Safety
General Safety
Our lab at TAS is classified as Biosafety Level 1, which means the lab uses microbes that pose little to no risk to healthy individuals. Hazardous chemicals and solutions are used at a minimal level and in well ventilated areas such as fume hoods in the TAS research lab. For instance, we do not use EtBr for gel electrophoresis. Instead, we use a safer nucleic acid stain called Seeing Safe DNA Dye. Nevertheless, all chemicals and solutions are still treated with all due respect, care, and caution. MSDS are stored on our lab computers and in a folder on the lab technician's desk.
For the vast majority of experiments, we work with a safe and common lab strain, Escherichia coli (E. coli) K-12 DH5alpha, and follow safety rules set by our lab instructor. We have our own biosafety committee, which consists of three research teachers Dr. Jonathan Hsu, Mr. Jude Clapper, and Dr. Nicholas Ward. They oversee proper work area conditions by checking on disposal of Petri dishes and liquid wastes, sanitation, and teaching proper laboratory techniques. Our guidelines, taken from National Yang-Ming University's Center of Environmental Protection and Safety, cover many safety rules and procedures ranging from lab specific rules to behavior. For example, we prohibit food, open-toed shoes and drinks in the lab. We also have a thorough clean up procedure. We also have a thorough clean up procedure. For example, all bacterial liquid wastes are bleached and used tips are autoclaved before disposal. Other lab wastes are carefully packaged and then sent to a disposal facility to ensure no hazardous materials are mismanaged.
In Taiwan, there are many laws and regulations regarding biosafety in research labs. These can be found in the National Yang-Ming University's Center of Environmental Protection and Safety and Health page.
Project-Specific Biosafety
Synthetic Viral RNA Targets
Instead of using actual viruses, which are dangerous and restricted without exception, we used synthetic viral RNA targets. The purpose of using a synthetic RNA target is to simply mimic the presence of a virus to see if our assay suggests that we can diagnose viral diseases.
We selected a 36 nucleotide gene fragment from the genes of our viruses of interest. The detailed process of the selection of these genes are outlined in the experimental section and the specific sequences are listed below.
As these synthetic viral RNA targets do not have the full genetic information necessary to formulate any protein of the virus, nor do they have the sequence for any other essential proteins to function like a virus, papers have cited the use of these types of sequences as synthetic, non-viral, and non-replicating (Corman et al., 2020; Hamidi & Ghourchian, 2015; Schott et al., 2016). It is also notable that our assay does not provide any enzymes that will amplify the target sequence. Amplification takes place by synthesizing DNA through a standard and safe padlock probe template (Hamidi et al., 2015). Thus, these sequences are considered safe and have been checked in and approved by the iGEM Safety Committee.
| Virus name |
Sequence |
| SARS-CoV-2 (β-coronavirus) |
5'-AUAUUUCAACUGAAAUCUAUCAGGCCGGUAGCACAC-3' |
| Influenza Type A (H1N1pdm09) |
5'-UAGCUCCUCAUAAUUGAUGAAAUCUCCUGGGUAACA-3' |
| Influenza Type B (Victoria Lineage) |
5'-CAGAGGGUAUUUUCCCUGUGCAUUUUGGUCUGCCCA-3' |
Table 1: Synthetic Viral RNA Targets for SARS-CoV-2, Influenza A (H1N1pdm09), Influenza B (Victoria Lineage)
Diagnostic Results & Hardware Disposal
If this project was to become a consumer product such as a testing kit, the risks would include the effects of possible false positives and false negatives. For example, one who experiences a false positive diagnosis might undergo treatment for a virus they actually don't have, posing possible harm to the person. One who experiences a false negative diagnosis may be free to wander back into the human community, where the individual could infect others. To minimize those risks, we plan to increase the sensitivity and specificity of our test, as per FDA test kit guidelines researched, to ensure accurate diagnosis. We will also advise possible individuals tested to also confirm their results through another round of testing, whether that is with our kit or through another methodology. We hope these possible methods would reduce the harmful impacts of false diagnosis on not only the individual, but the community.
As the used test kit could be unsafe due to sample collection of possibly infected individuals, the manual will come with instructions regarding how to correctly throw away the device after use. There are three steps to which customers should follow with the disposal of the used device. The customers should first put the device in a disinfecting solution for three hours, wash the device thoroughly with soap and water, and lastly microwave the device to ensure safety (Disinfection Technology and Strategies for COVID-19 Hospital and Bio-Medical Waste Management, n.d.) Since no elements in the test kit is biologically hazardous, the package label does not need to follow CDC’s guideline of biologically hazardous objects (CDC LC Quick Learn: Recognize the Four Biosafety Levels , n.d.).
References
Corman, V. M., Landt, O., Kaiser, M., Molenkamp, R., Meijer, A., Chu, D. K., Bleicker, T., Brünink, S., Schneider, J., Schmidt, M. L., Mulders, D. G., Haagmans, B. L., van der Veer, B., van den Brink, S., Wijsman, L., Goderski, G., Romette, J.-L., Ellis, J., Zambon, M., … Drosten, C. (2020). Detection of 2019 novel coronavirus (2019-nCoV) by real-time RT-PCR.Eurosurveillance,25(3). https://doi.org/10.2807/1560-7917.ES.2020.25.3.2000045
Hamidi, S. V., & Ghourchian, H. (2015). Colorimetric monitoring of rolling circle amplification for detection of H5N1 influenza virus using metal indicator. Biosensors and Bioelectronics, 72, 121–126. https://doi.org/10.1016/j.bios.2015.04.078
Hamidi, S. V., Ghourchian, H., & Tavoosidana, G. (2015). Real-time detection of H 5 N 1 influenza virus through hyperbranched rolling circle amplification. The Analyst, 140(5), 1502–1509. https://doi.org/10.1039/C4AN01954G
Schott, J. W., Morgan, M., Galla, M., & Schambach, A. (2016). Viral and Synthetic RNA Vector Technologies and Applications. Molecular Therapy, 24(9), 1513–1527. https://doi.org/10.1038/mt.2016.143
