In accordance to the iGEM safety rules, our team used a cellular organism classified in the Risk Group 1. For the purpose of cloning and expression, Escherichia coli K-12-strain derivates were primarily applied in the lab as the main experimental subjects. Standard plasmids with standard antibiotic resistance markers were designed to enhance functions of recombinants selection. To ensure the safety of ourselves and the project, we worked in a BSL-1 (Biosafety type 1) laboratory, following standard safety protocols (e.g. masks, gloves, googles, and protective clothing). The lab also includes the “clean-to-dirty” workflow area, which means that lab staffs do not return to a “clean” area after working in a “dirty” area. For instance, entering PCR room is forbidden after working with DNA isolation or bacterial cultures in DNA room.
One of our project’s part which raised concern about biosecurity is the synthetic hand-made Affitoxin (i.e. the binding domain of AB diphtheria exotoxin) design. The details regarding this part would be discussed in this page. The fact that this part originates from the modification of diphtheria toxin raises concern about safety and security, including the possibilities of dual-use research. Regarding safety issues, the main purpose of the creation of this part is to eliminate the toxic domain of the exotoxin, leaving only the binding domain to be used to test our chimeric HB-EGF/Tar receptor. However, this condition still leaves a security issue regarding dual-use possibilities. Additionally, the creation of this part, if successful, means that we have only isolated the necessary domains needed for binding assays. The full sequence could be used to construct another chimeric protein in which the Affitoxin is linked to a more harmful toxin or material and used to deliver that harmful part into the cytoplasm of human cells containing HB-EGF surface receptor. To handle this, our team are required to comply with rules of iGEM safety committee to be allowed to not publish the full sequence in the iGEM UI 2018 Wiki pages.
