Team:Paris Bettencourt/Safety
Safety in the Lab
The lab and training
French research is governed by French laws and should follow specific rules for the manipulation, experimentation and release of genetically modified organisms. Therefore, during our iGEM projects, we were advised by our PIs and instructors, and we followed the current regulations for all biological and chemical manipulations. (1) We performed all of our experiments in a laboratory of biosafety level 1 (the lowest) equipped with open benches, a chemical fume hood, and a laminar flow hood.
All the team members who worked in the wet-lab section of the project received safety guidelines by the lab manager Paulina Ejsmont and online safety-security training by the institution INSERM. For the specific experiments that required the use of more sophisticated machines like a flow-cytometer, instructors supervised the manipulations. All the necessary Personal Protective Equipment (PPE) were worn to ensure the safety of the experimenters and the cleaning of the laboratory was realized daily to avoid any contamination. All the microorganisms were manipulated under special sterile conditions: under a clean hood, with gloves and airflow. All the chemical products with special risks were manipulated under the chemical flow hood, with aeration, one-usage gloves, and if necessary glasses. Hazardous chemicals were handled one by one to prevent risks linked to their mixing. Hazardous substances were stored following SDS (Safety Data Sheet) recommendations. Highly-concentrated powders were stored in small quantities, hazardous chemicals were stored in separate storage units depending on their type.
The experiments and the products used were subjected to validation. In addition, the material and methods section of the papers we used for these experiments were carefully studied to avoid handling errors.
The chemicals
Antibiotics: To select bacteria, we used several antibiotics (including Kanamycin, Chloramphenicol…..) that could have hazardous effects on human health at high concentration and in case of inhalation. Stocks were made using contrentaded powder stored in small volumes to prevent danger in case of spilling. Working stocks are not considered hazardous when manipulated with care. No members of the team were identified as sensitive to antibiotics therefore, no specific measures needed to be implemented more than normal safety measures and PPE.
Others: All the other chemicals with special risks (e.g DAPI being carcinogenic) were manipulated with special precautions under a chemical fume hood and after training with our lab manager and/or our PIs. Precaution of usage provided by suppliers were carefully respected.
The strains
During the experiments, different organisms have been manipulated. Since we only worked with Prokaryotes and phages, no special permits or requirements where needed. First, we received 3 different strains of E. coli. The E. coli K-12 MG1655 and the E. coli K-12 MinB are strains that we received from two of our professors from the CRI institute. The first is the regular laboratory adapted non-pathogenic one and the second is a mutant with a knockout of the min operon. We also asked the iGEM Marburg team from 2015 who also worked on minicells to send us their E. coli K-12 TB43 mutant minicell producing strain. Most of our work was done on E. coli MG1655, a non-mutant strain especially for genetic engineering and the other ones were used for characterization mostly. Some of the organisms we used have the property of synthesizing minicells. They are considered bacterial vesicles or non-living organisms. However, during manipulations and storage, we ensured the same precautions as we had when manipulating bacteria.
E. coli K12 strains:
- MG1655 from the CRI collection
- TB43 from Marburg collection
- MinB from Ariel Lindner collection
Minicells produced by TB43 strain
Minicells produced by E. coli K12 MinB strain
Minicells produced by E. coli K12 MinB strain
λ-Phage (replication deficient phage)