Throughout our project, we obeyed the iGEM safety rules. When conducting our experiments, all of the organisms used for developing our product stayed in our lab. To ensure safety during future use of our device outside the lab, we made sure that no biomaterials were spread to the environment.
Safety Training inside the Laboratory
Before we entered the lab, all of our team members had taken the "Laboratory Bio-Safety Training" course, "Laboratory Safety & Hygiene Training" course and “Hazard Communication Training” course and received respective certificates from NCTU. Furthermore, all experiments were strictly examined by the NCTU laboratory management system to ensure all procedures were safe.
Level and Rule of the laboratory
Our experiments were done in a BSL1 laboratory. Also, when we performed experiments in the laboratory, our advisers provided training regarding the correct operation of instruments, methods preventing containment, and ways to deal with laboratory waste. What's more, they supervised us during our experiments to ensure safety. All equipment used to experiment with and contain E. coli were sterilized with 75% alcohol or bleach properly, and experiments were operated in laminar flow hood to prevent any leakage or pollution of bacteria. We familiarized ourselves with and followed Taiwan bio-safety laws.
Escherichia coli Nissle 1917
E. coli Nissle (EcN) is a nonpathogenic bacterium isolated by Alfred Nissle in 1917. For nearly 100 years, the EcN strain has been used as the active pharmaceutical ingredient in a licensed medicinal product distributed in several countries. Because EcN is non-pathogenic, we can prevent the dog from being hurt.
Dog's gastric acid
Dog's gastric acid has a low pH value of about 1-2, which is much acidic than the proper pH for Escherichia coli, so this is very difficult for DenTeeth to live in a dog's stomach.
Inhibition of Growth of DenTeeth Test
To prove that DenTeeth can be inhibited by gastric acid, we conducted an Inhibition of Growth of DenTeeth Test. Because the HCl concentration of the animal's gastric acid ranged from 0.15M to 0.05M, we added 200μL HCl(aq) from 0.3M to 0.1M with equal DenTeeth and observed whether the DenTeeth in different concentrations of HCl(aq) would be inhibited. In addition, we chose ddH2O and HClO(aq) as our negative control and positive control, respectively.
The DenTeeth's biobrick could successfully degrade in any range from 0.1M to 0.3M HCl(aq). Only the sample added with ddH2O has the band in 4685 b.p., which was the length of our biobrick. The colony PCR result proved that the biobrick could not have the chance to transfer to another bacteria.
- Michael de Vrese , J Schrezenmeir.(2008)”Probiotics, prebiotics, and synbiotics7”Advances in Biochemical Engineering 111:1-66