Context

Kaneka Corporation utilizes microorganisms to produce coenzyme Q10 and biopolymers. We learned of the current situation through the interview with this company: strain leakage is occurring overseas. We wondered if Optopass could be effective to prevent unauthorized production in the situation where the factory manager is absent. We spoke with them for the purpose of clarifying how Optopass could be applied to industrial processes. We conducted this interview on September 30.

Interview: What we learnt

We learned about the production flow at Kaneka Corporation and recognized challenges at each stage. In the inactivation process based on the Cartagena Protocol, products are heat-treated in the culture bed for a certain period of time before being sent to the next purification stage. According to Kaneka, uniform irradiation and irradiation time were critical for using the light-induced kill switch instead of heat treatment. They also said that if a function that could control gene expression time-dependently and a mechanism that would allow genetically modified organisms to die in their lifespan were developed, the cost would be reduced because the sterilization process would no longer be necessary. As for the purification process, they wanted to simplify it as much as possible. They proposed a system in which microorganisms are recombined to prevent the accumulation of substances other than the target substance, and their activity is regulated by light. In this case, they would be more likely to use it if the effect of light irradiation lasts longer.

They also indicated a risk after unlocking the passcode. When closing down overseas plants, there is some risk of leakage of unlocked microorganisms. They hoped for a system in which, even after activation, the activity would be lost if certain conditions were not met.

Harvest: What we investigated after the interview

From this human practice to Kaneka, we found areas for improvement when implementing Optopass at the industrial level. This conversation clarified the difficulty of using the same light-illuminating equipment as in the lab and the necessity to examine how the light should be applied. In this process, we need to consider the impact on the culture. In particular, it is essential to conduct simulations to determine whether it is possible to illuminate the entire apparatus.

The same organization as above.

Context: Why we interviewed

Biobanks are institutions that promote research that links medical and genomic data in order to develop new medical treatments. We have been thinking about how Optopass could be used in society, particularly in medicine. As a result, we asked Tohoku Medical Megabank Organization, Tohoku University, running a major biobank in Japan, about what approach would be appropriate for applying Optopass to the medical field and what challenges there would be in implementation.

Before HP

The Optopass system could be used to safeguard the organism's genetic information. Therefore, we assumed it would be possible to prevent the leaking of genomic information related to patient privacy from the organism containing the gene, such as a genome-based drug. Furthermore, we thought that it could be used to protect genetic information during transportation, such as when delivering organisms containing genes useful in the field of pharmaceuticals to other researchers.

Interview: What they suggested

Initially, we considered the implementation idea of using Optopass to protect individual gene information. However, according to the Tohoku Medical Megabank Organization, it is often not individual genes but the entire metabolic system that is important to protect in the field of drug discovery, and drug discovery often does not use enzymatic synthesis. Therefore, the proposal to use Optopass to apply security to the genes of enzymes that work in some reactions was found to be less effective because it is difficult to keep the whole system secret.

Suggestion

When we asked how Optopass could be used in different ways, they said that by designing a "Dummy System," it could become a general-purpose technology to protect useful microorganisms on a "strain" basis, whereas the current Optopass method could only protect information gene-by-gene.

In this "Dummy System", the target strain and dummy strain coexist at first, and when exposed to light in the correct order, the dummies die and the target can be isolated. Although preventing the target strain from being sequenced is difficult, the idea is that this would allow "camouflage" with the dummy, making it more difficult to hit the sequence that one wants to protect. However, we have to keep in mind that it is also possible to destroy the hardware which contains the strains and isolate it by using selection to extract only the target strain that one wants to protect.

Action: How we reshaped our project after the interview

As its potential was expanded and the suggestion was to use it on a "strain basis", modeling was carried out to verify the feasibility of Optopass as the Dummy System. We designed a Dummy System where the microorganism with the light receptive genes can protect the security of the target organism by competitively being selected against the target strain when exposed to the wrong light or when the hardware is destroyed and exposed to a wide range of light, and the DNA sequence of the target would be cut and rendered unreadable as a precaution against isolation. More information can be found on Dummy System section in Modeling page.

Field

Recombinase design, Light inducer design

Context: Why we interviewed

Dr. Kawano specializes in optogenetics and is working with dCas9 and recombinase, which we were considering at the time to record order. The purpose of this interview was to get feedback on the technical feasibility and caveats about the overall project plan. In particular, we wanted to know whether dCas9 or the recombinase system was more realistic as a method of recording order from an expert's perspective. We conducted this interview on June 17.

