Lactobacillus Union

Early in our iGEM 2022, to collaborate with the Latin Amerca iGEM teams, we made contact with many of them and organized the LATAM Meetup collaboration, which enabled us to meet many other Latin iGEMers and get to know about their projects. During the presentations Chassei got our eyes, the Costa Rica team’s idea consisted in the characterization of Lactobacillus casei, allowing the use of its great potential. On the discussion day of the meetup, the topics that we talked the most revolved around our chassis, since both teams were working with the same chassi genre.

Thinking about the topics in the discussion, we realized our teams could help each other, so we invited them for a more specific talk about our projects. This was the first stepping stone for our partnership throughout the competition, often in contact, aiming to solve together the problems we were facing.

Development of an effective Lactobacillus Transformation Protocol

One of the facts that make Lactobacillus hard to work with, besides the lack of characterization, is the difficulty to achieve its successful transformation. Our team had some obstacles transforming Lactobacillus acidophilus, but we succeeded with the help of a professor. However, Chassei members were struggling to transform Lactobacillus casei: they had tried 3 protocols and could not transform their chassi. With that, they got in touch with our team and we analyzed the protocols they were trying to use. Considering our team's experience, we identified and adapted the problematic steps in the protocols that failed, developing a transformation protocol for Lactobacillus! Using the new protocol Chassei proved its reproducibility and could continue with their project development.

Check out the protocol we have created together:

pLBov Plasmid Reproducibility Test

In addition, Chassei sent us their own designed pLBov plasmid to test in Lactobacillus acidophilus, for the transformation and expression, for us to validate its function in another Lactobacillus species. Unfortunately the sample did not arrive in time for the conclusion of the experiments and final validation.

Development of Killswitches

Other discussions that we had with Chassei were related with the biosecurity of our microorganisms in their applications. Our team was having trouble designing an efficient Killswitch, that would prevent the spreading of our genetic circuits and GMO in the environment. From that, we talked to team Chassei and worked together to build our team's Killswitch. Firstly, they shared with us their experiences with their Killswitch development. Then, we received suggestions about the different types of inputs and approaches that could be used for our circuit and we evaluated which ones would fit better for what we needed. They helped us to find inputs that consisted of the common physiologic human gut and external environmental signals. After this, our team crossed data of literature articles and iGEM registry and chose the inputs and biological parts used. The partnership helped us understand the key points about the Killswitch constructions. Because of this, we decided to make two Killswitch circuits, one for the vector and other for the host, to avoid the genetic material conjugation between bacteria. Chassei team members also helped us with the literature research, and with the choice of their Biobrick endolysin as the specific toxin for our host. Check out the result of this partnership, our Killswitches, in the Safety page.

We would like to thank Team Chassei for this partnership, where we overcame shared and individual problems. The good exchange of experiences generated impactful results, providing one more step for the better use of the Lactobacillus for the next iGEM teams and the science community in general.