Safety is a prerequisite for scientific experiments, and we attach great importance to project safety and laboratory safety.

Secure laboratory

The laboratory of Peking 2023 is a BSL-1 laboratory with more than 10 years of operating experience and complete experimental facilities, including biosafety cabinets and chemical fume hoods. Laboratory safety is supervised by Associate Professor Qingsong Wang.

At the beginning, Peking 2023 conducted a four-day laboratory professional training for all staff, learning laboratory safety rules, instrument operation, basic experimental skills, and emergency measures. Before conducting experiments independently, students need to be guided by experienced senior students to conduct experiments. In addition, Peking 2023 has established a laboratory management system and is equipped with an experienced safety officer. Specifically, we have the following measures (also mentioned in the safety form):

· Personal Protective Equipment / PPE

· Inventory controls

· Physical access controls

· Data access controls

· Lone Worker or Out of Hours policy

· Waste management system

Secure project design

Microbial chassis selection

At the beginning of the project, Peking 2023 intended to select VNP20009 (a strain of Salmonella) as our chassis due to its high affinity to tumour tissues. However, it is modified by a pathogenic bacterium, which is not conducive to our experimental operation and future applications.

Therefore, we chose Escherichia coli Nissle 1917 (EcN) as the main microbial chassis. EcN is considered a non-pathogenic strain of E. coli and is often used as a probiotic. Many microbial therapies use EcN as a chassis. Peking 2023 hopes to make EcN obtain tumour affinity close to VNP20009 through precise AND gate design.

Knocked out the Asd gene

Peking 2023 constructed a diaminopimelic acid (DAP) auxotrophy recombinant bacterium strain by knocking out the asd gene of EcN, which encodes aspartate beta-semialdehyde dehydrogenase, a key enzyme in the biosynthesis pathway of DAP (the basic component of peptidoglycan on the cell wall). DAP is essential for the cell wall of EcN. It would die without exogenous DAP added, eliminating the possibility of leaky expression.