Overview

　　Our team is deeply convinced that all biological experiments carry some risk to the environment and experimenter. Under the guideline of IGEM Headquarter, we strictly follow a high standard of biosafety and responsible biological engineering. We also take note of the Laboratory biosafety manual published by WHO to ensure our lab work conducting in compliance with safety regulation. Moreover, before conducting experiments in the lab, all team members have undergone lab training by our PI and team members in NYMU Taipei 2016. The training included laboratory emergency response procedures, operation protocols for all kinds of machines, and biological waste decontamination and disposal protocols. As one of the members of the synthetic biology community, we are responsible for abiding by the common rule.

Biosafety

Organisms We Used

　　We used cyanobacteria Synechococcus sp. PCC 6803, cyanobacteria Synechococcus sp. PCC 7942 and E.coli DH5α in our project. We extracted NrtA gene from cyanobacteria Synechococcus sp. PCC 6803. Next, we transformed NrtA gene into E.coli. Finally, we co-cultured the engineered E.coli with microalgae to induce the nitrogen starvation. Nitrogen starvation enlarged the oil production of algae. After extracting oil from microalgae, we killed E.coli to prevent contamination and prevent the danger of spreading engineered E.coli. To deal with the safety of E.coli, we constructed a suicide mechanism.

Strain information of E.coli

　　According to the Globally Harmonized System (GHS), E.coli DH5α is not a dangerous substance. Base on Centers for Disease Control and Prevention (CDC) Guidelines (Biosafety in Microbiological and Biomedical Laboratories, 5th Edition¹), E.coli DH5α is Biological Safety Level One organism.

Strain information of PCC 6803 and PCC 7942

　　PCC 6803 and PCC 7942 are the most widely used model organisms for photosynthesis research.^2,3 Both belong to the biosafety level one organism.

Suicide Mechanism

　　Being afraid of causing environmental danger, we designed a suicide mechanism, with the aim of killing E.coli. We designed suicide genes, endolysin and holing to clear away all of engineered E.coli, ensuring the safety. Endolysin and holin we used are similar to the mechanism used by team PeKing (2014 iGEM Beijing). Holin can induce holes on cell membrane. Endolysin can pass through the membrane and decompose peptidoglycan if holin successfully forms holes on cell membrane. After the cell membrane and cell wall are destroyed, E.coli will be lysed.

Lab safety

　　All experiments were done in Biosafety Level 1 (BSL1) laboratory. Our lab is equipped with safety facilities, such as emergency shower and fire extinguishers, and we conduct experiments under Laboratory Safe Hygiene Precautions of our school.⁴ To always remind us the importance of safety, the safety insert and postcard are hung at where can be seen easily in the lab. Furthermore, all experiment protocols were checked and guided by our PI, Dr. Chou before we conducted.

Lab training

　　Every members of wet lab in NYMU iGEM team had finished lab safety lesson and training twice, which are directed by professors who instructed us step by step. Thus, members could conduct experiments properly and safely.

Protection

　　In the laboratory, each member wears laboratory coat and gloves with long pants and shoes which cover feet entirely. Before going out of the experimental zone, we take off the coat and gloves, and wash our hands with antibacterial hand wash.

Waste

　　After experiments, we gather chemical and bacteria mixture into a bottle and add bleach. Besides, we sterilize trash with autoclave and put it at the specific area for contaminative trash. All the wastes in our lab are handled seriously and strictly to prevent contamination.

Reference

1. U.S. Department of Health and Human Services. (2009). Biosafety in Microbiological and Biomedical Laboratories (5th ed.). HHS Publication No. (CDC) 21-1112
2. Bertram M. Berla, Rajib Saha, Cheryl M Immethun, Costas D. Maranas, Tae Seok Moon, and Himadri B. Pakrasi. (2013). Synthetic biology of cyanobacteria: unique challenges and opportunities. Front Microbiol, 13(4), 246.
3. Raul Muñoz, Cristina Gonzalez-Fernandez. (2017). Microalgae-Based Biofuels and Bioproducts: From Feedstock Cultivation to End-Products. Woodhead Publishing.
4. Taiwan National Yang Ming University. (2010). Laboratory Safe Hygiene Precautions. (http://ces.web.ym.edu.tw/ezfiles/151/1151/img/183/laboratorysafehygieneprecautions-991001.pdf)