Genetically modified organisms escaping from lab is a serious problem since it will bring unpredictable impacts to our ecosystem. In our part design and production line, we were very conscious of biosafety. The yeast will pass the following process to achieve complete destruction. We also dealt cautiously with waste produced from the experiment and participated in safety training to ensure everyone conducting experiments had a minimal probability to cause biohazard pollution.

Filtering system

  The average size of the P. pastoris used is about 4–6 μm [1], and the filter we used is 33 kDa, which is equivalent to the size of tens of nanometers [2], more than sufficient to trap all the yeast.

Heating system

  According to the literature [3], heating at 70 °C for 30 minutes is sufficient to kill the cells. We used a temperature of 110 ° C for 10 minutes to kill any cells that might pass through the filter.

Following were our heating test:

Figure1: P. pastoris heated at 30 °C for 30 minutes

Figure2: P. pastoris heated at 50 °C for 30 minutes

Figure3: P. pastoris heated at 70 °C for 30 minutes

  We found that our P. pastoris not survival after were heated at 50 °C for 30 minutes. Thus, our production line heat process would kill P. pastoris passing through the filter.

Reference

1. 1. Gmeiner, C., Saadati, A., Maresch, D., Krasteva, S., Frank, M., Altmann, F., … Spadiut, O. (2015). Development of a fed-batch process for a recombinant Pichia pastoris Δoch1 strain expressing a plant peroxidase. Microbial Cell Factories, 14(1).
   doi:10.1186/s12934-014-0183-3
2. Bacher, G., Szymanski, W. W., Kaufman, S. L., Zöllner, P., Blaas, D., & Allmaier, G. (2001). Charge-reduced nano electrospray ionization combined with differential mobility analysis of peptides, proteins, glycoproteins, noncovalent protein complexes and viruses.
   Journal of Mass Spectrometry, 36(9), 1038–1052.
   doi:10.1002/jms.208
3. 3. Martínez, D., Menéndez, C., Echemendia, F. M., Hernández, L., Sobrino, A., & Trujillo, L. E. (2015). Kinetics of sucrose hydrolysis by immobilized recombinant Pichia pastoris cells in a batch reactors. J Microb Biochem Technol, 7, 294-6.