After setting our goal to decompose synthetic dye with laccases, we sought to find out more information of biological sewage treatment and practical problems that required to be addressed. We also wanted to find out how our project could be applied to real world sewage treatment, and what change should we make to improve our project.

Therefore, we visited Hanghua sewage plant（World Trade Center, Beijing, China). Hanghua sewage plant was experienced in using biology method to decompose pollutant while they also had used other ways, including chemical and physical methods. Mr. Li, the manager of the sewage plant, told us lots of information we needed.

During the interview, we found out the situation of each current methods.

First of all, we learned that their previous chemical and physical techniques such as active carbon adsorption and oxidation method, and being expensive in cost for both methods, the sewage plant had to switched to biological treatment.

Hanghua sewage plant mainly used integrated sewage treatment device, in which a critical step was to add bacteria to decompose pollutant. Specially engineered bacteria were able to reproduce rapidly in sewage water, and this allowed the factory to process waste water with relatively lower financial requirement.

However, the bacteria used in sewage treatment may release toxic gases such as H2S. The plant had to turn on the ventilation system periodically, which consumed large amount of electric power and produced loud noise. As time went by, the advantage of lower cost is offsetted by electricity bills and the higher payment for their workers working under unfriendly environment.

In traditional biological treatment method, bacteria are usually injected into waste water directly, floating, growing, as well as continuously being removed by the outflow. The dynamic balance of bacteria concentration might be broken by increasing the total flow of sewage water. Under extreme circumstances, growth rate of bacteria might be overwhelmed by the loss rate through outflow. Moreover, workers in the sewage treatment plant told us, due to wear and tear since 2003 and other undiscovered problems, the total ability of sewage treatment had dropped 40%. To address this issue, we needed to develop a sustainable consumptive material.

By discovering shortcomings of traditional treatment method, we found some critical factors that were worth considering in our project.

The first factor was the low concentration of bacteria/enzyme during treatment. A high flow rate may wash away bacteria fast, lowering the concentration of bacteria/enzymes as well as the chance and duration of contact between pollutant and enzymes. This reminded us to adjust our project in two aspects:
(1) Attach bacteria/Laccase to some fixed matrix, preventing them from being washed away by water flow.
(2) Use lots of small plastic beads to expand surface area, increasing the chance of contact between pollutant and enzymes.

In combination of the two aspects, we may achieve a higher concentration of laccase and longer duration of contact between pollutant and enzymes. Moreover, because the laccase can process the sewage without releasing toxic gas, it will eliminate the unnecessary cost of using extraction pump and extra employee payment, which as a result can enable factory to utilize the biggest advantage of biology method with inexpensive price.

The second factor to consider was the competition among bacteria species. Through human practice, we realized that the composition of sewage disposal was very complicated. There were usually more than one species of microorganism competing in the same environment, which may negatively affect the growth of target bacteria. By allowing our bacteria to produce biofilm, they could get an advantage of growth by sharing nutrients and getting sheltered from competitors.

Finally, we came up with the idea, using PHA plastic beads coated with biofilm and biofilm producing bacteria, then display laccase enzyme on biofilm by adding SpyTag to Biofilm and SpyCatcher to CotA laccase. PHA can also act as a carbon source to bacteria, maintaining growth of bacteria and production of biofilm for longer period. This may further reduce cost by replacing biomaterials less frequently.

Our public education includes two sections: on campus and off campus activities.

On campus, we design and implement a layered education system, starting from series of lectures to all students, interviewing some students and evaluating their potential of being iGEMers, then our synthetic biology club offering them training in genetic engineering.

Off campus, we established the collaboration with sewage plant for exchanging the instructive information including biology, plant and policy. We strongly believe that the practicability of our project would attract more plants even business companies to cooperate with us.

We conducted lectures of iGEM projects and synthetic biology techniques to students on Tuesday and Thursday weekly. At the start of our education program, we briefly introduced our project's purpose, effectiveness and future potential contribution to the environment. We also invited students who were interested in improving the environment by synthetic biology technologies. After that, we held a series of lectures explaining the molecular mechanism, and prove practicability of our project to the students who followed up our project.

For the whole process of education, we put a high value on the effectiveness of lecture. Therefore, we required students who is willing to cooperate our work to register their WeChat account, and asked randomly 75% of students each time to share their perspectives, so we could adjust and improve the content of our lecture. We also interviewed students to find future iGEMers.

Furthermore, in our synthetic biology club, we introduced some basic methods and techniques of genetic engineering such as DNA extraction, restriction enzyme digestion, Primer design, and PCR, reserving talented students for our further iGEM projects.

In general, our on-campus education program comprehensively passed on our experience in iGEM and synthetic biology knowledge. Our program had also triggered students’ concern on environmental problems, as well as their passion for learning synthetic biology and becoming a qualified iGEMer.

During the visit of a sewage treatment plant, we briefly introduced our project to work leaders, then we showed reliability of our project and resolution to protect the environment. On one side, sewage treatment plant helped us conduct the design of our program by providing first-hand information of sewage plant. On the other side, we offered more information of our modified biology treatment. As a result, both sides were looking forward to establishing a long-term collaboration.

In conclusion, through the biology method, we found that the expensive cost and decreasing effectiveness are two main problems for traditional biological treatment method. These experiences from human practice has inspired us with the idea of designing our Biofilm × Laccase system. In the near future, we will build a filter with PHA beads coated with laccase linked biofilm and optimize the filter for real world sewage treatment.