Difference between revisions of "Team:CHINA CD UESTC/hp education"

The previous studies showed that the enzymatic biofuel cell (EBFC) has more advantages on operation and function over the ordinary biofuel cell (BFC). EBFC has three main advantages: 1) High efficiency of energy conversion; 2) Green alternative energy; 3) Characteristics fitted the biosensor. We found two high-efficiency oxidases, glucose oxidase and laccase, which can be used in anode region and cathode region of EBFC respectively. At the same time, we learned that laccase have some other advantages like ^[3] : 1) It comes from natural bacteria or plants, so it’s friendly to environment; 2) It has high-efficiency oxidability; 3) Laccase can oxidize a broad range of substrates so that it can be used in sewage disposal. It is definitely the most suitable choice to put laccase on the cathode.

Therefore, we set out to transform the cathode with laccase. For the purpose of visualizing the location and concentration of laccase, we combined RFP with laccase. After that, we designed a way of enriching laccase on the cathode--using magnetosomes！

Along with the development of times and population growth, energy consumption is increasing rapidly. Up to now, the thermal power is the main source. According to BP Energy Outlook 2035 , we can find that the world's fossil fuel reserves are declining ^[4] , and the largest shift of shares give us an insight into the most likely shape of the future energy landscape! (Figure 1)

Figure 1. There have been some rapid shifts in fuel shares in power generation in the past: oil gaining in the 1960s and losing in the 1970s; nuclear picking up in the 1970s/80s and falling in the 2000s; gas rising through the 1990s and 2000s. In the Outlook, the largest shifts are the increase in the renewables share and the decline in the coal share ^[4] .

Among numerous renewable energy, bioenergy is a kind of clean renewable energy and a potential excellent substitute for fossil fuel. With the advance of biotechnology biofuel cell (BFC) which can convert the chemical energy of fuel into electric energy with enzyme or microbial tissue as a catalyst, has been researched widely.

Previous iGEM teams had done some studies about microbial fuel cell (MFC). iGEM13_Bielefeld-Germany made an Escherichia coli Fuel Cell platform to provide an efficient electron transfer from the bacteria to the electrode. iGEM14_LZU-China cloned a NO3-sensor sequence and riboflavin producing genes into Escherichia coli for anode and a gene coding chromate (VI) reductase Yief was cloned into E.coli for cathode. iGEM14_SCAU-China boosted up the level of intracellular NAD+ for higher electron transfer rate.

Biofuel cell is divided into microbial fuel cell and enzymatic biofuel cell (EBFC). EBFC is a special kind of fuel cell which uses organics as fuels and enzymes as catalysts. EBFC is generally separated into anode region and cathode region by proton exchange membrane. Fuels are oxidized under the action of enzyme in the anode region. Oxygen is reduced in the cathode region.

EBFC have broad application prospect, so we want to create a new type of device to develop the bioenergy.

The main configurations of enzymatic fuel cells involve bioanodes based on glucose oxidase, glucose dehydrogenase or lactate oxidase and biocathodes based on copper oxidases such as laccase, tyrosinase or bilirubin oxidase. This concept was initiated by Mano et al. who implanted microbioelectrodes based on osmium redox hydrogels, in a grape obtaining thus 2.4mW at 0.54v ^[5] .

Laccases is a kind of copper-containing oxidoreductase. In the reduction reaction, the electron from the oxidation is transferred to the other three copper ions. These ions form a trinuclearic cluster, which transfers electrons to the terminal electron acceptor oxygen ^[6] .

Meanwhile, laccase has the property of oxidizing a wide range of substrates e.g., phenolic compounds, so it can be used in sewage disposal. Our project used these two enzymes and transformed the cathode. We constructed the expression vector of RFP + laccase and transformed it into E. Coli. The red fluorescence produced by RFP can be used as an indication of laccase’s concentration and activity. According to the method of electron transfer, EBFC can be divided into electronic media electrodes and direct electrochemical electrodes. Considered that the latter has high catalytic efficiency and small restriction by environment, we tried to enrich the laccase on the cathode to enhance the redox potential of our EFBC.

