Safety

Project Design Safety

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An important part of doing research responsibly is doing it safely in accordance with all the policies and rules set up by the organisations around us. iGEM's policies are there to keep us and the public safe, so for any activities we wanted to do that could have gone against these policies, we had to both submit a check in form to iGEM, but also clear with our university. These check in forms were approved by iGEM and are explained below.

Activity	What it challenged	Why we could not use an alternative	Why it was approved
Taking our Cas12a/13a proteins outside the laboratory in the context of a simulated test we could send to teams to try to collaborate.	Release beyond containment policy as they are a product of a GM organism (BL21(DE3)).	These Cas proteins were a key part in our test and could not be replaced for a simulated test, compared to blood spiked with the targets vs real samples of infection.	No GMO would be released as we have a detailed lysis then filtering protocol in the process of purifying these proteins. No risk in regards to the actual proteins.
Using blood (defibrinated horse blood or pork blood from a butchers) to simulate how our test results (fluorescence) can show up within blood from a cow. It would be used to see how much blood blocks the signal, and tests with blood spiked with the targets we are looking for.	Using a sample from an animal.	As we are using fluorescence and testing trying to take out the colour from the samples, we needed to use actual blood as it has the red blood cells that we would need to deal with, not just a red dye.	The blood samples we got are either food safe, or have no risks related as they are from a supplier and used at our university with no risk identified.

Laboratory Safety

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We all had training as team members on how to work appropriately in a Level 2 biosafety lab. In our project we only used Level 1 organisms (E.coli strains DH5 alpha, BL21 (DE3), Rosetta) and although we are looking at how to detect Mycobacterium bovis, we do not use any large parts of its genome or anything that codes for whole proteins or virulence factors. We still followed the safety requirements of the lab, and any COSHH and SOP forms, which are detailed below.

Laboratory areas	Chemical Fume Hoods - Used for any volatile and hazardous substances as has airflow away from user to protect them. Biological Safety Cabinets - Used for any protocols that need to be kept sterile as air is filtered so limits contamination, and everything is sprayed with 70% ethanol prior to entering. Lab Bench - Where any other protocols take place, if the COSHH/Risk assessment/SOP does not require use of the above.
Waste disposal	All biological solid waste is autoclaved (by a trained technician and not us) at 121 °C for 20 min prior to disposal. (Red Bins) Biological liquid waste is treated with the lab grade multipurpose disinfectant Virkon for at least 6 hours before disposal down the sink. Hazardous chemical liquid waste (e.g. Ni resin) is disposed of in the correct brown bottle in the fume hood and removed by a commercial waste company. Hazardous solid chemical waste is disposed of in the correct bin for incineration by a commercial waste company. (Yellow Bins) Spills are cleaned up in accordance to the relevant COSHH forms, and sterilised with 70% ethanol where required.
Personal protective equipment	Nitrile gloves (EN 374) and a lab coat are always worn at a minimum. Safety glasses (EN 166) are worn in accordance with relevant COSHH forms to the protocol.
Protocols	We have all been trained in good microbiological techniques such as changing pipette tips when changing liquid taken up etc to prevent contamination.

Hazards and Risks

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With all activity that takes place within the lab, there is a level of risk attached. Whilst lab work was taking place, hazards were identified throughout the project, and minimised to the best of our ability to lower the risk of these activities. If any activity did cause harm, the relevant safety training had been done so all team members knew where to go and what to do in the event of an injury. Below are some examples of the risks we identified in the COSHH forms or SOPs we filled out for our protocols and how we minimised the risk.

Activity	Hazard	How it was minimised	What to do in event of injury
Melting/ dissolving agar	Superboiling in microwave	Members instructed to only microwave for a few seconds at a time to limit the possibility of boiling over, and to swirl to agitate the mixture.	If skin was burnt by hot liquid, rinse under cool water.
Agarose powder	Can irritate respiratory tract if inhaled	Try not to agitate powder to cause dust in the air.	Take victim outside to breathe fresh air and contact a doctor if symptoms persist.
Ammonium chloride powder	Causes serious eye irritation and harmful if swallowed	Normal use of the powder should not result in contact with eyes or swallowing, but powder should not be agitated and cause dust.	Rinse out eyes with clean water and drink water if swallowed, and contact a doctor if feeling unwell.
SYBR Safe DNA gel stain	Causes skin and eye irritation	Use in the fume hood with gloves and goggles. Any waste is disposed of in yellow chemical waste bin.	Rinse out eyes with clean water and drink water if swallowed, and contact a doctor if feeling unwell.
Hydrochloric acid	Corrosive to metals, causes eye and skin burns/damage and maybe respiratory irritation.	Stored in glass containers, used in fume hood, goggles (plus lab coat and gloves) worn when using it.	Move to fresh air if experiencing respiratory irritation and rinse eyes and skin with plenty of water and remove contaminated clothing.
Running gel electrophoresis from power pack	Electric shock	Training by an experienced person before use and checking all leads and tanks are plugged in correctly before use.	Turn off electric supply and call 999 for additional help if injuries are severe.