Team:Tsinghua/Safety
Safty
Safety workshop
As a lab of biosafety level 2, we joined safety workshop, interacting with other teams whose projects are of biosafety level 2 as well, in order to figure out a more efficient way to improve our lab safety. After this meeting, we, the ones whose project is about anti-biofilm, established detailed lab safety rules for anti-biofilm community (ABC). This rules also have been carried out by all the students joining the project.
Safty WorkShop we took
Safty Form
Contact info：
Team: Tsinghua
Name: Qinglin Zeng
Email: 985586986@qq.com
Part 1: Overview:
Our project is to use E.coli to construct an in vivo nitric oxide generator to inhibit the biofilm,especially the biofilm of P.aeruginosa in the PVC pipelines. This system will be induced to function by the C4-HSL signaling molecules generated by the existing biofilm, in order to start the function of nitric oxide synthetase (NOS). Meanwhile we plan to introduce report gene(EGFP controlled by NO induced expression promoter Phmp) to the P.aeruginosa to identify the NO sensing. Considering the dark environment of the pipeline, we decided to use blue light sensitive kill switch to prevent our engineered bacteria from leaking.
Overview Tsinghua iGEM Lab and Part of the Lab Rules
In Tsinghua iGEM Lab, all equipment will be placed according to regulations
Part 2: Identifying possible risks
3. Which whole organisms, including viruses and cell lines, are you planning to use or using in your project?
We are planning to use Escherichia coli (BL21/DH5a based on K-12 strain), Pseudomonas aeruginosa (PAO1), Bacillus subtilis(168) in our experiment.
4. What risks could these organisms pose to you or your colleagues in the laboratory, or to your community or the environment if they escape the lab?
All these Normal strains will not pose threat to the environment or people in most of the cases.Potential risks of wounds and respiratory tract infection caused by Pseudomonas aeruginosa (PAO1).However, if which carrying antibiotic resistant gene escapes, the tolerance strain or this tolerance gene might spread.
5. What organisms are you using as chasses in your project?
1) Normal E.coli will not pose threat to the environment or people. However, if the E.coli carrying antibiotic resistant gene escapes, the tolerance strain or this tolerance gene might spread.
2) Pseudomonas aeruginosa
6. What risks could your chassis pose to you or your colleagues in the laboratory, or to your community or the environment if they escape the lab?
The unmodified E.coli will not pose threat to environment and people. However, if the E.coli with plasmids that carry the antibiotic resistant gene escape, the tolerance strain or this tolerance gene might spread. Meanwhile the Pseudomonas aeruginosa that carries certain resistant gene (kan or amp) may have potential risk to the environment.
7. which chemicals are you using in your project that might be hazardous?
Nitric oxide releasing agents (SNP)
8. as part of your project, are you are planning to make/have made new parts or substantively changed existing parts in the registery?
Yes.
9. Part information is submitted in a spreadsheet.
We've already uploaded the spreadsheet.
10. What experiments will you do with your organisms and parts?
In our first project, the RhlR-pRhl related parts will be used to generate an C4-HSL signal switch. We will use C4-HSL to test the efficiency of the switch in order to perform only face the existing biofilm. The NOS related parts will be used to generate a controllable NO releaser.
hmp-mCherry/EGFP system will be introduced to P.aeruginosa in order to prove the sensing of the NO signal.
light induced kill-switch (based on LEVI-ccdB) will be introduced to the engineered E.coli.
11. What risks could arise from these experiments?
The first risk is the recycle of the modified germs, before the suicide model is added, all the experiment performance should be take in appropriate situation(clean bench, gloves and masks) The NO produced by bacteria is relatively low(pmol) and will be harmful. However considering the using of NO donor reagents such as SNP, all the experiments related to NO are conducted under good ventilation environment.
12.Are you collecting any data about people, such as their opinions, quotations, health information, gender, behavior, attitudes, or concerns?
Yes.
13. Imagine that your project was fully developed into a real product that real people could use. How would people use it?
In the natural environment.
14. What safety, security or ethical risks would be involved with such a use?
The main risk of this application is the escape of transgenic E. coli. It would introduce synthetic genetic material to the environment, causing unpredictable gene transition in nature, including resistance for certain antibiotics. As our expectation, the engineered E.coli will be applied to clean the biofilm of P.aeruginosa in pipeline, which may increase the potential risk of contamination to the environment.
Part 3: Managing the risks you have identified
15. How will experts overseeing your project help to manage any of the risks you identified in this form?
The Tsinghua iGEM lab is part of the Center for Laboratory Training in Life Sciences in Tsinghua University. Lab instructor Li will oversee our experiments. Also we will work with instructors from Chen Lab, the laboratory of our Primary PI that specializes in microbiology research. Last but not least, we have a security commmittee evaluating the risks that might happen in our experiments.
16. What rules or guidance cover your work which could help to manage any of the risks you identified in Part 2 of this form (in particular Question 10)?
http://www.gov.cn/zhengce/content/2008-03/28/content_6264.htm
https://www.tsinghua.edu.cn/qhdxsysysbc/info/1031/1159.htm
17. Have your team members received any safety training?
Yes, we have already received safety training and we have already received security training.
18. Please select the topics that you learned about (or will learn about) in your safety training.
Lab access and rules (including appropriate clothing, eating and drinking, etc.
Responsible individuals (such as lab or departmental specialist or institutional biosafety officer)
Differences between biosafety levels
Biosafety equipment (such as biosafety cabinets)
Good microbial technique (such as lab practices)
Disinfection and sterilization
Emergency procedures
Transport rules
Physical biosecurity
Personnel biosecurity
Dual-use and experiments of concern
Data biosecurity
Chemicals, fire and electrical safety
19. Which work areas do you use / are you using to handle biological materials?
Open bench, Biosafety cabinet
20. What is the biosafety Level of your work space?
Level 2.
21. What other risk management tools will cover you work?
Accident reporting (measures to record any accidents)
Personal Protective Equipment (including lab coats, gloves, eye protection, etc)
An inventory control system (measures to track who has what materials and where they are)
Access controls (measures to control who can access your work spaces, or where materials are kept)
Medical surveillance (measures to find out if you get sick because of something you were using)
Waste management system (measures to make sure waste is not hazardous before it leaves your institution)
Special procedures or protocols that address safety or security
22. How will the rules, training, containment and other procedures and practices help to manage any of the risks you identified in Part 2 of this form (in particular Question 10)?
We will not use pathogenic strains of E. coli. The experiment related to PAO1 will only be conducted in biosafety cabinet.
Experiments will only be conducted when gloves and other protections are worn properly.
Wastes containing living bacteria or harmful reagents will be sterilized before being disposed, or sent to qualified institution for disposal.
Only iGEMers have can access our lab to carry out experiments.
We also have security rules which all iGEMers will obey. The rules are confirmed by lab instructor and all iGEMers.
Part 4: Compliance with iGEM’s rules and policies
23. Are you planning to/ have released any organism or product derived from your project?
No.
24. Are you planning to use, or using any animals (including insects and invertebrates) not on the Whitelist?
No.
25. Are you planning to use / have used any vertebrates (e.g. rats, mice, guinea pigs, hamsters) or higher order invertebrates (e.g. cuttlefish, octopus, squid, lobster)?
No.
26. Are you planning to use, or using any parts not on the Whitelist?
No.
27. Are you planning to carry out any of the activities not on the Whitelist?
No.
28. Are you planning to use, or using any parts or organisms obtained from outside the lab or regular suppliers?
No.
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