Interview: What they suggested

Dr. Kawano taught us the merits and demerits of dCas9 and recombinase. The details are as follows:

dCas9

＋　 Highly scalable by increasing gRNA types

－　 Off-target is more likely to occur

－　 gRNA degradation is fast and unstable

Recombinase

＋　 Order information remains in the sequence after cell division and is reliably stored

＋　 Irreversible and stable as a system

＋　 Off-target is not likely to occur

－　 Low scalability

In addition, he told us about the existence of blue-light-active recombinase and the possibility of red-inducible promoters responding to blue light. When asked for his opinion on the project as a whole, he indicated that leakage of recombinase was the most critical bottleneck.

He also introduced us to an expert, Dr. Rei NARIKAWA, to investigate a third candidate color, green light receptors.

Harvest: What we learned and investigated about their suggestion

We compared dCas9 with the recombinase system and judged that the latter is suitable for our project. We confirmed that there are ten different recombinases and determined that it was scalable enough as a prototype. For example, a ten digit cipher in three colors could be made in 1,536 different ways.

Action: How we reshaped our project after the interview

We chose to use a recombinase system to make a prototype of the Optopass project ,which aims to use the order of light for a cipher. The reasons are that it is irreversible and stable as a system, it is less prone to off-target events, and it is simpler to implement. We also carefully studied leakage in the recombinase system in our modeling (see Modeling page).

Field

Education

We visited LEGO School Setagaya and had discussions with two instructors, as well as with three instructors in other schools, connected simultaneously via Zoom.

Through this human practice, the concept of Genochemy has drastically improved.

Context: Why we interviewed them

We wanted to introduce Genochemy to their instructors, have them use it, and get feedback. LEGO School teaches Robotics Programming to children. Although there is a difference between Genochemy for living creatures and LEGO for robots, they use a block programming tool called LEGO Spike for their education, and we thought that they might share the same issues with us, such as it being difficult to teach those who are not familiar with robots and programs.

Interview: What they suggested to us

The concept of the LEGO Spike tool is "low entrance, high ceiling," which means that it is easy enough for anyone to begin but has the potential to take the user to a very high level. It is a concept that we can strongly feel empathy with, as we want to target a wide range of people. We got a lot of concrete advice that we can apply to Genochemy.

• They pointed out various problems with the current page. For example, they said that the explanation of each block was too short and difficult to understand.
• They also commented on the user experience. They elaborated on the point that limiting the functionality to some extent makes it easier to understand. For example, they pointed out that the promoter block currently can be connected to the left side, but if the promoter block is used on the left end of the gene circuit in most cases, it would make more sense to design it so that it cannot be connected to the right side of another block (just as you cannot connect a block on top of an Events block in Scratch). It would make it clearer.
• We also asked for opinions on tutorials for beginners. We found that we should explain in detail what genes are and what DNA is.
• It is important to motivate the user. It should be something that makes people say "I want to make this today!", or "I could do this!"
• They showed us the screen of LEGO Spike and explained how it is used in teaching. Based on these, we worked together to come up with the following new concept for Genochemy.

This new screen proposal has many improvements over the current Genochemy.

(1) In Genochemy, the description in the block is currently very short, for example, "Constitutive" for the constitutive promoter, but in Scratch and LEGO Spike, the description is written in sentences. We were advised that this would be easier to grasp the image, so we will make the change. In response, the design of the block will drastically be changed so that multiple lines of explanation could be added.

(2) It was pointed out that the blocks in the current tray section are just lined up, and although the types of blocks can be recognized by their shapes and colors, they are not explicitly grouped, so it is difficult to understand what the blocks are. Therefore, we will modify it to display the name of the block type.

(3) To motivate the users, instead of making all the blocks available from the beginning, they will be unlocked as they progress through the task. This way, they can be surprised, like "Now I can do this!" "I didn't know it was possible to do this!".

(4) Rather than asking people to theoretically create a circuit with a certain kind of function, it is better to introduce social issues and what organisms can do to solve them, and then ask people to create their own genetic circuits, thus bringing it closer to the context of the real world. We think they will understand the usefulness of synthetic biology in solving social problems.

(5) The place where Genomy (Genochemy's virtual organism) exists is called Lab, but it was pointed out that the space was abstract and difficult to understand. Therefore, we will make it possible to display concrete experimental apparatuses such as petri dishes. It would also be good to allow the user to choose the experimental apparatuses, and the number of available apparatuses will increase as the user progresses through the tasks, just as in the case of blocks.

(6) To give it a sense of reality, we are also thinking of adding animation effects that show the transformation process and the DNA being expressed when starting an experiment.

These changes bring Genochemy closer to our goal of making synthetic biology accessible to everyone. More people will be able to learn synthetic biology in a way that is easy to understand, accessible, and fun. Because of the size of these changes, we were not able to finish the development during the period, but they will be implemented in the near future.