Team:OUC-China
Our team is committed to developing a directional intelligent screening platform. Every member of our lab clearly knows the necessity of ensuring the safety of the experimental environment, and has been strictly abiding by the rules and regulations of our lab. On this page, we will introduce what we do to be responsible for the environment and our colleagues in the experiment.
Safe Project Design
The development of our DISP project is based on two microorganisms: the Aureobasidium melanogenum P16 and the E.coli DH5α. Before we choose our chassis organisms, we have checked the data from China CDC, NIH Guidelines and American Type Culture Collection (ATCC) product sheets in detail to ensure our chassis organisms are selected from Risk Group 1 in our design. It means our chassis organisms do not cause disease in healthy adult humans. In addition to this, all genes we have utilized are coming from Arabidopsis thaliana, the circuit we build are harmless for the growth of cassis organisms and relatively safe for applications.
We have also checked the toxicity of each molecule in accordance with the Hazardous Substances Data Bank (HSDB). All molecules are harmless to human.
For the sake of environments, all the chassis organisms are sterilized before being abandoned to prevent genetic from leaking.
Safe lab work
We work at a neat lab named “Lab for Microinnovation and Enterprise”, which is a BSL-2 laboratory. Before working in the lab, we are supposed to be trained, not only for the general lab safety rules, but also for the standard experimental operation avoiding unnecessary damage. There is no zero risk to the experimenter in any experiment, hence we are properly equipped with facilities, such as latex gloves, nitrile gloves, goggles, lab coats to protect us from biotic and abiotic hazards.
Furthermore, several operational zones are divided scientifically for special experiments like A zone where is weighing the drugs, preparing the medium and sterilizing, B zone where is the recording experimental note and organizing experimental data area, C zone where is a molecular operation area, D zone where is a polyacrylamide gel electrophoresis area. Meanwhile, our lab is also equipped with a sink and an emergency eyewash, this will keep us clean and ready for emergencies.
This year, we have revised the laboratory safety manual and enriched its content. At the end of each experiment, the experimenter of the day will clean up and ensure the safety of the laboratory, and sign the experiment safety record book.
To our delight, we also replaced our clean bench this year, ensuring a clean experience environment.
Practical lab safety procedures
We have made a series of laboratory regulations to make sure that it is important to protect us from biotic and abiotic hazards and we use it every day.
1.Before the experiment
1) Dress in a standard way, wear lab clothes and gloves, and do not wear slippers or loose hairs.
2)Beverages (including water) and food are not allowed to enter the laboratory.
3)Be familiar with the instrument before the experiment to avoid damage to it due to unfamiliar operation.
2.Experiments
1)In case of injury, drop the work at hand for the first time to deal with it.
2)Operate in strict accordance with the experimental specification and the operating specification of the instrument to avoid errors or experimental failure or damage to the instrument due to personal reasons
3. After the experiment
1)Clean the table and tidy up the experimental supplies.
2)Waste drugs should be disposed of according to specific requirements.
3)Do not take anything out of the laboratory for biosafety issues.
Improvement of preliminary design
Toxic protein
After communicating with other teams, we found that using YopE as the suicide protein in our design not only posed a security threat to the platform's R&D personnel, but also posed a greater security problem to the staff of the microbial factory. After communicating with the DUT-China team to test the sacB toxic protein in their project, we decided to use sacB as the final suicide protein of our platform because of its good killing effect and better biosafety. The sacB gene encodes a secretory sugar polysaccharide, which can catalyze the hydrolysis of sucrose into glucose and fructose, and polymerize fructose into high molecular weight fructose and the accumulation of high molecular weight fructans has potential toxic effects on cells, which can cause cell death. So we decided to test the toxic protein to see if it is capable to replace YopE.
Prevent escape
Considering that the engineering bacteria designed on the platform will be used in the fermentation tank of the factory, in order to prevent a large number of production engineering bacteria from escaping into the environment and causing pollution, we decided to design a switch to prevent escape after communicating with other iGEM teams. We design another kill riboswitch switch which can detect the concentration of an important element in culture medium If it spreads into environment accidentally, our kill switch can detect the low concentration of the important element and kill the engineered bacteria. This suicide switch is coupled with the copy number system. We designed to place it in the reintroduced PFK gene. When the engineering bacteria are stored in the fermentation tank, because there are enough detected substances, the PFK gene is normally expressed. When the engineering bacteria escape to the environment, the lack of detected substances causes the PFK gene expression to be closed and the engineering bacteria to die.
