Introduction:

The Shym_BIL_NIS team believes that safety is a key factor in the development of science, establishing what is harmful, identifying deviations of values and morals, being responsible for the conservation of entire ecosystems and the whole of humanity, adhering to certain norms, measures and standards. Reliability and safety, these characteristics that our team wanted to reflect in our work. For more information, check out our safety form.

    BSL-2 plus:
  • Physical separation from access corridors
  • Self-closing, double-door access
  • Exhausted air not recirculated
  • Negative airflow into laboratory
  • Entry through airlock or anteroom
  • Hand washing sink near laboratory exit
    BSL-2 practice plus:
  • Controlled access
  • Decontamination of all waste
  • Decontamination of laboratory clothing before laundering
    Primary barriers:
  • BSCs or other physical containment devices used for all open manipulations of agents
  • PPE: Protective laboratory clothing, gloves, face, eye and respiratory protection, as needed

1. Each member of the team got acquainted with the safety techniques for working in the BSL-2 safety level laboratory.

2. The main protocols of this level were also carried out in details, such as: waste separation, sterilization of glassware, and use of personal protection equipment.

Our team understood that working in the laboratory required special attention, so we carefully studied all the necessary protocols before working.

Microorganisms review:

Our team knew that working with microorganisms requires a lot of responsibility, because there is always a risk of releasing them into the environment, while the laboratory work, which can become a big threat to public safety. For this reason, plasmid transformation was performed in non-pathogenic and well-characterized strains of Escherichia coli: BL21 (DE3), Rosetta (DE3, gami) and DH5 alpha. In addition, the subsequent transformation was carried out in Pseudomonas putida KT2440 cells, and this strain is certified as “generally recognized safe", with stable properties and universal metabolism. These strains do not cause serious diseases or other problems neither for people, nor the environment and belong to BSL-1 organisms.

Some experiments require the usage of harmful substances and toxins. Our team tried to adhere to all safety standards when using these substances. Antibiotics were used in the transformation of DNA sections into bacterial plasmid as experimental indicators. Our team made sure that when working with antibiotics, they did not get into the eyes and skin areas to avoid allergic reactions. All the work was done on the basis of certain safety instructions. UV light was used to sterilize all tools and used items after working with cells in the safety cabinet.

We have transformed the gene responsible for the expression of the PETase enzyme in the Pseudomonas putida KT2440 bacterium, and based on theory, this enzyme decomposes PET to monomeric mono-2-hydroxyethyl terephthalate and is an ester. Due to its complex structure and strong hydrogen bonds between ethylene glycol and terephthalic acid molecules, it does not pose any harm to the environment, since compounds of this type are widespread in everyday life and are used in mass consumption.