in the culture were disposed of properly. For our chassis and transformations, we used E. coli K12 derivatives, XL1 blue cells, and Stellar cells.
Regarding equipment safety, precautions were taken with the hydrogen and nitrogen tanks used to fill the anaerobic chambers. The tanks were secured at all times as well as regularly inspected for safety. The anaerobic chambers are equipped with electronic gas leak sensors, so any malfunctions with the machinery can be stopped with these safety features. The Hydrogen and nitrogen tanks were tied and weighed down during the tank changes.
The lab contained radioactive materials. The team did not use these materials as part of the project, and took an online safety course that taught how to be a safe ancillary user.
The PIs are Dr. Thomas Santangelo and Dr. Christie Peebles, and the safety officer is Claudia Gentry-Weeks. Before entry to the laboratory, every student was required to perform online certifications in biosafety lab work, hazardous waste generation, Responsible Conduct of Research, and being an ancillary user of radiological materials. The team also received a workshop on microbiological methods and biosafety including autoclave safety, proper use of PPE, sterile technique, spill cleanup, and disposal of biohazardous waste. Three separate laboratory settings were utilized for the project, and specific safety protocols were followed in each.
Weekly laboratory safety inspections of hazardous waste satellite accumulation areas were performed by the lab members, and the team successfully complied with the required items. The Colorado State University Biosafety Office under the Vice President for Research Office provided oversight for biosafety at our institution. The guidelines are from CSU's BSL1 section in CSU's biosafety handbook.