Team:BostonU/Mammalian synbio/Significance

Significance

Safety and Concerns

One obvious concern that spans both the scientific and ethical realms are the safety issues that come from working with mammalian cells. Our iGEM team has thoroughly addressed these and hope to spread awareness through protocols and tips here (link to protocols page, specifically mammalian). Every member of our team completed BSL1 and BSL2 laboratory safety training through BU. With regards to BSL1 we utilized K12 E. Coli cells, an abundantly worked with and minimally threatening chassis. WIth regards to BSL2, we worked only with HEK293T cells. These cells were contained within one specific tissue culturing room. More specifically they were stored in a CO2-regulated incubator that had no other types of cells in it or they were worked with in our sterile, O2-regulated hood equipped with UV light to sterilize after being worked in. Working with the cells it was imperative to sterilize anything that came from outside the hood, but more importantly in concerns to disposing of the cells the priority was to kill all remaining cells and contain any media or other reagents they had come in contact with inside a tightly closed flask which was all disposed of as biohazardous waste. Safety protocols for this type of cell culturing are very important not only to verify that the cells are viable but more importantly that when they are no longer useful for research that they are properly disposed of and do not escape into any outside environments.

A concern that comes up further down this process is: if this split protein methodology is feasible, is it safe for human therapeutic use? This question was especially pertinent to our team as we were already testing in mammalian cells and had seen some promising results. What we want to emphasize is that our research is fundamentally trying to make safer a process which could be life saving in the future. Genomic editing and manipulation techniques are only getting better everyday, however, our project is not specifically focusing on making them better. We are focusing on the actual implementation of them. For example, right now it is possible that SaCas9 will recognize the wrong sequence and cut out the wrong gene, which could be potentially fatal. But with our system in place, it would hopefully be less likely that this would happen. There would be less chances for SaCas9 to even bind and carry out its functionalities since most of the time in vivo it would be split apart and inert. So already the protein is inactive for most of the time, however if it was required to be active the inducible dimerization provides for a simple and specific way to bring together the protein to carry out its genomic editing properties. We want to emphasize that this research is meant to directly address the natural, random problems that arise with any protein, and attempt to make them more regular and reliable with temporal control.

While we are excited to do foundational research in mammalian synthetic biology for such downstream genome editing and manipulation, we realized that this type of research is not heavily conducted in the iGEM community. However, many teams do intensive research for potential health and therapeutic applications and we feel it is important for this to get translated into mammalian systems for more precise downstream applications. We understand that there are many limitations to utilizing mammalian cells, but believe it is important to continue and intensify the dialogue in iGEM to promote it.

To better understand the climate in iGEM, we initially analyzed how many previous iGEM teams have used mammalian cells over the years.

It is clear above that the amount of teams using mammalian chassis are fairly static over the years, while other chassis like E. Coli increase greatly. It is obvious that there are many problems and impediments in using mammalian cells, and although we do not expect teams to immediately start working with them, we find it imperative to dig deeper and not only understand what the problems are but provide feasible feedback to potentially fix it.