This year, we did a lot work in our lab and beyond the bench concerning our project – a brand-new IGEM BLOOD ALCOOL METER (iMeter). We questioned the reliability of the breathalyzer test in the field sobriety test. We researched the news and found out the factors influencing the breath test such as individual metabolic differences and diabetes diseases, as well as in some situations the police cannot force people to breathe such as suspects’ refusal and people dead or in coma.
In our human practices, we surveyed the alcohol consumers (i.e., the school teachers, the parents, the general public) and the experts (i.e., the police, the judges, the health care workers, the wine tester) by the questionnaire concerning the sobriety issues. Moreover, we visited them in the police station, in the court and in the hospital to discuss the issue of “drinking and driving” and the idea of our project. Furthermore, we hosted a drink “party” assisted by the school’s military instructor and a police officer to let the school’s principal and teachers to experience the breathalyzer used in the field sobriety test. And all of the opinions and encouragements got from the activities did help us design our iGEM product and do a better iGEM project.
In our lab work, we completed 11 new BioBricks including improving an existing part. We did protein analysis and enzyme activity assay to prove our concept. In addition, we performed modeling to test our enzyme’s efficiency in an electrochemical simulator, which was helped by biotech companies working on blood glucose meter innovation, BIONIME Corp. and Zensor R&D Corp. Overall, this is an amazing year for us, MINGDAO iGEM 2016. See you in the Giant Jamboree!
- We have registered MINGDAO as an iGEM TEAM 2016, and had a great summer in our biolab and beyond the bench. Furthermore, we’re going to have fun at the Giant Jamboree
- We have established MINGDAO’s Wiki that clearly documents our project and human practice activities.
- We have clearly declared the attributions of our members in each aspect of your project.
- We have documented the eleven novel standard BioBrick Parts and Devices central to our project. [BBa_K1991000, BBa_K1991001, BBa_K1991002, BBa_K1991003, BBa_K1991004, BBa_K1991005, BBa_K1991006, BBa_K1991007, BBa_K1991008, BBa_K1991009, BBa_K1991010]
- We have submitted our BioBrick parts to the iGEM Registry.
- We have completed the Judging form and the Safety form.
- We have prepared PPT slides and Poster to present at the Giant Jamboree.
- We have experimentally validated that four new BioBrick Parts and Devices of our own design and construction work as expected. We also documented the characterization of this part in the Main Page section of the Registry entry for that Part/Device. [BBa_K1991003, BBa_K1991007, BBa_K1991009, BBa_K1991010]
- In the Collaboration, we have helped a registered iGEM team of NCTU-FORMOSA 2016 in a significant way by validating the protein expression of one of their parts using SDS-PAGE and Coomassie Blue staining.
- In the Human Practices, our team has investigated on alcohol consumers (e.g., the school’s teachers, the member’s parents, the general public, etc.) and experts (e.g., the police officers, the judges, the health care workers, the wine tasters, etc.), as well as addressed problems with a survey in the context of our project concerning the field sobriety test. In addition, we educated the public about the issue of drunk driving by hosting a sobriety test experience for school’s staffs/teachers in the campus assisted by the school military instructor and the police officer. Moreover, we exchanged the idea and shared iGEM experience with other iGEM teams in a meet-up.
- We expanded the Human Practices activity by integrated the investigated issues into the design and the execution of our project. For example, we found the problems from the news against the breathalyzer test such as the individual metabolic differences, diabetes disease, suspects’ refusal, people dead or in coma. We’re inspired by Blood Glucose Meter from the data in the questionnaire concerning the sobriety test. We transformed the idea to design and execute our project and collaborated with a biotech company to create our IGEM Blood Alcohol Meter (iMeter).
- We improved of a previously existing BioBrick Part [BBa_K1694002] created by NCTU-FORMOSA in 2015 and entered the information in the part's page on the Registry.
“An alternative version with BamHI site: If you'd like to directly use this part as a vector/backbone for cloning a fusion protein, you should be noticed that an extra NcoI site is present on Cm resistance gene. BBa_K1991004 replaced NcoI site behind LO with BamHI site and provides an alternative choice for cloning a LO fusion protein.”
- We demonstrated a functional proof of concept of our project consisting of the BioBrick device [BBa_K1991009].
- To show our project working under real-world conditions. We demonstrated our whole system working under simulated conditions in the lab.
This year, we made the impossible possible. With all of our human practices activities and the achievements from the lab work, we believe MINGDAO is eligible for special prizes at the Jamborees.
