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From 2013hs.igem.org

Contents 1 Safety 2 If The Project Goes According to Plan: 3 If The Project Does Not Go According To Plan: 4 Biosafety Review Board at St Pauls:

Safety

On top of all of the safety requirements and mitigation strategies discussed below, all lab-work conformed to and was carried out in accordance with the SPS safety requirements. All lab-work was supervised by members of staff and lab technicians at all times. The SPS lab safety policy is below. The general school safety policy can be found here: [1]

1. Your own safety and that of others depends upon sensible behaviour in the laboratories. The Health and Safety at Work Act holds teachers and pupils responsible for their actions if others are harmed by them.

2. You should never run, push other people or throw anything when you are in a laboratory.

3. When you are doing practical work always work methodically, using care and common sense. Rushing leads to accidents.

4. If you are not sure of what you are supposed to do, ask for help. Listen to, and act on, any verbal instructions your teacher may give.

5. You should only go into a laboratory if you have permission to do so.

6. Apparatus and chemicals may only be removed from a laboratory if you have permission to do so.

7. Don't touch apparatus, electrical equipment or chemicals (or the controls of the gas, water and electricity supplies) unless you have been told to do so.

8. If you have any doubts about the safety of any equipment you are expected to use e.g. a jagged piece of glassware, consult your teacher.

9. Report ANY accident, however slight, to your teacher.

10. Waste, or surplus, materials should be disposed of as instructed by your teacher.

Do not throw solids into sinks.

Do not pour non-aqueous liquids into sinks unless told to do so.

Do not return chemicals or solutions to their original storage containers.

11. Don't put hot objects directly onto a wooden bench.

Make sure anything burning or smouldering e.g. a splint, is fully extinguished.

Allow hot equipment or substances to cool before you touch them, store them or throw them away.

12. Don't sit or lean on a laboratory bench, as you or your clothing may come into contact with harmful substances. If you spill any substance accidentally, clear it up after consulting your teacher.

13. Avoid contact of chemicals with your skin. Wash your hands after laboratory work; this is especially important before meals.

14. If chemicals or apparatus are left on the bench from the previous lesson do not touch them but immediately inform your teacher.

15. Always leave your bag in a safe place in the laboratory, so that it cannot cause an obstruction.

16. Wear the eye protection provided at all times during Biology and Chemistry practical work, and when you are told to do so in Physics.

17. Long hair must be tied back during practicals.

18. Ties and shirts should be tucked in during practicals.

19. Don't eat or drink in laboratories unless invited to do so by the teacher in charge.

20. You should know the meaning of the hazard warning symbols found on storage containers.

Organism Used:

The organism we are using is an E.coli K12 derivative; they are BL21. They are in risk group one as they have had their pathogenic material taken out of them and so it is highly unlikely that they will cause disease in healthy adult humans.

Organisms parts are derived from and their respective risk groups:

Letter Code for Part	BioBrick Reference	Function	Organism Derived From	Risk Group
A	BBa_K418003	Promoter	E. coli strain NEB 10-beta	1
B	BBa_B0033	RBS	E. coli strain NEB 10-beta	1
C	BBa_C0040	TetR gene	E. coli strain NEB 10-beta	1
D	BBa_C0072	Lacl gene	P22 bacteriophage	1
E	BBa_E1010	RFP gene	Discosoma striata	1
F	BBa_B0015	Terminator	E. coli strain NEB 10-beta	1
J	BBa_K091104	Promoter	E. coli strain NEB 10-beta	1
K	BBa_B0034	RBS	E. coli strain NEB 10-beta	1
L	BBa_E0030	YFP gene	Aequorea victoria	1

If The Project Goes According to Plan:

Risk to Team members:

Very small risk. The E.coli has its pathogenic material taken out of it and so has a “low but not non-existent virulence”. The risk to team members is therefore small since, as with all organisms classified in risk group 1, it is not known to consistently cause disease in healthy adults. Furthermore, any risk to team members is further mitigated through washing hands with alcohol rub before and after practical work and wearing gloves and safety glasses at all times while working with the bacteria. The bench is also disinfected with Dettol (anti-microbial) before and after lab-work.

