Team:EPFL/HP/Mini-Kit
Educational Cell-Free Mini Kit (ECFK)
Ever since the workshop in Cambridge, we had plenty of ideas on how to realize the Cell-Free Mini Kit intended for demonstrations at local high schools. Furthermore, doing an educational kit relying on the cell-free technic for protein expression systems would offer the great advantage of cheap, simple experiences and the possibility to easily store the kit. Hence,in August, the team reached out to several high schools located nearby EPFL to ensure they had interests in our kit and that they would add it to their classes. A few days later we received encouraging responses from two high schools in Geneva, College Calvin and Ecole Moser, who told us to provide them with a more detailed proposal as soon as we would have it ready.
The ECFK consists of several tubes containing different lysate reaction components, ideal to show a range of biomolecular processes. There are two kits: one of them is freeze dried, which enables simpler manipulations and storage for high schools with low budget; the other one isn't freeze dried, which enables students to learn how to pipette correctly and perform more elaborate experiments. The mini kit also contains a detailed description of the experiments that can be performed with it as well as suggestions of topics (such as cell-free biology) that the teacher could find interesting to include in his biology lectures. In fact, the ECFK is a diverse toolbox suitable for the demonstration of biological principles to groups with basic knowledge in biology, as the teacher is free to provide the students with an adequate background for their grade level.
After having set our mind on what the Cell-Free Mini Kit should look like in the end, we reached out to EPFL's biosafety department to make sure we were complying with national safety requirements. Stéphane Karlen, the main coordinator for biological safety on campus, made a risk evaluation of the kit and concluded that the Cell-Free Educational Mini Kit didn't represent a risk for the user nor the environment.
Details on the Cell-Free Mini Kit
The Cell-Free Mini Kit includes the material and experimental setup for the following four questions :
- Presence of the ß-galactosidase enzyme in a cell used for lysate production
- Expression of protein in a cell-free extract
- Interest of energy supply for biochemical reactions
- Polymerase operating mode
- Repression of a gene
The experiments will include the following procedures :
- Detection of ß-galactosidase enzyme presence in a lysate made of cell culture by colorimetric assay
- Study of cell-free in vitro expression of ß-galactosidase and kinetics of ß-galactosidase activity
- Showing the importance of an energy source by performing cell-free experiments with and without energy solution
- Testing different polymerases in cell-free environment with different promoters
- Study the principle of gene repression using a riboswitch which can be expressed in the presence of a trigger
Components for the abovementioned reactions include :
- E.Coli cell lysate of BL21, DH5alpha and M15 strains
- Energy solution, a mixture containing amino acids, salts, reducing agents
- Buffer A, contains Tris, magnesium and potassium glutamate
- Substrate, chlorophenol Red-β-D-galactopyranoside
- Linear DNA templates to be expressed in vitro (for example LacZ)
- Our toehold/trigger, which is a construct of single and double stranded DNA templates
The kit exists in two versions: either all the materials are freeze-dried and delivered in tubes containing the separate freeze-dried components, or they are distributed in their liquid form using dry ice.
Visit and feedback from high school teachers
In October, two high school teachers from the College Calvin in Geneva came to EPFL to meet with two iGEM EPFL members and discuss the possible integration of the Cell-Free Educational Mini Kit in their high school's biology curriculum. We gave them a live demonstration of the experiments provided by the Mini Kit and discussed the impact it would have on the high school courses of their students. The conversation was very interesting, as we were until that very moment not sure how well the high school labs were equipped - would they have micro-pipets, -80°C freezers, incubators? The teachers were glad to respond to all of our questions, were very enthusiastic and already gave us a first feedback. In fact, instead of working with freeze-dried components, they taught us that many high schools have the capacity to handle the components in their fluid form as their labs have micro-pipets and access to -80°C freezers; we thus adapted our protocols in order to render the mini kit suitable for such handling. We managed to include many other aspects of their feedback into our Minikit, although two of them persist until now: The central questions of each experiment might be a bit too easy for high school students and it would be great to have two versions of the instructions, one for the students and one for the teacher.
"Votre projet me paraît passionnant et je serais très heureux de parler avec vous des implications et applications dans les classes. Il pourrait faire l'objet d'une publication dans le cadre du projet Bio-Tremplins de la faculté des sciences et du DIP Genève qui s'adresse aux enseignants et touche près de 400 personnes à travers la francophonie."
