Choosing a non-pathogenic chassis

Bacterial strain: Escherichia coli K12

Escherichia coli K-12 are not a threat to human, animal (chickens, pigs, and calves), plants or the environment, according to the 1997 Final Risk Assessment of Environmental Protection Agency (EPA). “Any concerns in terms of health considerations are mitigated by its poor ability to disseminate, colonize the colon and establish infections in a murine model” (Smith et al., 2010). In addition, the chance of the insertion gene mutating the bacterial strain to be hazardous is minimal as the inserting genetic materials have to meet the EPA’s criteria (limited in size, well characterized, free of certain nucleotide sequences, and poor mobility).

Choosing parts that will not harm humans/ animals/ plants

Inserts:

Our GEOs are composed of standard promoters, ribosomal binding sites, terminators and fluorescent reporters, or have no known risks. The function and known homologs are classified by BLAST (http://blast.ncbi.nlm.nih.gov/Blast.cgi) and researched (mainly via PubMed). For more information on researcher, public, and environmental safety refer to Safety Q & A

Working with a system that can be substituted for a safer suitable system compatible with the intended work is illegal as well as immoral, even if the alternative is more difficult or expensive to acquire. In our project, safety is favored before monetary issues therefore we always attempt to locate a better host, donor, vector, insert, or system for humans, animals and the environment.

Since Ethidium bromide is a very toxic agent and acts as a mutagen (intercalates double stranded DNA), we have changed our nucleic acid staining molecules to Invitrogen SYBR Safe, which is significantly less toxic and better for the environment (may have direct disposal into the wastewater drainage systems). In case EtBr or Invitrogen SYBR Safe affects DNA biological processes, these substances are added in minimal concentration to the agar solution before it solidifies.