There is also only a very small risk posed to the team members by any laboratory equipment or chemicals. The laboratory equipment used included:

1. Microcentrifuge

2. Plate incubator

3. Broth Incubator and Shaker

4. Gel electrophoresis apparatus

5. Micro pipettes

6. Mini-prep kit

Among the chemicals used were the antibiotics chloramphenicol, Ampicillin, Kanamycin and Tetracycline. Gloves and safety glasses were worn at all times while the chemicals were being handled. Teachers and lab technicians supervised the handling and making of antibiotic stock and team members washed their hands immediately afterwards.

Risk to the Public

Very small risk. It is highly unlikely that that the public will be exposed to the bacteria before the project is finished through bacteria escaping from the lab – workbenches are sprayed and wiped down with Dettol (anti-microbial) after lab-work and all team members wash their hands with soap and alcohol rub before leaving the lab. All waste is disposed of in specially allocated waste bins, the contents of which are autoclaved and then incinerated. We work in a class one containment laboratory.

Once the project is complete, there is still a very small risk to the public; the fact that the bacteria are in risk group one means that it is highly unlikely that they will cause disease in a healthy adult. Moreover, though the bacteria are intended to be used in food testing, this is expected to occur in a laboratory and the samples of food tested will not be the same as any food the public might ingest. The public are therefore expected to never encounter the bacteria themselves.

Risk to the Environment:

There is only a very small risk to the environment. It is highly unlikely that the bacteria will escape from the laboratory as it is a class one containment laboratory – all waste is autoclaved and then incinerated and benches are disinfected while all team members wash their hands with both soap and alcohol rub.

If the bacteria were to escape into the environment, then it is highly unlikely that there will be significant environmental damage or a significant detrimental effect on the local ecosystem. Though the E.coli have the potential to infect mammals, it is unlikely that they will cause disease as they have had their pathogenic material removed and are in risk group one. They are therefore unlikely to upset local ecosystems in the unlikely event of the bacteria finding their way into the external environment.

If The Project Does Not Go According To Plan:

Risk to Team Members

There is only a very small risk to team members if the project does not go according to plan – the bacteria itself is only in risk group one and none of the parts or biobricks used are harmful or could cause the bacteria to develop into a more dangerous, pathogenic organism. The project will not go according to plan only if the bacteria do not take up the biobricks in the mutated plasmids we provide it with. Whether this happens or not, the bacteria will remain relatively harmless.

Team members further mitigate the risk to themselves by washing their hands with both soap and alcohol rub before and after practical procedures, wiping down the benches with anti-microbial disinfectant before and after the procedures and by wearing safety glasses and gloves at all times during the procedures as previously discussed.

Risk to the Public

There is only a very small risk to the public if the project does not go according to plan. It is highly unlikely that containment procedures will go wrong and that organisms or parts will be released from the laboratory – it is a class one containment laboratory and all methods to contain the bacteria are as previously discussed. If both the project does not go according to plan and the bacteria or parts escape from the laboratory then there is still only a small risk to the public. Neither the bacteria nor the parts are inherently dangerous, the bacteria being in risk group one and all parts being derived from organisms that are in risk group one.

There is little risk to security through malicious misuse of the bacteria or the parts. The only people with access to the parts and the bacteria are the team members and the team supervisors and advisers. The laboratory doors are locked at all times when no one is working in the lab and there is always at least one supervisor present when lab-work is being carried out.

If the bacteria or the parts were to be hijacked or obtained by foul means, there is still only limited potential for malicious misuse; the bacteria are in risk group one and have had their pathogenic material taken out of them. None of the biobricks are inherently dangerous either. Therefore any damage through misuse would mainly be economic in nature: the project is designed to be used in food and allergy testing and it would require a lot of effort to develop the bacteria so that they became dangerously pathogenic.

Risk to the Environment

There is only a very small risk to the environment. Even though E.coli is able to infect mammals, the bacteria we are using has had its pathogenic material taken out of it and so it is unlikely that it will be able to cause disease. This is certainly the case in humans and it is likely that the same can apply for most mammals. It is therefore unlikely that the E.coli would upset local ecosystems if it were to escape. It is also highly unlikely that the E.coli or any parts will escape since it is a class one containment laboratory. The same applies for if the project does not go according to plan – if this happens then the bacteria will not have taken up all of the biobricks and so will remain as they were when we started work. Either way, they are relatively harmless.

Biosafety Review Board at St Pauls:

We have an internal review board that is chaired by Dr. Jon Bennett. Any proposals or suggested changes are first put to him to approve. This also ensures that there are always at least two people to judge and assess decisions on safety and biosafety in the laboratory.