"Your project seems fascinating to me and I would be happy to talk to you about implications and applications in class rooms. It could be part of a publication in line with the project Bio-Tremplins of the science faculty of the university of Geneva that is addressed to teachers and will touch up to 400 people all over the french speaking part of Switzerland."
"Je parle chaque année du projet iGEM à mes élèves de 4OS, alors si pour une fois il y a moyen de faire plus que de regarder une vidéo, je serai ravi de le faire."
"Every year I talk about the iGEM project to my last year high schools students, hence if, for once, it is possible to do more than watching a video, I would be happy to do it."
"Quel projet captivant et motivant sur lequel vous travaillez ! Il est toujours sympathique d’avoir des nouvelles d’anciens élèves et d’être informés de ce qui se fait dans les laboratoires. Nous sommes intéressés par votre kit."
"What a captivating and motivating project on which you are working ! It is always nice to hear from former students and learn more about what is being done in laboratories. We are interested in your kit."
A sustainable solution for high schools
The next issue we had to settle was the supply of the components needed for the Mini Kit for high schools in a sustainable way that would last long after the end of iGEM 2017 competition. The team will indeed not have access to the lab spaces anymore after November 2017 and will therefore not be able to manufacture and deliver components of the Cell-Free Mini Kit. We looked into more details for the following two solutions:
- Having one of the labs at EPFL produce the lysate and energy solution; for example, as suggested by our supervisors themselves, within the scope of the laboratory initiation course for second year students at the faculty of Life Sciences, which would have made our project worth of a whole set of in-lab practical sessions at EPFL
- Contacting the company responsible for delivering laboratory materials to the high schools
The second point caught our attention after the meeting with the teachers. They gave us the contacts of BiOutils - (french for bio-tools), whose mission is to promote the teaching of modern biology at local high schools and supply them with the required materials and competences. We have established a collaboration with BiOutils: we currently are in contact with this platform and are exchanging protocols for lysate production and energy solution in order to make the to production and supply of all the components of the Cell-Free Mini Kit for high schools on a larger scale achievable once the iGEM competition will be over. This solution fits our expectancies for the mini kit in the most satisfactory way as it will then enter a list of supported cantonal experiments for high schools, and biology teachers of the whole Canton of Geneva would be able to order and use the ECFK in their class.
We are very glad to have found such broad support and interest in the idea of our Cell-Free Educational Mini Kit and continue to work in the hope that it will become permanently integrated in local high school curricula. With the help of BiOutils, the Mini Kit will be produced on a larger scale and will be accessible for every high school in the region. Its future use will be ensured after the Jamboree and will, so do we hope, create a long lasting impact on education by generating infatuation for little known science fields amongst high school students in addition to bringing cell-free biology into swiss educational systems.
Feedback
Our interactions with the high school teachers were very valuable for us to gather opinions and reactions in regard to the Mini Kit we wanted to propose. After exchanging the protocols with the teachers from College Calvin, they suggested to us that lyophilisation of the components would not be needed, as they were equipped to directly manipulate the solutions. They told us that it would be better for the course since the students would be the ones pipetting the solutions and preparing the mixtures and that it would make the Mini Kit more interactive. We of course agreed with this statement and went on to modify our Cell-Free Educational Mini Kit protocols to include manipulation of the solutions directly and describe the amounts of reagents needed for delivery. Hence, after this discussion, we now propose two different kits: one lyophilised, enabling easy storage for low ressources high schools, and one non-lyophilised, for high schools that could have access to pipettes and adequate storage facilities for the kit.
After the visit, the two teachers told us they were very eager to try out a test-kit such as to integrate the kit before the current school high school semester ends in February. We promised to provide them with five test kits as soon as we would come back from the Giant Jamboree in Boston so that they could get started with experiments!
The company BiOutils wanted to know how the individual components could be synthesized or produced, as they would taking over this part for the high schools. We quickly provided them with our detailed protocols on how to make lysate, which cell lines are optimal for cell-free protein expression, how to prepare the energy solution and which substrates and buffers are needed for the reactions.
After an hour-long conversation and several e-mails exchanged with the Ecole Moserin Geneva, we had established that Ecole Moser would like to include the Cell-Free Mini Kit in their curriculum but could only do so once the next academic year begins (starting Summer 2018). They are also being supplied by BiOutils and asked us if it were possible to expand the Mini Kit with certain experiments for more advanced biology courses. This suggestion was very much welcomed by us and we set out to write a manual that may serve as a roadmap for generating new mini kit experiments. At the moment we are discussing with them the experiments of the mini kit they find most useful and the way to realise them.