Figure 2. An electrochemical phenol biosensor based on the immobilization of laccase (Lac) on the surface of copper capped magnetic core–shell (Fe3O4–SiO2) nanoparticles (MNPs) ^[7] .

We obtained the laccase from BBa_K863005 . Traditional chemical approaches ^[7] of fixing laccase may affect the activity of laccase and are toxicological. So we hoped to find a better method!

Of course using synthetic biological methods is a great idea to achieve our goal. Magnetotactic bacteria (MTB), a kind of bacteria that can be attracted by magnet, are a superexcellent choice for us. We noticed that MTB contains a fantastic structure-- magnteosome. It is a magnetic nano materials covered by biofilm. And the magnetosome is essential to magnetotaxis.After our investigation, we decided to connect laccase to the magnetosomes’ membrane to enrich them on the cathode surface.

Figure 3. Transmission electron microscopy images of several different MTB showing their distinctive cell and magnetosome crystal compositions and morphologies. Scale bars = 500 nm in bacterial images and 100 nm in magnetosomes images ^[8] .

After our investigation, we decided to connect laccases to the magnetosome's membrane to gather them on the cathode surface.

But there are two problems for us to solve. On one hand, MTB are anaerobic, it means they are hard to be cultured. On the other hand, it is difficult to modify them. So, we were aiming to construct a magnetosome expression system in E.coli to solve those problems. According to a paper in Nature nanotechonlogy , we confirmed that transferring four related operons can make other bacteria magnetotactic ^[9] .

Finally, we co-transferred all the vectors we constructed to make E.coli produce magnetosomes carrying laccase. The special magnetosomes would be used into our EBFC to improve the electron transfer efficiency!

Purpose:

As we were alarmed to learn that most of Chinese believe that synthetic biology is harmful to human health. To combat this problem, we developed a dual-focus, comprehensive educational program targeting students. We thought that educate students is the most effective way to correct people’s conceptions. The team planned and carried out a series of iGEM educational lectures for aged 5-13 years old as part of the University of Electronic Science and Technology’s Education programs event.

1: we went to kindergarten to teach the children some biological knowledge by way of science course.
2: we held an interesting biology lecture on the experimental primary school of UESTC.
3: we extended iGEM to the village schools in order to spread iGEM to remote areas which is low-education. We wanted to teach them more things they do not understand. Viewed from another aspect, we would like to propagation iGEM to wherever our capacity allows.

On May 8th 2015

Members of the Hong Kong University Medicine School visited UESTC Life Science and Technology School. We iGEM experiment team took the opportunity to talk about our project with Professor Zhaoyi Hu. Knowing the main processes of our project, Prof.Hu said, "It seems that the competition not only requires you players have a certain professional basis and innovation capability, but also is a test of students' learning and thinking comprehensive ability." Then he put forward his concerns especially in biosafety aspects, namely, whether the bacteria source itself is safe or pathogenic, and if gene modified organisms are safe.

In the aspect of biosafety, our project have achieved three aspects as following:
1. The safety of bacteria source: DH5α and BL21 (DE3) we use are common bacterial strains;
2. The safety of transgenosis: The genes we will transfer are nonpathogenic and nonpolluting. There are little adverse impacts on environment and rare probability of transgene escape.
3. The safety of experimental operation: We players have formed a strict operation habit, such as sterilization of transgenosis bacterias, protection of experimenters.

In the end, Prof.Hu told us the general processes of sample treatment in medical research as well as separation and preservation methods of some bacteria strains.

Professor Zhaoyi Hu is head of Microbiology Department of Hong Kong University Medicine School, Infection and Immunity Joint Research Center of Hong Kong University. Their results research have a broad impact on international academic circles, especially the medical community .