“To better understand the problems concerning field (alcohol) sobriety tests, in addition to the lab works, our Human Practices are even more abundant and attractive. On the purpose of understanding law regulations, the breathalyzer devices and the practices, we met judges in the local court, got help from the police officers at the police station and visited nurses and technicians in the hospital. An online survey on the sobriety tests was designed and investigated from the public. And in our campus, we hosted a drink “party” to the teachers and the principal to let them understand and experience the sobriety test. Based on these integrated knowledge, we designed our prototype and collaborated with BIONIME Biotech Corp. to develop a novel blood alcohol meter. We also presented our work on an iGEM team meetup with NCTU_Formosa, NTHU_Taiwan, CGU_Taiwan. With these efforts, we hope we can win the prize.”
“To let our iGEM project go to the public and educate ourselves and junior students in Mingdao High School, we hosted a variety of activities. First, concerning the issue of alcohol sobriety test, we designed a questionnaire to survey the public's opinion and experiences including general public, school teachers, policemen, judges, health care workers and our parents. Second, we hosted a wine drinking “party”, and the school military instructors and a police officer help us to perform sobriety test using the breathalyzer to the teachers and our principal. On the education part, we visited the top research institutes in Academia Sinica, Taiwan. Moreover, our advisors and instructors taught biotech skills in the workshops. In addition, we share the iGEM spirits to the junior students with lectures and lab experiments. We do a lot on Education and Public Engagement, we hope we can get the prize.”
“We did make a lot of efforts on modeling with the help from BIONIME Biotech Corp, a blood glucose innovator. We performed experiments in electrochemical analysis, including cyclic voltammetry (CV) and amperometry. We conducted CV to study the electrochemical properties of our enzyme. The results showed that our work is functional and the redox reaction proceed smoothly. After that, amperometry was used to monitor the changes of current in a given period. The results of amperometry provided the data needed for drawing a calibration curve describing the concentration of alcohol and the current generated. The results we acquired gave us a deeper understanding of the connections between the alcohol concentration and the current produced. That helped us design our prototype and create the ideal environment for the chemical reaction to proceed efficiently and effectively. For those achievements, one cannot deny that we are fully qualified for the prize.”
“This year we are focusing on the problem of drunk driving. We find out that the breath sobriety test contains many problems. We’d like to apply the idea of Blood Glucose Meter (BGM) to a whole new design. In BGM, glucose reacts with glucose oxidase (GOX) on the electrode to produce electron. As a result, the device can measure the glucose concentration based on current intensity. To measure alcohol in our project, we engineered E. coli to produce alcohol oxidase (AOX) to catalyze the oxidation of alcohol just like GOX dose on glucose. Furthermore, we made a fusion protein with Lpp-Omp that make AOX displayed on the bacterial surface without need to purify the protein. After the electrochemical studies and prototype design, we created a noviel iGEM Blood Alcohol Meter (iMeter), which is portable, easy-to-use, accurate and time-saving compared to the sobriety test system used today.”
Best New Basic Part:
BBa_K1991001, Alcohol oxidase (AOX) gene from Pichia pastoris was optimized for protein expression in E. coli. The enzyme we proved can react with alcohol in a dose-dependent manner and be applied in biosensors.
Best New Composite Part:
BBa_K1991009, AOX was driven by a constitutive promoter and a strong RBS that could be displayed on E. coli cell surface by fusing with Lpp-OmpA outer membrane proteins. We confirmed the protein expression and enzyme activity, as well as proved the concept by demonstrating the chemical properties in an electrochemical simulator.
Best Part Collection
[BBa_K1991000, BBa_K1991001, BBa_K1991002, BBa_K1991003, BBa_K1991004, BBa_K1991005, BBa_K1991006, BBa_K1991007, BBa_K1991008, BBa_K1991009, BBa_K1991010] “Alcohol oxidase (AOX) was applied in biosensors which catalyzes the oxidation of alcohol followed by hydrogen peroxide production. There are two AOX genes present in a methylotrophic yeast, Pichia pastoris. The gene codon was optimized for expression in E. coli. The DNA sequences were synthesized by IDT and cloned onto pSB1C3 [BBa_K1991000, BBa_K1991001]. In addition, Lpp-OmpA outer membrane proteins, which can direct a fusion protein to display on the bacterial cell surface [BBa_K1991004, BBa_K1991007], were fused with AOX [BBa_K1991005, BBa_K1991006]. In our experimental results, the data showed the enzyme displayed on E. coli can react dose-dependently with various concentration of alcohol in the H2O2 production assay and modeling at the electrochemical analyzer [BBa_K1991002, BBa_K1991003, BBa_K1991008, BBa_K1991009, BBa_K1991010]”